APITOXINA & SISTEMA INMUNOLOGICO
| 1: Ann N Y Acad Sci. 2005 Nov;1056:279-92. |
Novel Drugs
and Vaccines Based on the Structure and Function of HIV Pathogenic Proteins
Including Nef.
Azad
AA.
Faculty of Health Sciences, Medical School, University of Cape Town, Anzio
Road, Observatory, 7925, Cape Town, South Africa. a_azad05@yahoo.com.au.
Evidence is presented to suggest that HIV-1 accessory protein Nef could be
involved in AIDS pathogenesis. When present in extracellular medium, Nef causes
the death of a wide variety of cells in vitro and may therefore be responsible
for the depletion of bystander cells in lymphoid tissues during HIV infection.
When present inside the cell, Nef could prevent the death of infected cells
and thereby contribute to increased viral load. Intracellular Nef does this
by preventing apoptosis of infected cells by either inhibiting proteins involved
in apoptosis or preventing the infected cells from being recognized by CTLs.
Neutralization of extracellular Nef could prevent the death of uninfected
immune cells and thereby the destruction of the immune system. Neutralization
of intracellular Nef could hasten the death of infected cells and help reduce
the viral load. Nef is therefore a very important molecular target for developing
therapeutics that slow progression to AIDS. The N-terminal region of Nef and
the naturally occurring bee venom mellitin have very similar primary and tertiary
structures, and they both act by destroying membranes. Chemical analogs of
a mellitin inhibitor prevent Nef-mediated cell death and inhibit the interaction
of Nef with cellular proteins involved in apoptosis. Naturally occurring bee
propolis also contains substances that prevent Nef-mediated cell lysis and
increases proliferation of CD4 cells in HIV-infected cultures. These chemical
compounds and natural products are water soluble and nontoxic and are therefore
potentially very useful candidate drugs.
PMID: 16387695 [PubMed - in process]
| 2: Eur J Immunol. 2005 Nov;35(11):3268-76. |
Prevention
of allergy by a recombinant multi-allergen vaccine with reduced IgE binding
and preserved T cell epitopes.
Karamloo F, Schmid-Grendelmeier P,
Kussebi F, Akdis M, Salagianni M,
von Beust BR,
Reimers A, Zumkehr J, Soldatova L,
Housley-Markovic Z,
Muller U, Kundig T, Kemeny DM, Spangfort MD,
Blaser K, Akdis CA.
Swiss
Novel approaches for the prevention of allergy are required, because of the
inevitably increasing prevalence of allergic diseases during the last 30 years.
Here, a recombinant chimeric protein, which comprises the whole amino acid
sequences of three bee venom major allergens has been engineered and used
in prevention of bee venom sensitization in mice. Phospholipase A2 (Api m
1), hyaluronidase (Api m 2) and melittin (Api m 3) fragments with overlapping
amino acids were assembled in a different order in the Api m (1/2/3) chimeric
protein, which preserved entire T cell epitopes, whereas B cell epitopes of
all three allergens were abrogated. Accordingly, IgE cross-linking leading
to mast cell and basophil mediator release was profoundly reduced in humans.
Supporting these findings, the Api m (1/2/3) induced 100 to 1000 times less
type-1 skin test reactivity in allergic patients. Treatment of mice with Api
m (1/2/3) led to a significant reduction of specific IgE development towards
native allergen, representing a protective vaccine effect in vivo. These results
demonstrate a novel prototype of a preventive allergy vaccine, which preserves
the entire T cell epitope repertoire, but bypasses induction of IgE against
native allergen, and side effects related to mast cell/basophil IgE FcepsilonRI
cross-linking in sensitized individuals.
| 3: In Vivo. 2005 Jul-Aug;19(4):801-5. |
Inhibitory effect
of whole bee venom in adjuvant-induced arthritis.
Lee
JY, Kang
SS, Kim
JH, Bae
CS, Choi
SH.
College of Veterinary Medicine and Research Institute of Veterinary Medicine,
Chungbuk National University, Republic of Korea.
The aim of this study was to assess the inhibitory effect of whole bee venom
(BV) on adjuvant-induced arthritis in the rat. Rats were divided into pre-apitherapy,
post-apitherapy and control experimental groups. The pre-apitherapy group
was subcutaneously stung with a honeybee (Apis mellifera L.) and the control
group was subcutaneously injected with 0.1 ml of physiological saline solution
one day prior to complete Freund's adjuvant (CFA) injection. The post-apitherapy
group was subcutaneously stung with a honeybee on day 14 after CFA injection.
When arthritis had developed in the rat, the post-apitherapy group was subcutaneously
administered whole BV every other day for a further 14 days. Clinical signs,
hematological values and radioglogical features were observed during the entire
experimental period. In the pre-apitherapy group, the development of inflammatory
edema and polyarthritis was inhibited. Significant differences in lameness
score, hind paw edema volume and radiological features were observed between
control and pre-apitherapy rats. White blood cell counts indicated that the
degree of leucocytosis was significantly different between the pre-apitherapy
and control groups (p < 0.01). Inflammatory edema, polyarthritis and bone
change into the right hind paw were effectively inhibited in pre-apitherapy
rats during the two-week period post-CFA injection. In conclusion, whole BV
was found to inhibit arthritic inflammation and bone changes in the rat. This
may be an alternative treatment for arthritis in humans.
PMID: 15999553 [PubMed - indexed for MEDLINE]
| 4: Brain Res. 2005 Jul 12;1049(2):210-6. |
The anti-inflammatory
effect of peripheral bee venom stimulation is mediated by central muscarinic
type 2 receptors and activation of sympathetic preganglionic neurons.
Yoon SY, Kim HW, Roh DH, Kwon YB, Jeong TO, Han HJ, Lee HJ, Choi SM, Ryu YH, Beitz AJ, Lee JH.
Department of Veterinary Physiology,
College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul
National University, Seoul 151-742, South Korea.
The anti-inflammatory effect (AI) induced by peripheral injection of diluted
bee venom (dBV) involves activation of spinal cord circuits and is mediated
by catecholamine release from adrenal medulla, but the precise neuronal mechanisms
involved are not fully understood. In a recent study, we demonstrated that
an increase in spinal acetylcholine is involved in mediating the anti-inflammatory
effect of dBV and that this mediation also involves adrenomedullary activation.
The present study utilized the mouse air pouch inflammation model to evaluate
the involvement of spinal acetylcholine receptors and sympathetic preganglionic
neurons (SPNs) in dBV's anti-inflammatory effect (dBVAI). Intrathecal (IT)
pretreatment with atropine (muscarinic cholinergic antagonist) but not hexamethonium
(nicotinic cholinergic antagonist) significantly suppressed dBVAI on zymosan-evoked
leukocyte migration. Subsequent experiments showed that IT pretreatment with
methoctramine (a muscarinic receptor type 2; M(2) antagonist), but not pirenzepine
(an M(1) antagonist) or 4-DAMP (an M(3) antagonist), suppressed the dBVAI.
In addition, dBV stimulation specifically increased Fos expression in SPNs
of the T7-T11, but not the T1-T6 or T12-L2 spinal cord segments, in animals
with zymosan-induced inflammation. Moreover, IT methoctramine pretreatment
suppressed this dBV-induced Fos expression specifically in SPNs of T7-T11
level. Peripheral sympathetic denervation using 6-hydroxydopamine (6-OHDA)
treatment (which spares sympathetic adrenal medullary innervation) did not
alter dBVAI. Collectively these results indicate that dBV stimulation leads
to spinal cord acetylcholine release that in turn acts on spinal M(2) receptors,
which via a hypothesized disinhibition mechanism activates SPNs that project
to the adrenal medulla. This activation ultimately leads to the release of
adrenal catecholamines that contribute to dBVAI.
PMID: 15953592 [PubMed - indexed for MEDLINE]
| 5: Int Immunopharmacol. 2005 Aug;5(9):1406-14. Epub 2005 Apr 20. |
Bee venom
modulates murine Th1/Th2 lineage development.
Nam
S, Ko E, Park
SK, Ko S, Jun
CY, Shin
MK, Hong
MC, Bae
H.
College of Oriental Medicine, Kyung Hee University, #1 Hoeki-dong Dongdaemoon-gu,
Seoul, 130-701, Korea.
Administration of bee venom (BV) elicits anti-inflammatory, anti-nociceptive
and anti-allergic effects in various animal models. This study was designed
to evaluate the direct effects of BV on helper T cell activities and on Th1/Th2
lineage development using both in vitro and in vivo conditions. In the Th1
skewed condition, BV increased the expression of IFN-gamma mRNA and enhanced
the expression of T-bet on purified CD4(+) T cells from splenocytes of BALB/c
mice. On the other hand, BV treatment did not alter the expression of IL-4
or GATA-
| 6: J Allergy Clin Immunol. 2005 May;115(5):1063-7. |
Increased
expression of osteopontin is associated with long-term bee venom immunotherapy.
Konno
S, Golden
DB, Schroeder
J, Hamilton
RG, Lichtenstein
LM, Huang
SK.
Johns
BACKGROUND: Venom allergen immunotherapy (VIT) is proven to be highly effective
for insect allergy, but the mechanisms and the biomarkers associated with
clinical efficacy remain elusive. OBJECTIVE: The aim of this study was to
identify candidate biomarkers associated with successful VIT. METHODS: Gene
chip array and clustering analyses of PBMCs from subjects with or without
VIT were performed. RESULTS: From gene chip array and clustering analyses,
an increased expression of osteopontin was found in patients who completed
5 to 6 years of VIT and discontinued therapy for 3 to 6 years (completed treatment
group) compared with the untreated group. A significantly higher level of
serum osteopontin was found in the completed treatment group compared with
the untreated group (n =
| 7: J Ethnopharmacol. 2005 May 13;99(1):157-60. |
Effects of
bee venom on the pro-inflammatory responses in RAW264.7 macrophage cell line.
Jang
HS, Kim
SK, Han
JB, Ahn
HJ, Bae
H, Min
BI.
Department of East-West Medicine, Graduate School, Kyung-Hee University, Seoul
130-701, South Korea.
The purpose of this study is to elucidate the molecular mechanism of anti-inflammatory
effect of bee venom (BV), which has been used for the treatment of various
inflammatory diseases in oriental medicine. With this aim, we examined the
effects of BV on the nitric oxide (NO) production by lipopolysaccharide (LPS)
or sodium nitroprusside in RAW264.7 macrophages. We further investigated the
effects of BV on the expression of inducible nitric oxide synthase (iNOS),
cyclooxygenase-2 (COX-2), nuclear factor-kappaB (NF-kappaB) and mitogen-activated
protein kinase (MAPK) with RT-PCR in LPS-stimulated RAW264.7 cells. BV suppressed
the NO production and decreased the levels of iNOS, COX-2, NF-kappaB and MAPK
mRNA in a dose-dependent manner. These results suggest that BV has an anti-inflammatory
effect by inhibiting iNOS and COX-2 expression, possibly through suppression
of NF-kappaB and MAPK expression.
PMID: 15848037 [PubMed - indexed for MEDLINE]
| 8: Clin Exp Allergy. 2005 Mar;35(3):367-73. |
Characterization
of the human T cell response to antigen 5 from Vespula vulgaris (Ves v 5).
Bohle B, Zwolfer B, Fischer GF, Seppala U, Kinaciyan T,
Bolwig C, Spangfort MD,
Ebner C.
Department of Pathophysiology,
Medical University of Vienna, VA-1090 Vienna, Austria. barbara.bohle@meduniwien.ac.at
BACKGROUND: The T cell reactivity to the major allergen of bee venom, phospholipase
A2, has been thoroughly characterized. In contrast, only little is known about
the human cellular response to major allergens from wasp venom. OBJECTIVE:
To characterize the human T cell response to antigen 5 from Vespula vulgaris,
Ves v 5. METHODS: Recombinant Ves v 5 was used to establish allergen-specific
T cell lines (TCL) and T cell clones (TCC) from the peripheral blood of vespid-allergic
and non-allergic individuals. Ves v 5-specific TCL were mapped for T cell
epitopes using overlapping synthetic peptides representing the complete amino
acid sequence of Ves v 5. Ves v 5-specific TCC were analysed for antigen-induced
secretion of IL-4, IFN-gamma and IL-10. RESULTS: Seventeen distinct T cell
epitopes were recognized by allergic individuals among which Ves v 5(181-192)
was identified as a dominant T cell epitope. Partially different epitopes
were observed in TCL from non-allergic subjects and the dominant epitope Ves
v 5(181-192) was not prevalent in these cultures. Ves v 5-specific TCC isolated
from allergic individuals did not show the typical T helper type 2 (Th2)-like
cytokine profile in response to specific stimulation, i.e. high amounts of
IL-4 and low IFN-gamma. TCC from non-allergic individuals showed a Th1-like
cytokine pattern. CONCLUSIONS: Our findings provide evidence that the allergic
T cell response to Ves v 5 is not Th2-dominated and that different immunogenic
sites on this major wasp venom allergen are recognized by allergic and non-allergic
individuals.
PMID: 15784117 [PubMed - indexed for MEDLINE]
| 9: FEBS Lett. 2005 Mar 14;579(7):1658-64. |
Cross-presentation
of a CMV pp65 epitope by human dendritic cells using bee venom PLA2 as a membrane-binding
vector.
Babon A, Almunia C, Boccaccio C,
Beaumelle B,
Gelb MH, Menez A, Maillere B, Abastado JP,
Salcedo M, Gillet D.
Protein Engineering and Research
Department (DIEP), bat 152, CEA-Saclay, 91191 Gif sur Yvette cedex, France.
We have used bee venom phospholipase A2 as a vector to load human dendritic
cells ex vivo with a major histocompatibility complex (MHC) class I-restricted
epitope fused to its C-terminus. The fusion protein bound to human monocyte-derived
dendritic cells and was internalized into early endosomes. In vitro immunization
experiments showed that these dendritic cells were able to generate specific
CD8 T cell lines against the epitope carried by the fusion protein. Cross-presentation
did not require proteasome, transporter associated with antigen processing,
or endosome proteases, but required newly synthesized MHC molecules. Comparison
of the antigen presentation pathway observed in this study to that followed
by other toxins used as vectors is discussed.
PMID: 15757657 [PubMed - indexed for MEDLINE]
| 10: J Allergy Clin Immunol. 2005 Feb;115(2):323-9. |
A major allergen
gene-fusion protein for potential usage in allergen-specific immunotherapy.
Kussebi F, Karamloo F, Rhyner C, Schmid-Grendelmeier P,
Salagianni M,
Mannhart C, Akdis M, Soldatova L,
Markovic-Housley Z,
Von Beust BR,
Kundig T, Kemeny DM, Blaser K, Crameri R, Akdis CA.
Swiss Institute of Allergy and
Asthma Research, Davos, Switzerland. fatimah.kussebi@charite.de
BACKGROUND: Specific immunotherapy is a common treatment of allergic diseases
and could potentially be applied to other immunologic disorders. Despite its
use in clinical practice, more defined and safer allergy vaccine preparations
are required. Differences between epitopes of IgE that recognize the 3-dimensional
structure of allergens and T cells that recognize linear amino acid sequences
provide a suitable tool for novel vaccine development for specific immunotherapy.
OBJECTIVE: The aim of the study was to delete B-cell epitopes and prevent
IgE crosslinking, but to preserve T-cell epitopes by fusion of 2 major allergens
of bee venom because of a change in the conformation. METHODS: By genetic
engineering, we produced a fusion protein composed of the 2 major bee venom
allergens: phospholipase A 2 (Api m 1) and hyaluronidase (Api m 2). RESULTS:
The Api m [1/2] fusion protein induced T-cell proliferation and both T H 1-type
and T H 2-type cytokine responses. In contrast, IgE reactivity was abolished,
and profoundly reduced basophil degranulation and type 1 skin test reactivity
was observed. Pretreatment of mice with Api m [1/2] fusion protein significantly
suppressed the development of specific IgE as well as other antibody isotypes
after immunization with the native allergen. CONCLUSION: The novel fusion
protein of 2 major allergens bypasses IgE binding and mast cell/basophil IgE
FcepsilonRI crosslinking and protects from IgE development.
PMID: 15696088 [PubMed - indexed for MEDLINE]
| 11: Arthritis Rheum. 2004 Nov;50(11):3504-15. |
Antiarthritic
effect of bee venom: inhibition of inflammation mediator generation by suppression
of NF-kappaB through interaction with the p50 subunit.
Park
HJ, Lee
SH, Son
DJ, Oh KW, Kim
KH, Song
HS, Kim
GJ, Oh GT, Yoon
do Y, Hong
JT.
College of Pharmacy, Chungbuk National University, 48 Gaesin-dong, Heungduk-gu,
Cheongju, Chungbuk 361-763, South Korea.
OBJECTIVE: To investigate the molecular mechanisms of the antiarthritic effects
of bee venom (BV) and melittin (a major component of BV) in a murine macrophage
cell line (Raw 264.7) and in synoviocytes obtained from patients with rheumatoid
arthritis. METHODS: We evaluated the antiarthritic effects of BV in a rat
model of carrageenan-induced acute edema in the paw and in a rat model of
chronic adjuvant-induced arthritis. The inhibitory effects of BV and melittin
on inflammatory gene expression were measured by Western blotting, and the
generation of prostaglandin E(2) (PGE(2)) and nitric oxide (NO) and the intracellular
calcium level were assayed. NF-kappaB DNA binding and transcriptional activity
were determined by gel mobility shift assay or by luciferase assay. Direct
binding of BV and melittin to the p50 subunit of NF-kappaB was determined
with a surface plasmon resonance analyzer. RESULTS: BV (0.8 and 1.6 mug/kg)
reduced the effects of carrageenan- and adjuvant-induced arthritis. This reducing
effect was consistent with the inhibitory effects of BV (0.5, 1, and 5 mug/ml)
and melittin (5 and 10 mug/ml) on lipopolysaccharide (LPS; 1 mug/ml)-induced
expression of cyclooxygenase 2, cytosolic phospholipase A(2), inducible NO
synthase, generation of PGE(2) and NO, and the intracellular calcium level.
BV and melittin prevented LPS-induced transcriptional and DNA binding activity
of NF-kappaB via the inhibition of IkappaB release and p50 translocation.
BV (affinity [K(d)] = 4.6 x 10(-6)M) and melittin (K(d) = 1.2 x 10(-8)M) bound
directly to p50. CONCLUSION: Target inactivation of NF-kappaB by directly
binding to the p50 subunit is an important mechanism of the antiarthritic
effects of BV.
PMID: 15529353 [PubMed - indexed for MEDLINE]
| 12: J Allergy Clin Immunol. 2004 Oct;114(4):943-50. |
Modulation
of allergic responses in mice by using biodegradable poly(lactide-co-glycolide)
microspheres.
Jilek
S, Walter
E, Merkle
HP, Corthesy
B.
Department of Chemistry and Applied BioSciences, Swiss Federal Institute of
Technology Zurich, Zurich, Switzerland.
BACKGROUND: Biodegradable poly(lactide- co -glycolide) (PLGA) microspheres
are a promising carrier for vaccine delivery capable of maturing antigen-presenting
cells to stimulate T-cell-mediated immune responses. However, the potential
of microspheres to downregulate an allergic response in vivo is unknown. OBJECTIVE:
The aim of this study was to determine whether microspheres could potentiate
DNA vaccination against allergy and to evaluate the immunomodulatory properties
of microspheres alone. METHODS: Mice were treated prophylactically with DNA-loaded
plain PLGA microspheres before sensitization with phospholipase A2 (PLA2),
the major allergen of bee venom. PLA2-specific IgG1, IgG2a, IgE in serum were
measured for 8.5 months, and splenocyte proliferative responses and cytokine
profiles were determined. Protection against anaphylaxis was evaluated after
injection of an otherwise lethal dose of PLA2. RESULTS: Phospholipase A2-specific
IgG1 and IgG2a production turned out to be 2 times higher using cationic microspheres
compared with anionic microspheres, but was not influenced by the presence
of DNA. In contrast, reduction in IgE production and T-cell hyporesponsiveness
were observed with all microsphere formulations. Recall challenge with PLA2
triggered combined expression of both IL-4 and IFN-gamma, together with sustained
expression of IL-10 that can explain the protective effect against anaphylaxis.
CONCLUSION: Our data suggest a dual mechanism that does initially rely on
a TH2 to TH1 immune deviation and then on IL-10-mediated suppression. This
is the first physiological demonstration that plain PLGA microspheres can
induce tolerance in mice for as long as 6 months postsensitization.
PMID: 15480340 [PubMed - indexed for MEDLINE]
| 13: Protein Sci. 2004 Nov;13(11):2970-8. Epub 2004 Sep 30. |
Alteration
of the tertiary structure of the major bee venom allergen Api m 1 by multiple
mutations is concomitant with low IgE reactivity.
Buhot
C, Chenal
A, Sanson
A, Pouvelle-Moratille
S, Gelb
MH, Menez
A, Gillet
D, Maillere
B.
Protein Engineering and Research Department, batiment 152, CEA-Saclay, 91191
Gif sur Yvette, France.
We have engineered a recombinant form of the major bee venom allergen (Api
m 1) with the final goal of reducing its IgE reactivity. This molecule (Api
mut) contains 24 mutations and one deletion of 10 amino acids. The successive
introduction of these sequence modifications led to a progressive loss of
specific IgE and IgG reactivity and did not reveal any immunodominant epitopes.
However, Api mut exhibited a clear loss of reactivity for Api m 1-specific
IgE and IgG. Injection of Api mut into mice induced specific antibody production.
This humoral response was as high as that induced by the Api m 1 but the cross-reactivity
of the antibodies was weak. As inferred by far UV circular dichroism, this
mutant was correctly folded. However, near UV circular dichroism and denaturation
curves of Api mut showed that it exhibits a dynamic tertiary structure and
that it is a highly flexible molecule. Finally, as all the sequence modifications
have been introduced outside the human and murine T cell epitope regions,
we investigated its T cell properties in mice. We showed that Api mut-specific
T lymphocytes induced in vivo were stimulated in vitro by both proteins. These
data provide new insights in the design of hypoallergenic molecules.
PMID: 15459335 [PubMed - indexed for MEDLINE]
| 14: Allergy. 2004 Oct;59(10):1110-7. |
The CD63
basophil activation test in Hymenoptera venom allergy: a prospective study.
Sturm
GJ, Bohm
E, Trummer
M, Weiglhofer
I, Heinemann
A, Aberer
W.
Department of Experimental and Clinical Pharmacology, University of Graz,
Graz, Austria.
BACKGROUND: The basophil activation test (BAT), which relies on flow cytometric
quantitation of the allergen-induced up-regulation of the granule-associated
marker CD63 in peripheral blood basophils, has been suggested to be a useful
approach in detecting responsiveness to allergens. The purpose of this study
was to establish the usefulness of the BAT with regard to the clinical history
and current diagnostic tools in Hymenoptera venom allergy using a prospective
study design. METHODS: Fifty-seven consecutive patients allergic to Hymenoptera
venom as defined by a systemic reaction after an insect sting, and 30 age-
and sex-matched control subjects with a negative history were included. The
degree and nature of sensitization was confirmed by skin testing, specific
immunoglobulin E (IgE), serum tryptase levels and BAT. In the nonallergic
control group only analysis of specific IgE and BAT were performed. Correlation
of BAT, skin test and specific IgE, respectively, with the clinical history
in the allergic group was termed as sensitivity and in the control group as
specificity. RESULTS: Twenty one of 23 (91.3%) bee venom allergic patients
and 29 of 34 (85.3%) patients allergic to wasp and hornet venom tested positive
in BAT. The overall sensitivity of BAT, specific IgE and skin tests were 87.7,
91.2 and 93.0%, respectively. The overall specificities were 86.7% for BAT
and 66.7% for specific IgE. No correlation between the severity of clinical
symptoms and the magnitude of basophil activation was observed. CONCLUSION:
The BAT seems to be an appropriate method to identify patients allergic to
bee or wasp venom with a comparable sensitivity to standard diagnostic regimens.
The higher specificity of BAT as compared with specific IgE makes this test
a useful tool in the diagnosis of Hymenoptera venom allergy.
Publication Types:
PMID: 15355471 [PubMed - indexed for MEDLINE]
| 15: Allergy. 2004 Oct;59(10):1102-9. |
The basophil
activation test in wasp venom allergy: sensitivity, specificity and monitoring
specific immunotherapy.
Erdmann
SM, Sachs
B, Kwiecien
R, Moll-Slodowy
S, Sauer
I, Merk
HF.
Department of Dermatology and Allergology, University Hospital of Aachen,
Aachen, Germany.
BACKGROUND: As in vitro diagnosis of wasp venom sensitization by specific
serum IgE has a sensitivity of only 60-80%, additional in vitro tests are
desirable. Basophil activation is associated with the expression of CD63 and
its measurement has been proposed as a novel in vitro test for immediate-type
allergy. Furthermore, to date, no in vitro test exists to monitor successful
specific immunotherapy (SIT) with wasp venom. Therefore, the potentially harmful
sting challenge is still recommended. OBJECTIVE: We compared the CD63-based
basophil activation test (BAT) in the diagnosis of wasp venom allergy with
skin tests and measurement of specific IgE. Furthermore, we investigated whether
BAT can predict the outcome of the sting challenge in patients on SIT. METHODS:
Fifty patients with a systemic reaction caused by a wasp sting and 20 controls
were studied. Intracutaneous tests were performed with wasp and bee venom
in the suspected allergics. Specific IgE was determined by the CAP-FEIA method
and basophil activation by flow cytometry upon double staining with anti-IgE/anti-CD63
mAb. Twenty-five patients were sting challenged 6 months after starting SIT
and the BAT was repeated before challenge. RESULTS: Sensitivity of the intracutaneous
tests, specific IgE and BAT was 100, 76, and 92%, respectively. Specificity
of specific IgE and the BAT was 85 and 80%, respectively. The cut-off for
a positive BAT was 15% CD63+ basophils. There was a positive correlation between
IgE reactivity to wasp venom and the number of CD63+ basophils (r = 0.65).
Although no patient had a systemic reaction upon sting challenge, in most
subjects basophil activation did not decrease when compared with the BAT before
SIT. CONCLUSIONS: Quantitation of basophil activation by CD63 expression is
a valuable new in vitro method for diagnosis of allergy to hymenopteran venoms.
The CD63-based BAT is a helpful tool for the complementation of routine diagnostic
tests such as specific IgE as it increases sensitivity of in vitro detection
of sensitization. However, this in vitro method does not offer an alternative
to the sting challenge in monitoring successful SIT.
PMID: 15355470 [PubMed - indexed for MEDLINE]
| 16: Am J Chin Med. 2004;32(3):361-7. |
Anti-inflammatory
effect of bee venom on type II collagen-induced arthritis.
Lee JD, Kim SY, Kim TW, Lee SH, Yang HI, Lee DI, Lee YH.
Research Group of Pain and Neuroscience
in Vision 2000 Project East-West Medical Research Institute, Kyung Hee University,
Seoul, Korea. ljdacu@khmc.or.kr
Bee venom (BV) has been used to relieve pain and reduce inflammation in traditional
Oriental medicine, especially in chronic inflammatory diseases such as rheumatoid
arthritis (RA). We previously reported that the BV injection into a traditional
acupuncture point (Zusanli) reduced arthritis-associated edema and nociceptive
responses in Freund's adjuvant-induced arthritis in rats (Kwon et al., 2001).
This study was designed to evaluate the anti-inflammatory and anti-cytokine
effect of BV on a murine type-II collagen-induced arthritis (CIA) model. Male
mice were immunized by spontaneous injection of 100 microg of an emulsion
of bovine type-II collagen and complete Freund's adjuvant (CFA), with a booster
injection after 2 weeks. In the experimental group, 0.1 ml BV was injected
at acupuncture point (Zusanli) near both knees twice a week for a total of
5 times. In the control group, normal saline was injected at the same frequencies.
These injections began 5 weeks after the first collagen injection. Starting
the 3rd week after the first collagen injection, we examined limb swelling
and severity of arthritis twice a week. At 8 weeks, mice were sacrificed and
synovial tissue was examined with the light microscope and serum cytokines
(IL-1beta and TNF-alpha) were measured by ELISA. The incidence of arthritis,
the mean arthritis index and the number of arthritic limbs were significantly
lower in the treatment compared to the control group (63% versus 75%, 3.4%
versus 8.5%, 23% versus 75%, respectively). Among the serum proinflammatory
cytokines, the production of TNF-alpha in the BV group was suppressed compared
to the control group (59 +/- 4.5 versus 99.5 +/- 6.5, p < 0.05), but IL-1beta
was not suppressed. The examination of the histopathology of the joints of
murine CIA showed decreased inflammation signs and less lymphocyte infiltration
after BV acupuncture therapy. Acupuncture therapy with BV suppressed the development
of arthritis and caused inhibition of the immune responses in type-II collagen-induced
arthritis.
PMID: 15344419 [PubMed - indexed for MEDLINE]
| 17: Clin Exp Allergy. 2003 Sep;33(9):1209-15. |
Impaired
secretion of interleukin-4 and interleukin-13 by allergen-specific T cells
correlates with defective nuclear expression of NF-AT2 and jun B: relevance
to immunotherapy.
Faith
A, Richards
DF, Verhoef
A, Lamb
JR, Lee
TH, Hawrylowicz
CM.
Department of Respiratory Medicine and Allergy, The Guy's, King's College
and St Thomas' Hospitals School of Medicine, London, UK. alex.faith@kcl.ac.uk
BACKGROUND: Allergen immunotherapy (IT) is a successful treatment associated
with decreased Th2 cytokine production by allergen-specific T cells. We have
previously demonstrated (Faith et al., J Immunol 1997; 159:53-57) that inhibition
of Th2 cytokine production in vitro correlates with impaired tyrosine kinase
activity through the TCR. The transcription factor complex, nuclear factor
of activated T cells (NF-AT), which regulates Th2 cytokine production is controlled
by the activity of tyrosine kinases. OBJECTIVE: To address whether decreased
Th2 cytokine production by allergen-specific CD4+ T cells following IT is
correlated with altered translocation and nuclear expression of the NF-AT
family member, NF-AT2, and the activator protein 1 (AP1) component of NF-AT,
jun B. METHODS: T cell lines specific for insect venom phospholipase A2 (PLA)
were derived from patients prior to and during conventional venom IT. Nuclear
expressions of NF-AT and jun B were assessed following stimulation through
the TCR. Th1 and Th2 cytokine and IL-10 production by insect venom-specific
T cells were also determined. Results were compared with a well-established
model system in which anergy was induced in cloned, allergen-specific Th2
cells. RESULTS: Impaired translocation and decreased expression of NF-AT2
and jun B were detected in PLA-specific T cell lines derived from bee venom-allergic
individuals following 16 weeks treatment compared to pre-treatment. These
results correlated with significantly reduced production of IL-4 and IL-13
and significantly increased production of IFN-gamma and IL-10 by PLA-specific
T cells. Impaired IL-4 and IL-13 production also correlated with defective
nuclear expression of NF-AT2/jun B in cloned, anergic allergen-specific Th2
cells. CONCLUSION: These results suggested that optimal production of IL-4
and IL-13 by allergen-specific T cells is dependent on the nuclear expression
of NF-AT2 and jun B. Thus, specific inhibition of NF-AT2/jun B might be an
option in novel and improved forms of allergen IT.
PMID: 12956740 [PubMed - indexed for MEDLINE]
| 18: J Allergy Clin Immunol. 2003 Jun;111(6):1255-61. |
Induction
of IL-10+CD4+CD25+ T cells by grass pollen immunotherapy.
Francis
JN, Till
SJ, Durham
SR.
Upper Respiratory Medicine, National Heart and Lung Institute, Imperial College,
Dovehouse Street, London SW3 6LY, UK.
BACKGROUND: Immunotherapy involves the modulation of allergen-specific T-cell
responses, either T(H)2-to-T(H)1 immune deviation or, in bee venom-treated
patients, induction of IL-10 production by CD4+CD25+ T cells. IL-10-producing
CD4+CD25+ regulatory T cells have emerged as potential mediators of immune
tolerance in numerous murine models of immunopathology. OBJECTIVE: The aim
of this study was to evaluate the role of IL-10 production and CD4+CD25+ T
cells in the response to grass pollen immunotherapy. METHODS: PBMCs were isolated
from patients after 1 year of grass pollen immunotherapy and from matched
untreated atopic and healthy control subjects. After 6 days of in vitro stimulation
with Phleum pratense, production of IL-10, IL-5, IL-4, and IFN-gamma and proliferation
and numbers of CD4+CD25+ T cells were measured. T cells were then stimulated
for a further 5 hours with phorbol 12-myristate 13-acetate and ionomycin and
assessed for intracellular IL-10 by means of flow cytometry. RESULTS: Patients
undergoing immunotherapy produced significantly more IL-10 than atopic control
subjects (patients undergoing immunotherapy, 116 +/- 21 pg/mL [n = 11]; atopic
patients, 30 +/- 5 pg/mL [n = 11]; P <.001), and the number of CD4+CD25+
cells identified after allergen stimulation was also greater in the immunotherapy
group. The numbers of CD4+CD25+ T cells correlated positively with activation
as measured by proliferation in both of the control groups but not in the
immunotherapy group. Moreover, only T cells from patients undergoing immunotherapy
were positive for intracellular IL-10, and these were almost exclusively CD4+CD25+
cells. CONCLUSION: Grass pollen immunotherapy results in a population of circulating
T cells that express the IL-10(+) CD4+CD25+ phenotype in response to allergen
restimulation.
Publication Types:
PMID: 12789226 [PubMed - indexed for MEDLINE]
| 19: Toxicon. 2003 Jun;41(7):861-70. |
Inhibition
of mammary carcinoma cell proliferation in vitro and tumor growth in vivo
by bee venom.
Orsolic
N, Sver
L, Verstovsek
S, Terzic
S, Basic
I.
Department of Animal Physiology, Faculty of Science, University of Zagreb,
Rooseveltov trg 6, 10 000 Zagreb, Croatia. norsolic@yahoo.com
The possible tumor growth- and metastasis-inhibiting effects of bee venom
in mice and in tumor cell cultures were studied. The tumor was a transplantable
mammary carcinoma (MCa) of CBA mouse. Intravenous administration of bee venom
to mice significantly reduced the number of metastases in the lung. However,
subcutaneous administration of bee venom did not reduce the number of lung
metastases, indicating that the antitumor effect of the venom could be highly
dependent on the route of injection as well as close contact between the components
of the venom and the tumor cells, as was shown by in vitro studies on MCa
cells. We also observed variations in immunological parameter induced by bee
venom. We proposed that bee venom has an indirect mechanism of tumor growth
inhibition and promotion of tumor rejection that is based on stimulation of
the local cellular immune responses in lymph nodes. Apoptosis, necrosis, and
lysis of tumor cells are other possible mechanisms by which bee venom inhibits
tumor growth.
PMID: 12782086 [PubMed - indexed for MEDLINE]
| 20: FASEB J. 2003 Jun;17(9):1026-35. |
T helper
(Th) 2 predominance in atopic diseases is due to preferential apoptosis of
circulating memory/effector Th1 cells.
Akdis
M, Trautmann
A, Klunker
S, Daigle
I, Kucuksezer
UC, Deglmann
W, Disch
R, Blaser
K, Akdis
CA.
Swiss Institute of Allergy and Asthma Research (SIAF), Obere Strasse 22, CH-7270
Davos, Switzerland.
T cells constitute a large population of cellular infiltrate in atopic/allergic
inflammation and a dysregulated, Th2-biased peripheral immune response appears
to be an important pathogenetic factor. In atopic dermatitis, circulating
cutaneous lymphocyte-associated antigen-bearing (CLA+) CD45RO+ T cells with
skin-specific homing property represent an activated memory/effector T cell
subset. They express high levels of Fas and Fas ligand and undergo activation-induced
apoptosis. The freshly purified CLA+ CD45RO+ T cells of atopic individuals
display distinct features of in vivo-triggered apoptosis such as pro-caspase
degradation and active caspase-8 formation. In particular, the Th1 compartment
of activated memory/effector T cells selectively undergoes activation-induced
cell death, skewing the immune response toward surviving Th2 cells in atopic
dermatitis patients. The apoptosis of circulating memory/effector T cells
was confined to atopic individuals whereas non-atopic patients such as psoriasis,
intrinsic-type asthma, contact dermatitis, intrinsic type of atopic dermatitis,
bee venom allergic patients, and healthy controls showed no evidence for enhanced
T cell apoptosis in vivo. These results define a novel mechanism for peripheral
Th2 response in atopic diseases.
PMID: 12773485 [PubMed - indexed for MEDLINE]
| 21: J Allergy Clin Immunol. 2003 Apr;111(4):854-61. |
Allergen-specific
T-cell tolerance induction with allergen-derived long synthetic peptides:
results of a phase I trial.
Fellrath
JM, Kettner
A, Dufour
N, Frigerio
C, Schneeberger
D, Leimgruber
A, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
BACKGROUND: There is a need to improve the safety and efficacy of allergen-specific
immunotherapy. Long synthetic peptide-based immunotherapy was proven safe,
immunogenic, and protective in preclinical trials. OBJECTIVE: To evaluate
the safety and immunogenicity of an allergen-derived long synthetic overlapping
peptide (LSP) immunotherapy, we designed a double-blind, placebo-controlled
phase I clinical trial in patients hypersensitive to bee venom. METHODS: Patients
from the active group were injected at day 0 with a mixture of 3 LSPs mapping
the entire PLA2 molecule, a major bee venom allergen, in a dose-escalating
protocol to a maintenance dose of 100 microg per peptide repeated at days
4, 7, 14, 42, and 70. The control group was injected with human albumin. RESULTS:
Whereas specific T-cell proliferation in the peptide group increased up to
day
PMID: 12704369 [PubMed - indexed for MEDLINE]
| 22: J Allergy Clin Immunol. 2003 Feb;111(2):426-8. |
Structural
similarity between the bee venom peptides and the immunodominant human myelin
basic proteins: role for pathogenesis of acute disseminated encephalomyelitis.
Dharmasaroja P.
Publication Types:
· Letter
PMID: 12589368 [PubMed - indexed for MEDLINE]
| 23: J Neuroendocrinol. 2003 Jan;15(1):93-6. |
The anti-inflammatory
effect of bee venom stimulation in a mouse air pouch model is mediated by
adrenal medullary activity.
Kwon YB, Kim HW, Ham TW, Yoon SY, Roh DH, Han HJ, Beitz AJ, Yang IS, Lee JH.
Department of Veterinary Physiology,
College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul
National University, Suwon, South Korea.
Cutaneous electrical or chemical stimulation can produce an anti-inflammatory
effect, which is dependent on adrenal medullary-sympathetic activation. We
have previously shown that peripheral injection of bee venom (BV) also produces
a significant anti-inflammatory effect that is neurally mediated. In the present
study, we examined whether this anti-inflammatory effect is also dependent
on the adrenal gland using the mouse inflammatory air pouch model. Subcutaneous
(s.c.) BV injection produced a marked suppression of leucocyte migration and
tumour necrosis factor (TNF)-alpha concentration induced by zymosan injection
into the air pouch. The role of the adrenal gland in this suppression was
evaluated in adrenalectomized mice. Adrenalectomy significantly reversed the
suppression of leucocyte migration and TNF-alpha elevation caused by BV. Serum
concentrations of corticosteroid were increased in mice with zymosan-induced
air-pouch inflammation and this increase was reduced by BV administration,
suggesting that adrenal corticosteroid release is not involved in mediating
the anti-inflammatory effects of BV. To test this hypothesis, the corticosteroid
receptor antagonist (RU486) was administered and found not to affect the BV-induced
inhibition of leucocyte migration. By contrast, pretreatment with the beta-adrenergic
antagonist propranolol reversed the BV-induced inhibitory effect on leucocyte
migration. These results suggest that the anti-inflammatory effect of s.c.
BV administration is mediated in part by the release of catecholamines from
the adrenal medulla.
PMID: 12535175 [PubMed - indexed for MEDLINE]
| 24: Eur J Immunol. 2002 Dec;32(12):3699-707. |
Emerging
principles for the design of promiscuous HLA-DR-restricted peptides: an example
from the major bee venom allergen.
Texier
C, Pouvelle-Moratille
S, Buhot
C, Castelli
FA, Pecquet
C, Menez
A, Leynadier
F, Maillere
B.
Protein Engineering and Research Department, CEA-Saclay, Gif sur Yvette, France.
Mechanisms underlying successful immunotherapy of allergic patients operate
at the level of CD4+ helper T cells. T cell epitopes from allergens may thus
constitute interesting molecules for immunotherapy, provided they are efficient
for all patients and are not recognized by IgE. In an attempt to define such
peptides for allergy to bee venom, we have investigated the capacity of peptides
encompassing the sequence of the major bee venom allergen to stimulate PBMC
from allergic patients and to react specifically with their IgE. The region
77-110 emerged as the most frequently T cell stimulating. We then analyzed
the binding modes of the sequence 81-97 for ten different HLA-DR molecules
and introduced punctual mutations to enhance the peptide affinity for these
molecules. Six different modes have been identified on the sequence 81-97,
one mode being common to eight HLA-DR molecules. Four HLA-DR molecules can
bind the P85-97 peptide by two different modes with an equivalent affinity.
The peptide N89L has a higher affinity for DRB1*0301 and DRB3*0101 and remains
as active as the native peptide towards the other HLA-DR molecules.
PMID: 12516563 [PubMed - indexed for MEDLINE]
| 25: Int Arch Allergy Immunol. 2002 Oct;129(2):160-8. |
Individual
hymenoptera venom compounds induce upregulation of the basophil activation
marker ectonucleotide pyrophosphatase/phosphodiesterase 3 (CD203c) in sensitized
patients.
Binder
M, Fierlbeck
G, King
T, Valent
P, Buhring
HJ.
Department of Internal Medicine II, Division of Hematology and Immunology,
University of Tubingen, Germany.
BACKGROUND: Bee and wasp venom extracts contain potent allergens capable of
inducing severe clinical reactions. To analyze immediate-type hypersensitivity
to defined hymenoptera venom components, a recently developed in vitro test
was applied that is based on the upregulation of CD203c expression on basophils.
METHODS: CD203c expression on blood basophils of 9 healthy donors and 39 patients
allergic to bee and/or wasp venom was analyzed by flow cytometry before and
after activation with the purified bee venom allergens phospholipase A2 (Api
m 1), hyaluronidase (Api m 2) and melittin (Api m 4), or the purified wasp
venom allergens phospholipase A1 (Ves v 1), hyaluronidase (Ves v 2) and the
recombinant antigen 5 (Ves v 5). Venom-induced CD203c upregulation on basophils
was compared with skin tests and assessment of specific IgE. Basophils of
nonresponders were preincubated with 10 ng/ml interleukin-3 (IL-3) prior to
allergen stimulation. RESULTS: CD203c upregulation on basophils was induced
by defined hymenoptera venom components in 35/39 patients with a diagnosed
allergy to wasp and/or bee venom. Twenty-seven of the 34 tested patients with
wasp allergy showed CD203c upregulation in response to Ves v 5, 26 of these
patients also reacted with Ves v 2 and 17 with Ves v 1. Nine of 13 patients
with bee allergy reacted with Api m 1, 13 individuals with Api m 2 and none
of these patients with the minor allergen Api m
PMID: 12403934 [PubMed - indexed for MEDLINE]
| 26: Biol Pharm Bull. 2002 Jun;25(6):710-7. |
Participation
of the arachidonic acid cascade pathway in macrophage binding/uptake of oxidized
low density lipoprotein.
Beppu M, Watanabe M, Sunohara M, Ohishi K, Mishima E, Kawachi H, Fujii M, Kikugawa K.
School of Pharmacy, Tokyo University
of Pharmacy and Life Science, Hachioji, Japan.
Arachidonic acid cascade inhibitors, including phospholipase A2 inhibitors,
dexamethasone and quinacrine (mepacrine), cyclooxygenase inhibitors, indomethacin
and aspirin, and lipoxygenase inhibitor AA861, prevented foam cell formation
and cholesterol accumulation in the incubation of thioglycollate-induced mouse
peritoneal macrophages with oxidized low density lipoprotein (LDL) at 37 degrees
C for 24 h. These inhibitors similarly prevented foam cell formation of fibronectin-
and Ca ionophore A23187-stimulated macrophages. Binding and/or uptake of Dil
(1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine)-acetyl LDL by macrophages
at 37 degrees C for 3h and arachidonic acid release from macrophages at 37
degrees C for 4h were inhibited by dexamethasone. Exogenously added phospholipase
A2 of bee venom and Crotalus adamanteous venom increased arachidonic acid
release during incubation for 2 h, and increased macrophage binding and/or
uptake of Dil-acetyl LDL at 37 degrees C for 3 h, and foam cell formation
at 37 degrees C for 24 h. Protein kinase inhibitors, ML-9 and staurosporine,
that inhibited macrophage binding and/or uptake of Dil-acetyl LDL did not
inhibit arachidonic acid release, indicating that protein phosphorylation
was not involved in the arachidonic acid pathway in the macrophage scavenger
receptor activation. Nordihydroguaiaretic acid that inhibited arachidonic
acid release prevented binding and/or uptake of Dil-acetyl LDL. The release
of arachidonic acid was not enhanced by fibronectin-stimulation, indicating
that Ca influx-dependent stimulation of macrophage activity was not through
the activation of phospholipase A2. These results indicate that, as well as
the fibronectin-stimulated Ca influx pathway and protein phosphorylation pathway,
the arachidonic acid pathway participated in the activation of macrophages
to bind and take up oxidized LDL.
PMID: 12081134 [PubMed - indexed for MEDLINE]
| 27: Toxicon. 2002 May;40(5):519-26. |
Role of melittin-like
region within phospholipase A(2)-activating protein in biological function.
Ribardo
DA, Kuhl
KR, Peterson
JW, Chopra
AK.
Department of Microbiology and Immunology, University of Texas Medical Branch,
301 University Boulevard, Galveston, TX 77555-1070, USA.
Phospholipase A(2)-activating protein (PLAA) has been implicated in the production
of prostaglandins (e.g. PGE(2)) via activation of phospholipases in various
stimulated cell types. Human PLAA, with 738 amino acid (aa) residues, contains
a region of 38% homology (aa 503-538) with the 26-aa long melittin peptide,
a major component of bee venom and a reported regulator of phospholipase A(2)
and phospholipase D activity. To learn more about the role of PLAA in the
production of eicosanoids and other inflammatory mediators, we synthesized
a murine PLAA peptide (36-aa long) having homology to melittin, as well as
to human and rat PLAA. The PLAA peptide and melittin increased the expression
of genes encoding the proinflammatory cytokine tumor necrosis factor alpha
(TNFalpha) and cyclooxygenase-2 (COX-2), which is involved in PGE(2) production.
We determined that the C-terminal region of the PLAA peptide (aa 515-538)
was essential, since truncation of the C-terminal end of the PLAA peptide
significantly reduced expression of genes encoding TNFalpha and COX-
| 28: Int Arch Allergy Immunol. 2001 Dec;126(4):335-42. |
Erratum in:
· Int Arch Allergy Immunol 2002 Apr;127(4):293.
Hymenoptera-venom-induced
upregulation of the basophil activation marker ecto-nucleotide pyrophosphatase/phosphodiesterase
Platz
IJ, Binder
M, Marxer
A, Lischka
G, Valent
P, Buhring
HJ.
University of Tubingen, Department of Internal Medicine II, Division of Hematology
and Immunology, Tubingen, Germany.
BACKGROUND: Bee and wasp venoms are potent allergens capable of inducing severe
clinical reactions. To detect immediate-type hypersensitivity to these allergens,
a rapid in vitro test was developed that relies on the upregulation of ecto-nucleotide
pyrophosphatase/phosphodiesterase 3 (E-NPP3) on activated basophils. METHODS:
Blood basophils of 13 healthy donors and 22 patients allergic to bee or wasp
venom were analyzed for E-NPP3 (CD203c) expression using monoclonal antibody
97A6. Basophils were analyzed by flow cytometry after activation with anti-IgE
antibody or allergen. Venom-induced E-NPP3 upregulation on basophils was compared
with diagnostic parameters, including skin tests and assessment of specific
IgE. In selected samples, the increase in E-NPP3 expression on activated basophils
was compared with histamine release and CD63 upregulation. RESULTS: In 20/22
patients sensitized to wasp or bee venom, E-NPP3 expression on basophils was
upregulated in response to activation by allergen or anti-IgE. The maximum
increase in E-NPP3 expression (above ten times of baseline) was achieved after
15 min of stimulation with 1 microg/ml of allergen or anti-IgE antibody. Sensitized
individuals who failed to upregulate E-NPP3 in response to IgE receptor cross-linking
also failed to induce histamine release and CD63 upregulation. CONCLUSIONS:
Flow cytometric determination of hymenoptera-venom-induced upregulation of
E-NPP3 is a novel in vitro test to identify sensitized individuals. Copyright
2002 S. Karger AG, Basel
Publication Types:
PMID: 11815741 [PubMed - indexed for MEDLINE]
| 29: Mol Pharmacol. 2001 Aug;60(2):341-7. |
A peptide
derived from bee venom-secreted phospholipase A2 inhibits replication of T-cell
tropic HIV-1 strains via interaction with the CXCR4 chemokine receptor.
Fenard
D, Lambeau
G, Maurin
T, Lefebvre
JC, Doglio
A.
Laboratoire de Virologie, Institut National
de la Sante et de
PMID: 11455021 [PubMed - indexed for MEDLINE]
| 30: Ther Umsch. 2001 May;58(5):274-7. |
[Principles of specific
immunotherapy of IgE-induced allergic reactions]
[Article in German]
Akdis
CA, Blaser
K.
Schweizerisches Institut fur Allergie- und Asthmaforschung (SIAF), Davos.
Allergen-specific immunotherapy (SIT) aims to selectively skew an allergic
immune response into a normal immunity. It appeared that the induction of
specific anergy in peripheral T cells and reactivation of anergized T cells
by microenvironmental cytokines represent two key steps in the mechanism of
SIT. In SIT of bee venom allergy the proliferative and cytokine responses
were significantly suppressed within seven days, simultaneously with an increase
in IL-10 production. IL-10 induces total anergy in T cells by autokrine interaction.
In addition, it can counter-regulate IgE and IgG4 synthesis. The addition
of blocking anti-IL-10 to stimulated PBMC fully reconstituted the proliferative
and cytokine responses in anergized T-cells. Again, particular cytokines are
able to reactivate anergic T cells to produce distinct IFN-gamma/IL-2 or IL-4/IL-13
dominated T cell cytokine patterns and direct by this way SIT towards successful
or unsuccessful treatment. The suppression of T cells by IL-10 is an active
biochemical process, which depends on the interaction of the ligated IL-10
receptor with the CD28 costimulatory signaling pathway in T cells.
PMID: 11407227 [PubMed - indexed for MEDLINE]
| 31: J Allergy Clin Immunol. 2001 May;107(5):914-20. |
Api m 6:
a new bee venom allergen.
Kettner
A, Hughes
GJ, Frutiger
S, Astori
M, Roggero
M, Spertini
F, Corradin
G.
Institute of Biochemistry, University of Lausanne, Lausanne, Switzerland.
BACKGROUND: Characterization of the primary structure of allergens is a prerequisite
for the design of new diagnostic and therapeutic tools for allergic diseases.
OBJECTIVE: The purpose of this study was the identification and characterization
of a low-molecular-weight, IgE-binding, bee venom (BV) allergen. METHODS:
BV proteins were separated by using size exclusion chromatography and HPLC.
IgE antibody binding to purified proteins was analyzed by means of immunoblotting,
and T-cell response was analyzed by means of proliferation assay. Amino acid
sequence was determined with 2 approaches, namely Edman degradation and carboxy
terminal analysis with mass spectrometry. RESULTS: Api m 6, which migrated
as an 8-kd band in SDS-PAGE, was frequently (42%) recognized by IgE from BV-hypersensitive
patients. In addition, PBMCs from BV-hypersensitive patients, as well as from
a normal control subject, proliferated in response to this allergen. Api m
6 exists as 4 isoforms of 7190, 7400, 7598, and 7808 d, respectively. Amino
acid sequences obtained from HPLC-purified preparations revealed that the
isoforms were constituted of a common central core of 67 residues, only differing
in the amino- and carboxy-terminal ends. Api m 6 showed no significant sequence
homology with known proteins. CONCLUSIONS: We have identified and sequenced
a new BV allergen that elicits a strong IgE and T-cell response in a large
number of BV-hypersensitive patients. Api m 6 should be considered in the
diagnostic and therapeutic approach of BV immunotherapy on the basis of peptides
or recombinant proteins.
PMID: 11344362 [PubMed - indexed for MEDLINE]
| 32: Int Arch Allergy Immunol. 2001 Jan-Mar;124(1-3):180-2. |
Mechanism
of IL-10-induced T cell inactivation in allergic inflammation and normal response
to allergens.
Akdis
CA, Joss
A, Akdis
M, Blaser
K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
akdism@siaf.unizh.ch
BACKGROUND: Induction of specific unresponsiveness (tolerance/anergy) in peripheral
T cells and recovery by cytokines from the tissue microenvironment represent
two key steps in specific immunotherapy (SIT) with whole allergen or antigenic
T cell peptides. METHODS: Antigen-specific T cell responses and molecular
mechanisms of T cell inactivation were investigated during conventional SIT,
T cell epitope peptide immunotherapy and natural exposure to bee venom in
allergic and hyperimmune individuals. RESULTS: T cell unresponsiveness, initiated
by autocrine action of IL-10, is characterized by suppressed proliferative
and cytokine responses. The unresponsive T cells can be reactivated by different
cytokines that may mimic the microenvironmental cytokine influence. IL-10
initiates peripheral tolerance by blocking the CD28 costimulatory signal in
T cells. Coprecipitation experiments reveal that upon stimulation CD28 and
IL-10 receptor are physically associated in T cells. Accordingly, IL-10 binding
to its receptor inhibits CD28 tyrosine phosphorylation, the initial step of
the CD28 signaling pathway. This leads to inhibition of phosphatidylinositol
3-kinase p85 binding to CD28. IL-10 only affects T cells that receive a stimulation
with low numbers of triggered T cell receptors and that require costimulatory
signals by CD28. CONCLUSION: These data demonstrate the pivotal role of autocrine
IL-10 and the interaction of its receptor with CD28 in the induction of T
cell tolerance as an immunoregulatory mechanism controlling antigen-specific
T cell responses. Copyright 2001 S. Karger
AG, Basel
Publication Types:
· Review
PMID: 11306962 [PubMed - indexed for MEDLINE]
| 33: J Immunol. 2001 Mar 1;166(5):3612-21. |
Antigen-independent
suppression of the allergic immune response to bee venom phospholipase A(2)
by DNA vaccination in CBA/J mice.
Jilek
S, Barbey
C, Spertini
F, Corthesy
B.
Division of Immunology and Allergy, R & D Laboratory, Centre Hospitalier
Universitaire Vaudois, Lausanne, Switzerland.
Phospholipase A(2) (PLA(2)) is one of the major honey bee venom allergens
for humans. To assess the long-term prevention of allergic reactions by DNA
vaccination, a PLA(2)-CBA/J mouse model was employed using empty or PLA(2)
sequence-carrying DNA plasmids. Early skin application of either DNA construct
before (prophylactic approach) or after (therapeutic approach) sensitization
with PLA(2)/alum led to reduced PLA(2)-specific IgE and IgG1 titers at 7 mo,
with concomitant rise in IgG2a and IgG3. Splenocytes recovered at 5-6 mo after
the last DNA administration exhibited a sustained IFN-gamma and IL-10 secretion
and reduced IL-4 production. Recall challenge with PLA(2) boosted IFN-gamma
and IL-10 secretion, suggesting the reactivation of quiescent memory Th1 lymphocytes.
Mice from the prophylactic groups were fully protected against anaphylaxis,
whereas 65% of the animals recovered in the therapeutic groups. Th1-polarized
immune responses were also active in mice vaccinated with an empty plasmid
32 wk before sensitization with another Ag (OVA). This is the first demonstration
that the Ag-coding sequence in DNA vaccine is not necessary to promote immune
modulation in naive and sensitized animals for a prolonged period, and has
relevance for the understanding of the innate and induced mechanisms underlying
gene immunotherapy in long-term treatment of allergy.
PMID: 11207323 [PubMed - indexed for MEDLINE]
| 34: Eur J Immunol. 2000 Dec;30(12):3533-41. |
Expression
of cutaneous lymphocyte-associated antigen on human CD4(+) and CD8(+) Th2
cells.
Akdis
M, Klunker
S, Schliz
M, Blaser
K, Akdis
CA.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
akdism@siaf.unizh.ch
The cutaneous lymphocyte-associated antigen (CLA) represents the homing receptor
involved in selective migration of memory/effector T cells to the skin. Numerous
reports demonstrated distinct CLA expression on Th1 cells. However, T cells
isolated from skin lesions and CLA(+) T cells circulating in peripheral blood
of atopic dermatitis patients expressed high IL-5 and IL-13. Accordingly,
we investigated the regulation of CLA on human type 1 and type 2 T cells.
CLA was induced on freshly generated Th1 and Tc1 cells only, but not on those
of type 2. Anti-CD3 stimulation was sufficient to induce CLA on Th2 cells
in the absence of serum in the culture medium. In serum containing medium,
IL-4 inhibited CLA and related alpha-fucosyltransferase mRNA expression. IL-12
and/or staphylococcal enterotoxin B (SEB) stimulation up-regulated CLA expression
on either Th2 and Tc2 cells. On stimulation with IL-12, CLA was expressed
on the surface of bee venom phospholipase A(2)-specific Th1, Th2, Th0 and
T regulatory 1 clones, representing non-skin-related antigen-specific T cells.
In addition, CLA could be re-induced on T cells that had lost CLA expression
upon resting. These results suggest that skin-selective homing is not restricted
to functional and phenotypic T cell subsets.
PMID: 11093173 [PubMed - indexed for MEDLINE]
| 35: J Immunol. 2000 Sep 15;165(6):3497-505. |
Inducing
tolerance by intranasal administration of long peptides in naive and primed
CBA/J mice.
Astori
M, von
Garnier C, Kettner
A, Dufour
N, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
To assess the capacity of a peptide-based immunotherapy to induce systemic
tolerance via the nasal route, we designed three long overlapping peptides
of 44-60 aa covering the entire sequence of phospholipase A2 (PLA2), a major
bee venom allergen. Both prophylactic and therapeutic intranasal administrations
of long peptides to PLA2-hypersensitive CBA/J mice induced specific T cell
tolerance to the native allergen. In prophylactic conditions, this tolerance
was marked by a suppression of subsequent specific IgE response, whereas the
therapeutic approach in presensitized mice induced a more than 60% decrease
in PLA2-specific IgE. This decline was associated with a shift in the cytokine
response toward a Th1 profile, as demonstrated by decreased PLA2-specific
IgG1 and enhanced IgG2a levels, and by a decline in the specific IL-4/IFN-gamma
ratios. T cell transfer from long peptide-tolerized mice to naive animals
abrogated the expected anti-PLA2 IgE and IgG1 Ab response, as well as specific
T cell proliferation, but enhanced specific IgG2a response upon sensitization
with PLA2. These events were strongly suggestive of a clonal anergy affecting
more profoundly Th2 than the Th1 subsets. In conclusion, these results demonstrate
that allergen-derived long peptides delivered via the nasal mucosa may offer
an alternative to immunotherapy with native allergens without the inherent
risk of systemic anaphylactic reactions. Moreover, long peptides, in contrast
to immunotherapy strategies based on short peptides, have the advantage of
covering all potential T cell epitopes, and may represent novel and safe tools
for the therapy of allergic diseases.
PMID: 10975871 [PubMed - indexed for MEDLINE]
| 36: Eur J Immunol. 2000 Jun;30(6):1638-45. |
Allergen-derived
long peptide immunotherapy down-regulates specific IgE response and protects
from anaphylaxis.
von
Garnier C, Astori
M, Kettner
A, Dufour
N, Heusser
C, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
To evaluate a long peptide-based allergy vaccine in a murine model, CBA/J
mice were sensitized with low dose alum-adsorbed phospholipase A2 (PLA2),
a major bee venom allergen. Presensitized mice were treated by daily i.p.
injections of a mixture of three long overlapping peptides (44- to 60-mer)
spanning the entire PLA2 molecule (100 microg/peptide) for 6 consecutive days.
This therapeutic approach induced a sharp drop in PLA2-specific IgE, an increase
in specific IgG2a, and a marked T cell hyporesponsiveness. T cell cytokine
secretion was characterized by a shift from a Th2 to a Th1 profile. Prophylactic
treatment of naive mice with long peptides prior to sensitization with PLA2
induced a comparable modulation of B and T cell responses. Upon i.p. challenge
with native PLA2, presensitized mice treated with the long peptide mixture
were fully protected from anaphylaxis. This indicated that allergen-derived
long overlapping peptides were safe and able to modulate an established Th2
response or to prevent its development. Furthermore, long peptide-based immunotherapy
provided clinical protection against anaphylaxis, thus appearing as a promising
approach of the therapy of allergic diseases.
PMID: 10898500 [PubMed - indexed for MEDLINE]
| 37: Int J Pancreatol. 2000 Feb;27(1):29-38. |
Role of various
phospholipases A2 and inhibitors in the pathogenesis and prevention of pancreatic
acinar cell necrosis: studies with isolated rat pancreatic acini.
Mossner
J, Wessig
C, Ogami
Y, Keim
V.
Department of Internal Medicine II, University of Leipzig, Germany. moej@server3.medizin.uni-leipzig.de
BACKGROUND: Phospholipase A2 (PLA2) may play a central role in the pathogenesis
of pancreatic acinar cell necrosis. Several questions, however, are unsolved:
Is acinar cell necrosis caused by PLA2 derived from infiltrating leukocytes
or from pancreatic PLA2 itself? Does PLA2 cause cellular lysis by the release
of lysolecithin from lecithin or by generation of free radicals? The aims
of this study were to determine which form of PLA2 is responsible for cellular
damage and how to inhibit its action. METHODS: Isolated rat pancreatic acini
were prepared by collagenase digestion. Newly synthesized proteins were labeled
by 35S-methionine. Acini were incubated in buffer to which various factors,
such as porcine pancreatic PLA2 or bee venom PLA2, homogenates of either leukocytes
or pancreatic homogenates, all with or without lecithin and with or without
potential inhibitors (aprotinin, 4-bromophenacylbromide, BM 16.2115, quinacrine,
various analogs of arachidonic acid), or free radicals (hydrogen peroxide,
xanthine/ xanthine oxidase) with or without allo-purinol or dismutase/catalase
were added. Cellular destruction was measured by the release of radiolabeled
proteins. RESULTS: PLA2 alone, free radicals, and granulocytes were not harmful
to acini within 30 min of incubation. Free radicals caused significant release
of radiolabeled proteins only after 3 h of incubation; this release could
be inhibited by scavengers. Incubation of pancreatic acini with PLA2 in combination
with lecithin caused rapid release of radiolabeled proteins. Addition of high
concentrations of enterokinase activated pancreatic homogenates both alone
and with lecithin caused release of cellular proteins, suggesting that pancreatic
PLA2 uses lecithin from pancreatic membranes as substrate. Almost all tested
potential inhibitors of PLA2 were unable to prevent the destruction caused
by either pancreatic or bee venom PLA2 and lecithin. However, HK
| 38: Arch Immunol Ther Exp (Warsz). 2000;48(2):107-10. |
Histamine receptor
expression on peripheral blood lymphocytes is influenced by specific and nonspecific
activation.
Zak-Ejmark T,
Kraus-Filarska M,
Malolepszy J,
Jankowska R,
Jutel M, Nowak IA, Nadobna G.
Department of Internal Medicine
and Allergology, University Medical School, Wroclaw, Poland.
Histamine is a physiological mediator which exerts both effector and regulatory
functions through its receptors on various cells. The aim of the study was
to investigate changes in histamine receptor expression on peripheral blood
lymphocytes affected by stimulation with both specific and nonspecific stimuli.
Lymphocytes were obtained from both healthy and allergic subjects. Cells were
incubated with various allergens (mixed grass pollen, Lolium perenne, Dermatophagoides
pteronyssinus 1, bee venom, phospholipase A2) and nonspecific (fMLP, PMA/ionomycin,
LPS) stimuli. The percentage of histamine-binding cells was determined with
a fluorescence microscope after incubation with histamine-fluorescein. In
control subjects histamine binding after stimulation with allergens was not
significantly changed. In contrast, in allergic subjects stimulation with
specific allergens resulted in significantly increased histamine binding.
Nonspecific stimulation caused increased histamine binding to lymphocytes
in both allergic subjects and healthy controls. We conclude that specific
and nonspecific activation of lymphocytes is associated with increased expression
of histamine receptors.
PMID: 10807051 [PubMed - indexed for MEDLINE]
| 39: Immunobiology. 2000 Jan;201(3-4):391-405. |
Lipopeptides as
immunoadjuvants and immunostimulants in mucosal immunization.
Baier
W, Masihi
N, Huber
M, Hoffmann
P, Bessler
WG.
Institut fur Molekulare Medizin und Zellforschung, Albert-Ludwigs-Universitat,
Freiburg, Germany.
In previous studies we have shown that lipopeptides constitute potent immunoadjuvants
in mice, rabbits and other species: in parenteral immunization, lipopeptide
adjuvants were comparable, or in some cases superior to Freund's adjuvant,
and were devoid of the side effects of this additive. Here we demonstrate
that lipopeptides also constitute adjuvants for mucosal immunizations. The
serum antibody responses against the wheat storage protein gliadin, the bee
venom constituent melittin, or the hen egg protein ovalbumin could in most
cases be enhanced more than 100-fold by the lipopeptide P3CSK4, applied via
the nasal route. An enhanced specific antibody level could also be detected
in supernatants of cell cultures prepared from spleens, Peyer's patches, lungs
and mesenteric lymph nodes of immunized mice. Moreover, the lipopeptide P3CSK4
enhanced chemiluminescence in mouse spleen cells and peritoneal macrophages
in vitro, indicating a macrophage-activating effect. Finally, nasal application
of lipopeptide increased protection against a lethal infection of influenza.
Our findings are of importance for the improvement of immunizations and might
lead to more effective vaccines.
PMID: 10776795 [PubMed - indexed for MEDLINE]
| 40: Allergy. 1999 Jul;54(7):742-7. |
The frequency
of phospholipase A2 binding of basophilic granulocytes does not decrease during
bee-venom-specific immunotherapy.
Irsch
J, Konig
C, Lohndorf
A, Tesch
H, Krieg
T, Merk
H, Radbruch
A, Hunzelmann
N.
Department of Genetics, University of Cologne, Germany.
BACKGROUND: The major allergenic component of bee venom is phospholipase A2
(PLA2). METHODS: In this study, PLA2 was used to analyze and enrich PLA2-binding
cells from peripheral blood by high gradient magnetic cell sorting. RESULTS:
In normal donors, the frequency of allergen (PLA2)-binding cells among peripheral
blood mononuclear cells (PBMC) as determined by flow cytometry is below 0.1%,
whereas in bee-venom-allergic patients, PLA2-binding cells are readily detectable
at frequencies of up to 2.3%. In severely bee-venom-allergic patients, many
basophilic granulocytes are present, as defined by anti-CD9, CD25, and CD38
mAb, comprising up to 95% of the PLA2-binding cells. From blood of allergic
and normal donors, about equal absolute numbers of allergen-binding CD19/21-positive
B cells can be enriched. Severe anaphylactic reactions (Mueller grade IV)
and failure of or adverse reactions during immunotherapy are associated with
high numbers of circulating allergen-binding basophils. Interestingly, in
the patients studied, the number of PLA2-binding basophilic granulocytes did
not markedly change during rush immunotherapy and up to 6 months of maintenance
immunotherapy. CONCLUSIONS: The specific and reproducible enrichment of PLA2-binding
cells provides a new tool for the analysis and monitoring of effector cells
in bee-venom-allergic patients with immediate-type hypersensitivity.
PMID: 10442531 [PubMed - indexed for MEDLINE]
| 41: Int Immunol. 1999 Aug;11(8):1313-26. |
On the diversity
and heterogeneity of H-2(d)-restricted determinants and T cell epitopes from
the major bee venom allergen.
Texier
C, Herve
M, Pouvelle
S, Menez
A, Maillere
B.
Departement d'Ingenierie et d'Etudes des Proteines, CEA-Saclay, 91191 Gif
sur Yvette, France.
One of the main limitations of using synthetic peptides for immunotherapy
in allergic patients is the difficulty to delineate the immunodominant T cell
epitopes which are necessarily dependent on HLA molecules. We have thus addressed
the question of the role of MHC II molecules in immunodominant epitopes selection
in the particular case of the major bee venom allergen (API m1). To exhaustively
and easily explore it, we used BALB/c mice whose H-2 haplotype is associated
with high IgE and IgG responses to API m1. By means of extensive sets of synthetic
peptides, we investigated the specificity of polyclonal T cells and monoclonal
hybridomas from mice immunized with API m1 and delineated four immunodominant
regions, restricted to either the I-E(d) or the I-A(d) molecule. All the peptides
were also tested for their capacity to bind to immunopurified MHC II molecules.
Eight determinants of high affinity were identified. They clustered into three
distinct regions and were largely overlapping. They included all the immunodominant
epitopes, but half of them were not capable of stimulating T cells. Strikingly,
interacting surfaces with either the TCR or MHC II molecule greatly differed
from one determinant to another. In one case, we observed that flanking regions
exerted a particular action on T cell stimulation which prevented the fine
epitope localization. Our results underline the diversity and complexity of
MHC II-restricted determinants and T cell epitopes from the major bee venom
allergen, even in a single haplotype. These data also participate in the development
of alternative approaches to conventional immunotherapy.
PMID: 10421789 [PubMed - indexed for MEDLINE]
| 42: Clin Exp Allergy. 1999 Mar;29(3):394-401. |
IgE and T-cell
responses to high-molecular weight allergens from bee venom.
Kettner
A, Henry
H, Hughes
GJ, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
BACKGROUND: Bee venom contains multiple allergens with a wide distribution
of molecular weight. In contrast with conventional bee venom desensitization,
peptide or recombinant allergen immunotherapy may have to take into account
patients' individual patterns of humoral or cellular response. OBJECTIVE:
To study immunoglobulin (Ig)E and T-cell responses to high-molecular weight
bee venom allergens >/= 50 kDa. METHODS: Bee venom proteins were separated
by size exclusion chromatography and fractions were characterized by one and
two-dimensional gel electrophoresis. IgE antibody binding to bee venom fractions
was analysed by immunoblotting and T-cell responses by proliferation assay.
RESULTS: Among 38 bee venom-hypersensitive patients, IgE recognition pattern
of bee venom allergens varied greatly. IgE bound mainly to phospholipase A2
and furthermore to several proteins >/= 50 kDa (50, 54, 69, 84 and 94 kDa).
N-terminal sequences of these proteins showed no homology with known proteins.
In addition, peripheral mononuclear cells from patients as well as from nonatopic
donors strongly proliferated in response to those proteins. CONCLUSIONS: Although
present in low amounts, high-molecular weight allergens from bee venom elicit
strong IgE and T-cell responses, and may need to be considered as clinically
relevant. Therefore, the development of peptide or recombinant protein-based
immunotherapy for bee venom allergy may require careful characterization of
such allergens.
PMID: 10202349 [PubMed - indexed for MEDLINE]
| 43: FASEB J. 1999 Apr;13(6):603-9. |
IL-10-induced
anergy in peripheral T cell and reactivation by microenvironmental cytokines:
two key steps in specific immunotherapy.
Akdis
CA, Blaser
K.
Swiss Institute of Allergy and Asthma Research, CH-7270 Davos, Switzerland.
akdisac@siaf.unizh.ch
Specific immunotherapy (SIT) is widely used for treatment of allergic diseases
and could potentially be applied in other immunological disorders. Induction
of specific unresponsiveness (anergy) in peripheral T cells and recovery by
cytokines from the tissue microenvironment represent two key steps in SIT
with whole allergen or antigenic T cell peptides (PIT). The anergy is directed
against the T cell epitopes of the respective antigen and characterized by
suppressed proliferative and cytokine responses. It is initiated by autocrine
action of IL-10, which is increasingly produced by the antigen-specific T
cells. Later in therapy, B cells and monocytes also produce IL-10. The anergic
T cells can be reactivated by different cytokines. Whereas IL-15 and IL-2
generate Th1 cytokine profile and an IgG4 antibody response, IL-4 reactivates
a Th2 cytokine pattern and IgE antibodies. Increased IL-10 suppresses IgE
and enhances IgG4 synthesis, resulting in a decreased antigen-specific IgE:IgG4
ratio, as observed normally in patients after SIT or PIT. The same state of
anergy against the major bee venom allergen, phospholipase A2, can be observed
in subjects naturally anergized after multiple bee stings. Together, these
data demonstrate the pivotal role of autocrine IL-
· Review
PMID: 10094921 [PubMed - indexed for MEDLINE]
| 44: J Pharmacol Exp Ther. 1999 Apr;289(1):166-72. |
Effects of
petrosaspongiolide M, a novel phospholipase A2 inhibitor, on acute and chronic
inflammation.
Garcia-Pastor P,
Randazzo A, Gomez-Paloma L,
Alcaraz MJ, Paya M.
Departamento de Farmacologia, Universidad de Valencia, Facultad de Farmacia,
Valencia, Spain.
The marine product petrosaspongiolide
M is a novel inhibitor of phospholipase A2 (PLA2), showing selectivity for
secretory PLA2 versus cytosolic PLA2, with a potency on the human synovial
enzyme (group II) similar to that of manoalide. This compound was more potent
than manoalide on bee venom PLA2 (group III) and had no effect on group I
enzymes (Naja naja and porcine pancreatic PLA2). Inhibition of PLA2 was also
observed in vivo in the zymosan-injected rat air pouch, on the secretory enzyme
accumulated in the pouch exudate. Petrosaspongiolide M decreased carrageenan
paw edema in mice after the oral administration of 5, 10, or 20 mg/kg. This
marine metabolite (0.01-1.0 micromol/pouch) induced a dose-dependent reduction
in the levels of prostaglandin (PG)E2, leukotriene B4, and tumor necrosis
factor-alpha in the mouse air pouch injected with zymosan 4 h after the stimulus.
It also had a weaker effect on cell migration. The inflammatory response of
adjuvant arthritis was reduced by petrosaspongiolide M, which also inhibited
leukotriene B4 levels in serum and PGE2 levels in paw homogenates. In contrast
with indomethacin, this marine compound did not reduce PGE2 levels in stomach
homogenates. Petrosaspongiolide M is a new inhibitor of secretory PLA2 in
vitro and in vivo, with anti-inflammatory properties in acute and chronic
inflammation.
PMID: 10087000 [PubMed - indexed for MEDLINE]
| 45: J Immunol. 1999 Feb 1;162(3):1836-42. |
An altered
peptide ligand specifically inhibits Th2 cytokine synthesis by abrogating
TCR signaling.
Faith
A, Akdis
CA, Akdis
M, Joss
A, Wymann
D, Blaser
K.
Swiss Institute of Allergy and Asthma Research, Davos, Switzerland. siaf@siaf.unizh.ch
Altered peptide ligands (APL) can modify T cell effector function by their
diversity in binding to the TCR or MHC class II-presenting molecules. The
capacity to inhibit Th2 cytokine production by allergen-specific T cells would
contribute to combating allergic inflammation. The presence of APL generated
by Ala-substitutions in a synthetic dodeca-peptide spanning an immunodominant
epitope of bee venom phospholipase A2 (PLA) was investigated in human T cells.
Four of five substituted peptides reduced proliferation, IL-4, and IFN-gamma
production by cloned PLA-specific Th0 cells proportionately. However, one
APL, PLA-F82A, inhibited IL-4 but had no effect on IFN-gamma production. This
uncoupling of IL-4 from IFN-gamma production was also observed on immunogenic
restimulation of the cloned T cells pre-exposed to the APL/APCs. It appeared
to result from lower affinity of binding to MHC class II by the APL compared
with the native peptide. The APL also inhibited IL-4 production by polyclonal
T cells. In consequence of the change in cytokine secretion, the production
of IgG4 in vitro increased by PLA-F82A stimulation, compared with the native
peptide. Exposure of the cloned T cells to either the APL or the native peptide,
in the absence of professional APC, induced anergy such that proliferation
and production of IL-4, IL-5, and IL-13 was abrogated on immunogenic rechallenge.
Defective T cell activation appeared to result from alterations in transmembrane
signaling through the TCR, specifically to lack of tyrosine phosphorylation
of the tyrosine kinase, ZAP-70.
PMID: 9973449 [PubMed - indexed for MEDLINE]
| 46: Biochim Biophys Acta. 1998 Sep 16;1425(1):74-80. |
Cytotoxicity of
pilosulin
Wu QX, King
MA, Donovan
GR, Alewood
D, Alewood
P, Sawyer
WH, Baldo
BA.
Molecular Immunology, Kolling Institute of Medical Research, Royal North Shore
Hospital, St. Leonards, NSW, Australia.
The synthetic peptide pilosulin 1, corresponding to the largest defined allergenic
polypeptide found in the venom of the jumper ant Myrmecia pilosula, inhibited
the incorporation of [methyl-3H]thymidine into proliferating Epstein-Barr
transformed (EBV) B-cells. The LD50 was four-fold lower in concentration than
melittin, a cytotoxic peptide found in honey bee venom. Loss of cell viability
was assessed by flow cytometry by measuring the proportion of cells that fluoresced
in the presence of the fluorescent dye 7-aminoactinomycin D. Examination of
proliferating EBV B-cells indicated that the cells lost viability within a
few minutes exposure to pilosulin 1. Partial peptides of pilosulin 1 were
less efficient in causing loss of cell viability and the results suggest that
the 22 N-terminal residues are critical to the cytotoxic activity of pilosulin
1. Normal blood white cells were also labile to pilosulin 1. T- and B-lymphocytes,
monocytes and natural killer cells, however, were more labile than granulocytes.
Analysis of pilosulin 1 using circular dichroism indicated that, in common
with melittin and other Hymenoptera venom toxins, it had the potential to
adopt an alpha-helical secondary structure.
PMID: 9813247 [PubMed - indexed for MEDLINE]
| 47: Clin Exp Allergy. 1998 Jul;28(7):839-49. |
Enzymatic
activity of soluble phospholipase A2 does not affect the specific IgE, IgG4
and cytokine responses in bee sting allergy.
Wymann
D, Akdis
CA, Blesken
T, Akdis
M, Crameri
R, Blaser
K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos.
BACKGROUND: The soluble bee venom phospholipase A2 (PLA) represents the major
allergen/antigen for allergic and hyperimmune individuals following bee sting.
A number of studies implicate enzymes, and PLA in particular, as potent allergens.
We have studied specific activation of T cells by enzymatically active and
inactive mutants of PLA, and secretion of cytokines regulating IgE and IgG4
antibody formation. METHODS: Recombinant (r) wild type PLA (rPLA-WT) and an
enzymatically inactive rPLA (rPLA-H34Q) were produced in Escherichia coli.
Eleven bee venom allergic patients and three hyperimmune, healthy individuals
were included in the study. After specific stimulation of PBMC with the rPLA
variants, proliferative response, IFNgamma, IL-2, IL-4, IL-5, and IL-13 production,
as well as total and PLA-specific IgE and IgG4 production, were analysed.
RESULTS: Similar levels of specific B cell recognition, proliferative and
cytokine responses were observed after stimulation with either enzymatically
active or inactive rPLA. In addition, equal amounts of antigen-specific and
total IgE and IgG4 antibodies were produced by stimulation with both forms
of rPLA. CONCLUSIONS: The enzymatic activity of PLA does not influence the
specific activation and cytokine production by T cells from bee venom-sensitized
or hyperimmune individuals, or the IgE/IgG4 antibodies synthesis by B cells
in vitro.
PMID: 9720818 [PubMed - indexed for MEDLINE]
| 48: Cytometry. 1998 Aug 1;32(4):268-73. |
Flow cytometric
analysis of cell killing by the jumper ant venom peptide pilosulin 1.
King
MA, Wu QX, Donovan
GR, Baldo
BA.
Department of Clinical Immunology, Royal North Shore Hospital, St. Leonards,
New South Wales, Australia. making@doh.health.nsw.gov.au
Pilosulin 1 is a synthetic 56-amino acid residue polypeptide that corresponds
to the largest allergenic polypeptide found in the venom of the jumper ant
Myrmecia pilosula. Initial experiments showed that pilosulin 1 lysed erythrocytes
and killed proliferating B cells. Herein, we describe how flow cytometry was
used to investigate the cytotoxicity of the peptide for human white blood
cells. Cells were labeled with fluorochrome-conjugated antibodies, incubated
with the peptide and 7-aminoactinomycin D (7-AAD), and then analyzed. The
effects of varying the peptide concentration, serum concentration, incubation
time, and incubation temperature were measured, and the cytotoxicity of pilosulin
1 was compared with that of the bee venom peptide melittin. The antibodies
and the 7-AAD enabled the identification of cell subpopulations and dead cells,
respectively. It was possible, using the appropriate mix of antibodies and
four-color analysis, to monitor the killing of three or more cell subpopulations
simultaneously. We found that 1) pilosulin 1 killed cells within minutes,
with kinetics similar to those of melittin; 2) pilosulin 1 was a slightly
more potent cytotoxic agent than melittin; 3) both pilosulin 1 and melittin
were more potent against mononuclear leukocytes than against granulocytes;
and 4) serum inhibited killing by either peptide.
PMID: 9701394 [PubMed - indexed for MEDLINE]
| 49: Clin Exp Allergy. 1997 Sep;27(9):1016-26. |
Comment in:
· Clin Exp Allergy. 1997 Sep;27(9):986-90.
Delineation of PLA2
epitopes using short or long overlapping synthetic peptides: interest for
specific immunotherapy.
Kammerer
R, Kettner
A, Chvatchko
Y, Dufour
N, Tiercy
JM, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
BACKGROUND: Venom immunotherapy is definitely indicated in severe systemic
anaphylactic reactions to bee stings, but is not devoided of risks of anaphylaxis.
Safer methods of immunotherapy need to be developed. OBJECTIVE: To delineate
phospholipase A2 T-cell epitopes using short 15mer vs long 40-60mer overlapping
peptides, and to approach the potential interest of a venom immunotherapy
based on the use of long peptides (1-60, 51-99, 90-134) mapping the whole
phospholipase A2 molecule vs a restricted number of immunodominant epitopes.
METHODS: Proliferation of a CD8+ T cell depleted peripheral blood mononuclear
cell fraction and short-term T-cell lines from unselected bee venom hypersensitive
patients in response to phospholipase A2 synthetic peptides. RESULTS: Whereas
T-cell proliferation to 15mer overlapping peptides was weak, T-cell response
to long overlapping peptides was in contrast vigorous in all patients, mostly
directed to C-terminal peptide 90-134. Our results did not support the concept
of rare dominant T-cell epitopes, and disclosed T-cell responses to multiple
epitopes in several patients. No significant IgE-binding to long overlapping
peptides was detected except in one patient against peptide 90-134. CONCLUSION:
15mer peptides might not be sensitive enough to fully delineate all potential
T-cell epitopes scattered along the allergen. Since they do not bind IgE in
vitro or only weakly, and taking into account a T-cell response frequently
directed to multiple epitopes, long overlapping peptides may represent ideal
tools for immunotherapy.
PMID: 9678833 [PubMed - indexed for MEDLINE]
| 50: Clin Exp Allergy. 1997 Sep;27(9):986-90. |
Comment on:
· Clin Exp Allergy. 1997 Sep;27(9):1016-26.
To bee or not to
be? T-cell responses to bee venom PLA2 in relation to anaphylaxis and immunotherapy.
McHugh SM.
Publication Types:
· Comment
PMID: 9678827 [PubMed - indexed for MEDLINE]
| 51: J Clin Invest. 1998 Jul 1;102(1):98-106. |
Role of interleukin
Akdis
CA, Blesken
T, Akdis
M, Wuthrich
B, Blaser
K.
Swiss Institute of Allergy and Asthma Research, CH-7270 Davos, Switzerland.
akdisac@isac@siaf.unizh.ch
The induction of allergen-specific anergy in peripheral T cells represents
a key step in specific immunotherapy (SIT). Here we demonstrate that the anergic
state results from increased IL-10 production. In bee venom (BV)-SIT the specific
proliferative and cytokine responses against the main allergen, the phospholipase
A2 (PLA), and T cell epitope-containing PLA peptides were significantly suppressed
after 7 d of treatment. Simultaneously, the production of IL-10 increased
during BV-SIT. After 28 d of BV-SIT the anergic state was established. Intracytoplasmic
cytokine staining of PBMC combined with surface marker detection revealed
that IL-10 was produced initially by activated CD4(+)CD25(+), allergen-specific
T cells, and followed by B cells and monocytes. Neutralization of IL-
|
52: J Allergy Clin Immunol. 1998 Jun;101( |
Successful
immunotherapy with T-cell epitope peptides of bee venom phospholipase A2 induces
specific T-cell anergy in patients allergic to bee venom.
Muller
U, Akdis
CA, Fricker
M, Akdis
M, Blesken
T, Bettens
F, Blaser
K.
Medical Division, Zieglerspital, Bern, Switzerland.
BACKGROUND: Specific immunotherapy with honeybee venom (BV) is highly effective,
but allergic side effects can occur during treatment. Immunotherapy with peptides
containing major T-cell epitopes of the relevant allergen or allergens provides
an alternative strategy without these problems. OBJECTIVE: The study investigates
the immunologic mechanisms and clinical effects of immunotherapy with T-cell
epitope peptides of the major BV allergen, the phospholipase A2 (PLA). METHODS:
Five patients with IgE-mediated systemic allergic reactions to bee stings
were treated with a mixture of three T-cell epitope peptides of PLA. Ten patients
allergic to BV receiving whole BV immunotherapy served as control subjects.
Increasing doses of the peptide mixture, up to a maintenance dose of 100 microg,
were administered subcutaneously within 2 months. The patients were then challenged
with PLA and 1 week later with a bee sting. The cellular and humoral immune
response was measured in vitro. RESULTS: No allergic side effects were caused
by the peptide immunotherapy, and all patients tolerated the challenge with
PLA without systemic allergic symptoms. Two patients developed mild systemic
allergic reactions after the bee sting challenge. After peptide immunotherapy,
specific proliferative responses to PLA and the peptides in peripheral blood
mononuclear cells were decreased in successfully treated patients. The production
of TH2 and TH1 cytokines was inhibited, and B cells were not affected in their
capacity to produce specific IgE and IgG4 antibodies. Their levels increased
after allergen challenge in favor of IgG4. CONCLUSIONS: Immunotherapy of BV
allergy with short T-cell peptides of PLA induces epitope-specific anergy
in peripheral T cells and changes the specific isotype ratio in a fashion
similar to that of conventional immunotherapy in successfully treated patients.
PMID: 9648701 [PubMed - indexed for MEDLINE]
| 53: J Investig Allergol Clin Immunol. 1998 Mar-Apr;8(2):109-14. |
Lymphocyte subpopulations,
cytokine release and specific immunoglobulin G in reactive and nonreactive
beekeepers.
Annila
I, Hurme
M, Miettinen
A, Kuusisto
P, Nieminen
MM.
Department of Respiratory Medicine, Medical School, University of Tampere,
Finland.
Although bee venom sensitization and systemic sting reactions are common among
beekeepers, the prediction of the severity of reactions has not yet been possible
with laboratory tests. The present study was designed to evaluate parameters
that might be clinically useful in estimation of systemic reactivity, and
parameters that could differentiate allergic beekeepers from sensitized subjects.
Thirty-two beekeepers were selected and placed into the following three groups:
anergic (n = 10), asymptomatic sensitized (n = 11), and allergic (n = 11).
Peripheral blood lymphocyte subpopulations, venom-specific immunoglobulin
(Ig) E and IgG and cytokine release by peripheral blood mononuclear cells
were measured. The ratio of stimulated interleukin-4 to stimulated interferon-gamma
was significantly higher in sensitized beekeepers than in allergic or anergic
subjects. Venom-specific IgG correlated significantly with the number of annual
stings (r = 0.575) and the years spent in beekeeping (r = 0.471). No significant
differences in the subpopulations of peripheral blood lymphocytes were found
between the study groups. We conclude that differences in the subpopulations
of peripheral blood lymphocytes are not associated with sensitization or systemic
reactivity. In asymptomatic sensitized beekeepers, T helper 2 T-cell dominance
is more pronounced than in allergic subjects. Bee venom specific IgG correlates
directly with the degree of exposure to bee venom.
PMID: 9615305 [PubMed - indexed for MEDLINE]
| 54: Izv Akad Nauk Ser Biol. 1998 Mar-Apr;(2):225-9. |
[Proliferation and
death of thymocytes caused by phospholipase A2 activator--melittin]
[Article in Russian]
Shaposhnikova
VV, Egorova
MV, Levitman
MKh, Kudriavtsev
AA, Sukharev
VI, Korystov
IuK.
Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences,
Pushchino, Russia.
The effects of mellitin, a component of bee venom activating phospholipase
A2, on proliferation and death of the rat thymocytes were studied in a wide
concentration range. Cell proliferation was estimated by the accumulation
of colchicine metaphases, Necrosis was estimated by cell lysis and Trypan
blue staining. Apoptosis was estimated by the type of DNA fragmentation, amount
of fragmented DNA, and percentage of cells with hypodiploid DNA set. Low concentrations
of mellitin (below 5 micrograms/ml) stimulated proliferation. At higher mellitin
concentrations, the thymocytes die by the primary necrosis type. Mellitin
did not induce apoptosis in the thymocytes within the concentration range
used: on the contrary, at high concentrations, it inhibited apoptosis of the
thymocytes in the control and after irradiation. Actinomycin D, inhibitor
of RNA synthesis, exerted no effect on the thymocyte death in the presence
of mellitin. It has been concluded that activation of phospholipase A2 may
induce necrosis, rather than apoptosis, and consequently, activation of phospholipase
A2 is not a necessary step in the signalling cascade that initiated apoptosis
in the thymocytes.
PMID: 9609959 [PubMed - indexed for MEDLINE]
| 55: Allergy. 1998 Mar;53(3):233-40. |
Systemic T-cell
unresponsiveness during rush bee-venom immunotherapy.
Segura
JA, Assenmacher
M, Irsch
J, Hunzelmann
N, Radbruch
A.
Institute for Genetics, University of Cologne, Germany.
By rush bee-venom immunotherapy, subjects reacting allergically to the venom
can be effectively anergized, although the mechanism of action is not known.
Here we analyzed the systemic effects of rush desensitization on the T cells
of allergic patients. In most patients, we found reduced frequencies of T
cells recalled to express CD69 and the cytokines interleukin (IL)-4 and interferon-gamma
(IFN-gamma) after stimulation of peripheral blood mononuclear cells with phorbol
12-myristate 13-acetate (PMA) and ionomycin, as compared with normal donors.
These frequencies are progressively reduced during immunotherapy. The frequency
of cells expressing IL-2 does not change. A few patients show a different
response to immunotherapy: frequencies of cells expressing CD69, IL-4, or
IFN-gamma do not change, and remain similar to those of normal donors. However,
the frequency of cells able to express IL-2 is increased. The analysis of
cytokine expression in CD45RO+ vs CD45RO- T-cell populations revealed differences
between normal and allergic donors. In allergic patients, higher frequencies
of IL-4- and IFN-gamma-expressing cells among the CD45RO- subpopulation were
found than in normal donors. This situation is not modified by immunotherapy.
The results reveal a certain degree of heterogeneity in the response of allergic
patients to bee-venom rush immunotherapy; however, all are clearly differentiated
from normal controls as judged by cytokine expression of CD45RO- T cells.
In most allergic patients, a considerable percentage of Th cells become unresponsive
to mitogenic stimulation, and may be responsible for the desensitization itself.
PMID: 9542602 [PubMed - indexed for MEDLINE]
| 56: Eur J Immunol. 1998 Mar;28(3):914-25. |
Differential
regulation of human T cell cytokine patterns and IgE and IgG4 responses by
conformational antigen variants.
Akdis
CA, Blesken
T, Wymann
D, Akdis
M, Blaser
K.
Swiss Institute of Allergy and Asthma Research, Davos, Switzerland. akdisac@siaf.unizh.ch
Bee venom phospholipase A2 (PLA) represents the major allergen and antigen
in allergic and non-allergic individuals sensitized to bee sting. We have
studied specific activation of peripheral T cells by different structural
and conformational variants of PLA and secretion of cytokines regulating IgE
and IgG4 antibody (Ab) formation. PLA molecules expressing the correctly folded
tertiary structure, which show high affinity to membrane phospholipids and
were recognized by Ab from bee sting allergic patients, induced high IL-4,
IL-5 and IL-13 production in peripheral blood mononuclear cell cultures. In
contrast, non-refolded recombinant PLA (rPLA) and reduced and alkylated native
PLA (nPLA) induced more IFN-gamma and IL-2 and higher proliferative responses.
Differences in proliferation and cytokine patterns among correctly folded
and non-refolded PLA resulted from conformation-dependent involvement of different
antigen-presenting cell (APC) types. Antigen (Ag)-presenting B cells recognized
PLA only in its natural conformation, stimulated Th2 type cytokines and induced
IgE Ab. Non-refolded PLA was recognized, processed and presented exclusively
by monocytes and induced a Th1 dominant cytokine profile leading to IgG4 production
by B cells. The possibility that production of particular cytokine patterns
and Ig isotype was influenced by the enzymatic activity of PLA was excluded
by using enzymatically inactive H34Q point-mutated, refolded rPLA. These findings
demonstrate the decisive role of specific Ag recognition by different APC,
depending on structural features, membrane phospholipid binding property and
the existence of conformational B cell epitopes, in the differential regulation
of memory IgE and IgG4 Ab. Furthermore, they show that a change from IgE-mediated
allergy to normal immunity against a major allergen can be induced by rPLA
variants that are not recognized by specific Ab and B cells but still carry
the T cell epitopes. These features may enable new applications for safer
immunotherapy.
PMID: 9541587 [PubMed - indexed for MEDLINE]
| 57: Adv Exp Med Biol. 1997;400A:365-73. |
PX-
Franson
RC, Rosenthal
MD.
Department of Biochemistry & Molecular Biophysics, Virginia Commonwealth
University, Richmond, USA.
Previously we reported that PGBx, a prostaglandin oligomer with anti-inflammatory
activity, inhibited 14 kDa phospholipase A2 (PLA2) activity and blocked arachidonic
acid mobilization in prelabeled human neutrophils (Biochim. Biophys. Acta
1006:272-277, 278-286, 1989) This study describes a new inhibitor of phospholipase
A2, PX-52, that also blocks agonist induced arachidonic acid mobilization
in prelabeled cells. PX-
| 58: Immunotechnology. 1995 Aug;1(2):115-25. |
Isolation and characterization
of allergen-binding cells from normal and allergic donors.
Irsch
J, Hunzelmann
N, Tesch
H, Merk
H, Maggi
E, Ruffilli
A, Radbruch
A.
Institute for Genetics, University of Cologne, Germany.
BACKGROUND: Flow cytometry of the immune system so far has been limited to
the analysis of subpopulations according to lineage markers. The cells involved
in a particular immune response could not be assayed due to their low frequency.
Here we show the potential of antigen-specific high gradient magnetic cell
sorting to enrich cells for visualisation in multiparameter cytometry, functional
studies and immortalization. OBJECTIVES: The aim of this study was the development
of an efficient technology for staining and isolation of antigen-binding cells
from human peripheral blood. In particular, allergen-specific cells from normal
and allergic donors should be analysed and compared to develop a cellular
diagnosis of allergy. STUDY DESIGN: The rare antigen-specific cells were sorted
by high-gradient magnetic cell sorting with MACS. Haptenized phospholipase
A2 (PLA2), the major allergen of bee venom, or haptenized ParoI, the major
allergenic component of Parietaria officinalis, were used as antigens. The
cells from normal and allergic donors, binding to the allergen were characterized
phenotypically by immuno-fluorescence. Allergen-specific B-cells were immortalized
by EBV transformation. RESULTS AND CONCLUSION: Allergen-specific cells can
be enriched from blood of both allergic and normal donors to purities of up
to 75%, by high gradient magnetic cell sorting. The specificity of labelling
with allergen was confirmed by establishing allergen-specific EBV-transformed
B-cell lines from the sorted cells. Clear differences exist in the cellular
composition of allergen-binding cells from normal compared to allergic donors.
In normal donors the allergen-binding cells are B-cells expressing CD19 and
CD21. In allergic donors, in addition to allergen-binding B-cells, occurring
in about equal absolute numbers as in normal donors, basophilic granulocytes
are labeled by allergen. These cells express CD38, CD9 and CD25 on their surface,
and stain for IgE.
PMID: 9373340 [PubMed - indexed for MEDLINE]
| 59: Eur J Immunol. 1997 Sep;27(9):2351-7. |
Glucocorticoids
inhibit human antigen-specific and enhance total IgE and IgG4 production due
to differential effects on T and B cells in vitro.
Akdis
CA, Blesken
T, Akdis
M, Alkan
SS, Heusser
CH, Blaser
K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
akdisac@siaf.unizh.ch
Although anti-inflammatory properties of glucocorticoids (GC) are well documented,
their activity in allergic diseases is still controversial. Recently, it has
been reported that GC can increase, both in vivo and in vitro, the polyclonal
production of total IgE. In this study we investigated the effects of GC on
the antigen (Ag)-specific IgE response in a human in vitro system with peripheral
blood mononuclear cells or B cells of bee venom-sensitized individuals that
allows the production of bee venom phospholipase A2 (PLA)-specific IgE and
IgG4 antibodies (Ab). PLA-specific Ab were induced by simultaneously activating
T cells and B cells specifically with allergen and polyclonally with anti-CD2
and soluble CD40 ligand (sCD40L) in the presence of interleukin (IL)-4. Indeed,
dexamethasone and prednisolone enhanced the formation of total IgE and IgG4
in PBMC, while the production of PLA-specific IgE and IgG4 Ab was selectively
inhibited in a dose-dependent manner. The suppressive effect of GC was mediated
during Ag-specific stimulation and T cell-B cell interaction. This was due
to GC suppressing specific T cell proliferation and cytokine production, whereas
neither allergen-specific nor total IgE and IgG4 production by sCD40L/IL-4-stimulated
pure B cells was affected. In contrast to GC, cyclosporine A inhibited both
total and PLA-specific IgE and IgG4 secretion in peripheral blood mononuclear
cells and B cell cultures. Further experiments showed that increase in nonspecific
total isotype response resulted from inhibition of IL-4 uptake by cells other
than B cells and sufficient availability of IL-4 to B cells for isotype switch
and synthesis. Furthermore, demonstration of opposite regulatory effects of
GC on specific and total isotype formation in vitro, including the inhibition
of allergy-relevant Ag-specific IgE response, may contribute to a better understanding
of apparently controversial observations, and explain why most allergic patients
benefit from GC therapy.
PMID: 9341780 [PubMed - indexed for MEDLINE]
| 60: J Investig Allergol Clin Immunol. 1997 Jul-Aug;7(4):217-24. |
Mononuclear blood
cell sulfidoleukotriene generation in the presence of interleukin-3 and whole
blood histamine release in honey bee and yellow jacket venom allergy.
Maly
FE, Marti-Wyss
S, Blumer
S, Cuhat-Stark
I, Wuthrich
B.
Institute of Clinical Chemistry, University Hospital Zurich, Switzerland.
Cross-linking IgE on basophils is known to cause both sulfidoleukotriene (sLT)
generation and histamine release. We recently developed an ELISA to determine
sulfidoleukotriene generation by blood mononuclear cells which employs pretreatment
with IL-3 to enhance leukotriene generation (cellular antigen stimulation
test, CAST). Here, we compared the CAST and whole blood histamine release
in response to honey bee/yellow jacket venom (BV/YJV) in 23 patients clinically
suspected of type-I allergy to these venoms. Of these, 17 were diagnosed as
"definitive venom allergics," defined by a positive skin test at
100 ng/ml of venom or less. The six in whom such skin reactivity was absent
were labelled "suspected venom allergics." Both venoms stimulated
sulfidoleukotriene generation and histamine release also from control individuals
(n = 10). In patients, insect venoms generally stimulated histamine release
and sulfidoleukotriene generation in excess of the mean + 3 SD of values obtained
with control individuals. However, about half of the patients reacted predominantly
with either histamine release or sulfidoleukotriene generation. No overall
correlation was found between threshold doses necessary to stimulate sulfidoleukotriene
generation (ThsLT) and histamine release (ThHist). (Linear correlation coefficients
between ThsLT and ThHist were -0.02 for honey bee venom and 0.13 for yellow
jacket, n = 23). Both findings are in contrast to the concept that these responses
occur in parallel. From results with "definitive venom allergics,"
CAST sensitivity was calculated as 100% for honey bee venom and 83% for yellow
jacket, and that of the histamine release assay as 62.5% for honey bee venom
and 50% for yellow jacket. Specificity of the CAST was calculated as 77% for
honey bee venom and 100% for yellow jacket, and that of the histamine release
assay as 44% for honey bee venom and 60% for yellow jacket. Thus, CAST results
are closer to skin test results than to those of the whole blood histamine
release assay.
PMID: 9330184 [PubMed - indexed for MEDLINE]
| 61: Naunyn Schmiedebergs Arch Pharmacol. 1997 Aug;356(2):233-9. |
Pharmacological
properties of Ca2+-activated K+ currents of ramified murine brain macrophages.
Eder
C, Klee
R, Heinemann
U.
Abteilung Neurophysiologie, Institut fur Physiologie der Charite, Humboldt
Universitat, Berlin, Germany.
Using the whole-cell configuration of the patch clamp technique, calcium-activated
potassium currents (I(K,Ca)) were investigated in ramified murine brain macrophages.
In order to induce I(K,Ca) the intracellular concentration of nominal free
Ca2+ was adjusted to 1 microM. The Ca2+-activated K+ current of brain macrophages
did not show any voltage dependence at test potentials between -120 and +30
mV. A tenfold change in extracellular K+ concentration shifted the reversal
potential of I(K,Ca) by 51 mV. The bee venom toxin apamin applied at concentrations
of up to 1 microM did not affect I(K,Ca). Ca2+-activated K+ currents of ramified
brain macrophages were highly sensitive to extracellularly applied charybdotoxin
(CTX). The half-maximal effective concentration of CTX was calculated to be
4.3 nM. In contrast to CTX, the scorpion toxin kaliotoxin did not inhibit
I(K,Ca) at concentrations between 1 and 50 nM. Tetraethylammonium (TEA) blocked
8.0% of I(K,Ca) at a concentration of
| 62: J Allergy Clin Immunol. 1997 Jul;100(1):96-103. |
Modulation
of T-cell response to phospholipase A2 and phospholipase A2-derived peptides
by conventional bee venom immunotherapy.
Kammerer
R, Chvatchko
Y, Kettner
A, Dufour
N, Corradin
G, Spertini
F.
Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois,
Lausanne, Switzerland.
BACKGROUND: Immunologic mechanisms of desensitization are still incompletely
understood. Safer methods of immunotherapy with reduced risks of anaphylaxis
need to be developed. OBJECTIVE: To study the effects of conventional venom
immunotherapy (VIT) on phospholipase A2(PLA2)-specific T cells and on T-cell
reactivity to short and long synthetic peptides that map the PLA2 molecule.
METHOD: Proliferation of a CD4+ cell-enriched peripheral blood mononuclear
cell fraction and cytokine secretion by T cell lines from patients hypersensitive
to bee venom and undergoing VIT in response to PLA2 and PLA2 synthetic peptides
were measured. RESULTS: T-cell proliferation in response to three synthetic
peptides, 40 to 60 amino acids long and mapping the entire PLA2 molecule with
an overlap of 10 residues (1 to 59, 51 to 99, and 90 to 134) steadily increased
during the first 14 weeks of VIT corresponding to the treatment period with
incremental doses of antigen. These results are in contrast to the low proliferation
indices obtained with short (15 amino acid-long) peptides, and the inability
to characterize the immunodominant region of the molecule with short peptides.
At the end of VIT (after 3 to 5 years), there was correspondingly, a marked
decrease in T cell responsiveness to PLA2 and to its long synthetic peptides.
This response was paralleled by a shift in the pattern of cytokine secretion
by T cell lines from a T(H0)-type to a T(H1)-type pattern. CONCLUSION: After
a transient increase in T-cell proliferation, late VIT was characterized by
T-cell hyporesponsiveness to allergen and by modulation of cytokine secretion
from a T(H0)-type to a T(H1)-type pattern. Because of their capacity to recruit
multiple T-cell epitopes, long peptides mapping the entire PLA2 molecule appear
to be efficient T cell stimulators and may represent potential candidates
for peptide immunotherapy.
PMID: 9257793 [PubMed - indexed for MEDLINE]
| 63: J Pharmacol Exp Ther. 1997 Jul;282(1):123-31. |
Variabilin:
a dual inhibitor of human secretory and cytosolic phospholipase A2 with anti-inflammatory
activity.
Escrig V, Ubeda A, Ferrandiz ML,
Darias J, Sanchez JM, Alcaraz MJ, Paya M.
Department of Pharmacology, University
of Valencia and Institute of Natural Products and Agrobiology, Tenerife, Spain.
The marine product variabilin was identified as a novel inhibitor of phospholipase
A2 (PLA2), which exhibited IC50 values of 6.9 microM and 7.9 microM for human
synovial secretory PLA2 and U937 cells cytosolic PLA2 activities, respectively.
This compound was less potent on bee venom or zymosan-injected rat air pouch
enzymes and failed to affect Naja naja venom PLA2. The production of leukotriene
B4 by human neutrophils stimulated with calcium ionophore A23187 was also
inhibited by variabilin, which was without effect on 5-lipoxygenase, cyclo-oxygenase
1 and cyclo-oxygenase 2 activities in cell-free assays. Other functions of
human neutrophils, such as degranulation and superoxide generation, were also
significantly reduced in vitro. Variabilin administered topically suppressed
the mouse ear edema induced by 12-O-tetradecanoylphorbol 13-acetate, whereas
the ear edema induced by arachidonic acid was unaffected; this suggests an
action previous to arachidonic acid metabolism. This compound administered
p.o. at 30 mg/kg and 45 mg/kg significantly inhibited mouse paw edema induced
by carrageenan and, at 0.01 to 1.0 micromol/pouch in the mouse air pouch injected
with zymosan, exerted a marked inhibition on PGE2 and leukotriene B4 levels
in exudates (ID50 values of approximately 0.028-0.029 micromol/pouch), without
affecting cell migration. Our results indicate that variabilin is an inhibitor
of human secretory and cytosolic PLA2 activities that controls eicosanoid
production in vitro and in vivo, inhibits neutrophil degranulation and superoxide
generation in vitro and shows anti-inflammatory activity after topical or
p.o. administration to mice.
PMID: 9223548 [PubMed - indexed for MEDLINE]
| 64: Int Arch Allergy Immunol. 1997 Mar;112(3):226-30. |
The murine (H-2k)
T-cell epitopes of bee venom phospholipase A2 Lie outside the active site
of the enzyme. Implications with respect to a paracrine activation of Th2
cells for an IgE antibody response.
Specht
C, Kolsch
E.
Institute for Immunology, University of Munster, Germany.
Recombinant, enzymatic active phospholipase A2 from bee venom (PLA2) is a
potent inducer of IgE antibody formation in CBA/J (H-2k) mice. In contrast,
a recombinant mutant protein lacking enzymatic activity due to an amino acid
exchange in the active site of the enzyme fails to induce IgE antibodies under
identical immunization conditions. Peptide mapping and T-cell stimulation
experiments with 18-mer overlapping peptides locate the T-helper cell-activating
epitopes in the C-terminal region of the PLA2 protein. No T-cell epitopes
are found in the area around position 34, the center of the enzymatically
active site. The data support a model in which initially an enzymatic activation
of mast cells or basophiles leads to IL-4 production which in a paracrine
way drives T-helper cells, concomitantly activated by antigen, into Th2 differentiation.
This ultimately favors B-cell activation for an IgE response.
PMID: 9066507 [PubMed - indexed for MEDLINE]
| 65: J Allergy Clin Immunol. 1997 Mar;99(3):345-53. |
Induction
and differential regulation of bee venom phospholipase A2-specific human IgE
and IgG4 antibodies in vitro requires allergen-specific and nonspecific activation
of T and B cells.
Akdis
CA, Blesken
T, Akdis
M, Alkan
SS, Wuthrich
B, Heusser
CH, Blaser
K.
Swiss Institute of Allergy and Asthma Research, Davos.
Investigations on the mechanisms of IgE regulation in vitro have been conducted
thus far in systems that allow the synthesis of total rather than specific
IgE. To study the regulatory prerequisites of antigen-specific IgE antibody
production, we have established a culture system that allows the generation
of bee venom phospholipase A2-specific IgE and IgG4 antibodies. Allergen-specific
IgE was induced by simultaneously activating T cells and B cells specifically
with allergen and polyclonally with anti-CD2 and soluble CD40 ligand in the
presence of IL-4. Additional stimulation of T cells through the CD2 activation
pathway by two different anti-CD2 monoclonal antibodies enhanced both the
allergen-specific and the total IgE and IgG4 responses. An optimal amount
of allergen (0.1 ng/ml) resulted in the induction of both allergen-specific
IgE and IgG4 antibodies. Higher antigen doses reduced allergen-specific antibodies
and enhanced total isotype production. This differential regulation of allergen-specific
and total isotypes reflects different allergen dose-dependent mechanisms in
specific and polyclonal activation of T and B cells. Although both isotypes
require IL-4 for initial induction, opposite regulatory effects by T cells
were observed for IgE and IgG4 antibody expression. In peripheral blood mononuclear
cell cultures stimulated with soluble CD40 ligand, IL-4, and phospholipase
A2, stimulation of T cells with higher amounts of anti-CD2 enhanced IgG4 in
parallel to increased IL-2 and interferon-gamma secretion but inhibited IgE
synthesis. These results provide evidence for differential regulation of allergen-specific
and total IgE and IgG4 by antigen concentration and demonstrate the pivotal
role of T cells controlling the synthesis of the IgE and IgG4 antibody isotypes.
PMID: 9058690 [PubMed - indexed for MEDLINE]
| 66: Eur J Immunol. 1997 Feb;27(2):515-21. |
The intensity of T cell receptor
engagement determines the cytokine pattern of human allergen-specific T helper
cells.
Carballido JM,
Faith A, Carballido-Perrig N,
Blaser K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos.
Enhanced production of T helper (Th)2 cytokines by allergen-specific Th cells
plays a major role in the induction and maintenance of IgE-mediated allergic
disorders. The mechanism that triggers this type of response in atopic individuals
is not fully understood. Allergen-specific human Th cell clones produce interleukin
(IL)-4 and low or undetectable levels of interferon (IFN)-gamma after stimulation
with low concentrations of antigen. However, these Th cell clones are capable
of generating significant amounts of IFN-gamma after optimal activation through
their T cell receptor (TcR). Allergen-specific Th cell clones isolated from
allergic individuals required higher doses of antigen to reach the plateau
of proliferation and to generate Th0 cytokine responses than their counterparts
isolated from nonallergic subjects. On the other hand, if allergen was replaced
by anti-CD3 monoclonal antibody (mAb), both allergic and nonallergic Th cell
clones attained the highest level of proliferation and significant IFN-gamma
production in response to equivalent concentrations of anti-CD3 mAb. These
results indicate that the strength of T cell ligation, which can be modulated
by the availability of the TcR ligand, controls the balance of Thl/Th2 cytokines
produced by memory Th cells in vitro. In the particular case of bee venom
phospholipase A2, it is shown that the expression of allergen-specific surface
Ig on antigen-presenting B cells has little influence on antigen uptake and
therefore in determining the levels of T cell activation and cytokine production.
Alternatively, the affinity of particular major histocompatibility complex
class II molecules on antigen-presenting cells for allergen-derived peptides
might determine the amount of specific ligand presented to the Th cells and
play a decisive role skewing the Th cell cytokine production towards Th1 or
Th2 phenotypes. These findings, which are consistent with the changes in cytokine
patterns observed following clinical hyposensitization, suggest that polarized
human Th2 cell subsets still retain the capacity to modulate their cytokine
pattern.
PMID: 9045925 [PubMed - indexed for MEDLINE]
| 67: J Immunol Methods. 1996 Dec 15;199(2):119-26. |
Quantitation of the cytosolic
phospholipase A2 (type IV) in isolated human peripheral blood eosinophils
by sandwich-ELISA.
Zhu X, Munoz NM, Rubio N, Herrnreiter A,
Mayer D, Douglas I, Leff AR.
Department of Medicine, University of Chicago, IL 60637, USA.
Sandwich enzyme-linked immunosorbent assay (sELISA) was developed for precise
quantitation of cytosolic phospholipase A2 (cPLA2 type IV) concentration in
isolated human peripheral blood eosinophils as an alternative to semiquantitative
chemiluminescent assay employing immunoprecipitation/Western blot analysis.
In this assay, monoclonal mouse anti-human cPLA2 antiserum was used as the
capture antibody, polyclonal rabbit anti-human cPLA2 antiserum as the secondary
antibody, and alkaline phosphatase-conjugated goat anti-rabbit IgG as the
tertiary, reporter antibody. Purified human cPLA2 (0-1000 ng/ml) dissolved
in Tris-HCl buffered saline was used as the standard protein. The detection
limit for cPLA2 in 10(6) eosinophils was 0.109 ng/ml, and coefficients of
inter- and intra-assay variation were 4.23% and 7.07%, respectively. There
was no cross-reactivity with other (secretory) isoforms of PLA2 (sPLA2 types
I-III) either from porcine pancreas, human synovial fluid, or bee venom. In
separate studies, the recovery of cPLA2 was > 83% when eosinophil lysate
was supplemented exogenously with two different concentrations of cPLA2. From
a total protein content of 22.3 +/- 1.7 micrograms/10(6) cells, the baseline
concentration of cPLA2 was 0.38 +/- 0.18 ng/10(6) cells in eosinophils obtained
from mildly atopic donors. Immunoblotting studies confirmed the complete specificity
for the type IV isoform as detected by sELISA. This sELISA method permits
the precise quantitative assessment of cPLA2 in nanogram quantities per million
cells, which has not previously been possible by immunoblotting analysis.
PMID: 8982353 [PubMed - indexed for MEDLINE]
| 68: Eur J Immunol. 1996 Dec;26(12):2972-80. |
Potential allergens stimulate
the release of mediators of the allergic response from cells of mast cell
lineage in the absence of sensitization with antigen-specific IgE.
Machado DC, Horton D, Harrop R, Peachell PT,
Helm BA.
Krebs Institute for Biomolecular Research, Department of Molecular Biology
and Biotechnology, University of Sheffield, GB.
A number of structurally diverse antigens preferentially stimulate the synthesis
of IgE antibodies, but no unifying principle has been proposed that explains
the nature of isotype selection. In the present study, we show that common
allergens present in bee venom, house dust mite emanations and parasite proteins
induce mast cell and basophil degranulation and stimulate interleukin-4 synthesis,
and secretion in the absence of antigen-specific IgE. These data point to
a linkage between the initial activation of cells of the innate immune system
and subsequent adaptive immune responses. They suggest that IgE-independent
mast cell and basophil degranulation is predictive of potential allergenicity
and can be evaluated by means of a cellular assay. Our study indicates that
non-immunological degranulation by prototypic allergens, such as bee venom
phospholipase A2 or proteases associated with house dust mite emanations,
is critically dependent on enzymatic activity. These findings have potentially
important implications for vaccine design in allergic and parasitic disease.
PMID: 8977293 [PubMed - indexed for MEDLINE]
| 69: Naunyn Schmiedebergs Arch Pharmacol. 1996 Nov;354(5):677-83. |
Inhibition of phospholipase
A2 activities and some inflammatory responses by the marine product ircinin.
Cholbi R, Ferrandiz ML,
Terencio MC,
De Rosa S, Alcaraz MJ, Paya M.
Department of Pharmacology, University of Valencia, Faculty of Pharmacy, Burjassot,
Spain.
The marine product ircinin has been tested for its effects on secretory and
cytosolic phospholipase A2 (PLA2) activities in vitro as well as for inhibition
of cellular functions in human neutrophils and inflammatory responses in mice.
Ircinin inhibited Naja naja venom, human synovial recombinant, bee venom and
zymosan-injected rat air pouch PLA2 with IC50 values in the microM range,
similar to those of the known inhibitor scalaradial. On the other hand, ircinin
was less active on cytosolic PLA2 from human monocytes and decreased potently
the release of LTB4 in human neutrophils. This marine product affected weakly
human neutrophil functions like superoxide generation and degranulation. In
the zymosan-injected rat air pouch ircinin inhibited in vivo the activity
of PLA2 present in exudates and reduced dose-dependently myeloperoxidase levels,
whereas cell migration was inhibited only at the highest dose tested. This
compound exerted a potent anti-oedematous effect after topical application
in the mouse ear oedema test. Ircinin is a new inhibitor of PLA2 activity
and our results suggest a potential role for this marine product as an inhibitor
of inflammatory processes.
PMID: 8938669 [PubMed - indexed for MEDLINE]
| 70: Clin Exp Allergy. 1996 Oct;26(10):1112-8. |
Comment in:
· Clin Exp Allergy. 1996 Oct;26(10):1101-4.
Influence of bee venom immunotherapy
on degranulation and leukotriene generation in human blood basophils.
Jutel M, Muller UR, Fricker M, Rihs S, Pichler WJ, Dahinden C.
Medical Division, Zieglerspital, Bern, Switzerland.
BACKGROUND: Rapid clinical tolerance can be induced over several hours by
very fast bee venom immunotherapy (VIT) protocols. OBJECTIVE: To investigate
the mechanisms underlying VIT we examined the changes of blood basophil responsiveness
during VIT. METHODS: Seven bee venom allergic patients with a history of severe
systemic reactions after a bee sting were investigated. A cumulative dose
of 111.1 micrograms bee venom (BV) was administered sc over 3.5 h under intensive
care conditions according to an ultra-rush protocol. The release of histamine
and the formation of leukotrienes in response to BV, major BV allergen Phospholipase
A2 (PLA), IgE receptor cross-linking with the use of monoclonal antibodies
against IgE and IgE receptor, as well as IgE independent activation in response
to C5a were determined in vitro before and after ultra-rush VIT. RESULTS:
We demonstrated a decrease of total histamine in peripheral blood leucocytes
just after VIT. Histamine release in response to all the stimuli used is not
affected by ultra-rush VIT, if expressed as per cent release of total histamine.
However, the absolute amount product released in response to stimulation was
decreased, particularly with allergen (BV, PLA). We also found a significant
reduction of LTC4 formation after VIT in samples stimulated with specific
allergen (BV, PLA). CONCLUSION: Blood basophils are a target for VIT, which
induces impaired release of both preformed and newly generated mediators.
However, we believe the basic mechanisms of rapid clinical tolerance induced
by ultra-rush VIT remain to be investigated.
PMID: 8911695 [PubMed - indexed for MEDLINE]
| 71: J Clin Invest. 1996 Oct 1;98(7):1676-83. |
Epitope-specific T cell
tolerance to phospholipase A2 in bee venom immunotherapy and recovery by IL-2
and IL-
Akdis CA, Akdis M, Blesken T, Wymann D, Alkan SS, Muller U, Blaser K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
Bee venom phospholipase A2 (PLA) is the major allergen in bee sting allergy.
It displays three peptide and a glycopeptide T cell epitopes, which are recognized
by both allergic and non-allergic bee venom sensitized subjects. In this study
PLA- and PLA epitope-specific T cell and cytokine responses in PBMC of bee
sting allergic patients were investigated before and after 2 mo of rush immunotherapy
with whole bee venom. After successful immunotherapy, PLA and T cell epitope
peptide-specific T cell proliferation was suppressed. In addition the PLA-
and peptide-induced secretion of type 2 (IL-4, IL-5, and IL-13), as well as
type 1 (IL-2 and IFN-gamma) cytokines were abolished, whereas tetanus toxoid-induced
cytokine production and proliferation remained unchanged. By culturing PBMC
with Ag in the presence of IL-2 or IL-15 the specifically tolerized T cell
response could be restored with respect to specific proliferation and secretion
of the type 1 T cell cytokines, IL-2 and IFN-gamma. In contrast, IL-4, IL-5,
and IL-13 remained suppressed. Treatment of tolerized T cells with IL-4 only
partially restored proliferation and induced formation of distinct type 2
cytokine pattern. In spite of the allergen-specific tolerance in T cells,
in vitro produced anti-PLA IgE and IgG4 Ab and their corresponding serum levels
slightly increased during immunotherapy, while the PLA-specific IgE/IgG4 ratio
changed in favor of IgG4. These findings indicate that bee venom immunotherapy
induces a state of peripheral tolerance in allergen-specific T cells, but
not in specific B cells. The state of T cell tolerance and cytokine pattern
can be in vitro modulated by the cytokines IL-2, IL-4, and IL-15, suggesting
the importance of microenvironmental cytokines leading to success or failure
in immunotherapy.
PMID: 8833918 [PubMed - indexed for MEDLINE]
| 72: Pediatr Allergy Immunol. 1996 Aug;7(3):109-16. |
Fetal peripheral blood mononuclear
cell proliferative responses to mitogenic and allergenic stimuli during gestation.
Jones AC, Miles EA, Warner JO, Colwell BM, Bryant TN, Warner JA.
Department of Child Health, University of Southampton, England.
Blood samples were obtained from fetuses and premature babies (n = 51) (15-34
weeks gestation) to determine at what stage the fetal immune system was able
to produce a positive proliferative response to common allergens. Peripheral
blood mononuclear cells (PBMC) were stimulated with the mitogen, phytohaemagglutinin
(PHA), and the allergens, house dust mite, cat fur, birch tree pollen, beta-lactoglobulin,
ovalbumin and bee venom (mellitin). Results were expressed as ratios of stimulated
to unstimulated 3H thymidine incorporation, and as percent positive responders.
There was an increase in proliferation ratio which correlated with increasing
gestational age for PHA (p < 0.0001), cat fur (p = 0.042), birch pollen
(p = 0.022) and beta-lactoglobulin (p = 0.006). The point in gestation when
cells from some individuals began responding to the allergens with a ratio
of 2.0 was at approximately 22 weeks. PBMC proliferative response ratios were
higher from samples from babies > 22 weeks gestation compared to < 22
weeks for the mitogen and all allergens, except mellitin. There was also a
greater proportion of positive responders from samples > 22 weeks compared
to < 22 weeks for the mitogen and all allergens, except mellitin. Maternal
exposure to birch pollen, which has a discrete season, was assessed to determine
whether exposure had occurred at 22 weeks gestation or beyond. Results showed
a higher proliferative response in infant cells stimulated with birch pollen
(p = 0.005) and higher proportion of positive responders (p = 0.01) in the
group of babies whose mothers had been exposed to birch pollen beyond 22 weeks,
compared to those whose mothers had not been so exposed. These results suggest
that in utero fetal exposure to an allergen from around 22 weeks gestation
may result in primary sensitisation to that allergen, leading to positive
proliferative responses, at birth.
PMID: 9116874 [PubMed - indexed for MEDLINE]
| 73: Eur J Immunol. 1996 Jun;26(6):1260-5. |
Kinetics and functional implications
of Th1 and Th2 cytokine production following activation of peripheral blood
mononuclear cells in primary culture.
McHugh S, Deighton J, Rifkin I, Ewan P.
Molecular Immunopathology Unit, Medical Research Council Centre, Cambridge,
GB.
The importance of cytokine production in some disease processes is now widely
recognized. To investigate temporal relationships between cytokines, we stimulated
peripheral blood mononuclear cells (PBMC) in vitro using the T cell mitogen
phytohemagglutinin (PHA) and various antigens chosen to induce predominantly
Th1 (streptokinase: streptodornase or purified protein derivative) or Th2
(Dermatophagoides pteronyssinus, bee or wasp venom: allergens in sensitive
subjects) responses. Cytokine production was measured by sensitive bioassays
or enzyme-linked immunosorbent assays. Of the 30 subjects studied, 10 were
normal and 20 individuals were allergic to either D. pteronyssinus (n = 10)
or bee venom (n = 10) (examined before specific allergen immunotherapy). We
examined the temporal profiles of a panel of cytokines produced in primary
culture. In PHA-driven cultures, cytokines were found to be sequentially produced
in the order interleukin (IL)-2, IL-4, IL-5, IL-3, interferon (IFN)-gamma,
IL-10, IL-6, IL-12 and tumor necrosis factor (TNF)-alpha. The response to
allergen in allergic patients was predominantly Th2 in nature, with the production
of IL-4, IL-5, IL-6 and IL-10, but little or no IFN-gamma. IL-2, IL-3, TNF-alpha
and IL-12 were also produced in low amounts. The response of both atopic and
normal subjects to recall bacterial antigens was predominantly Th1, with high
levels of IFN-gamma, IL-2 and TNF-alpha. The relevance of the order, amount
and speed of production, characteristic kinetics (production, consumption,
homeostatic regulation) and the cell source of the cytokines are discussed.
PMID: 8647202 [PubMed - indexed for MEDLINE]
| 74: J Immunol. 1996 Mar 1;156(5):1728-34. |
Biochemical characterization
of antigen-specific glycosylation-inhibiting factor from antigen-specific
suppressor T cells. I. Identification of a 55-kilodalton glycosylation-inhibiting
factor peptide with TCR alpha-chain determinant.
Nakano T, Ishii Y, Ishizaka K.
Division of Immunobiology,
Stimulation of OVA-specific suppressor T cell (Ts) hybridoma and bee venom
phospholipase A2 (PLA2)-specific Ts hybridoma with Ag-pulsed APC or by cross-linking
of CD3 resulted in the formation of Ag-specific glycosylation-inhibiting factor
(GIF). Affinity-purified Ag-specific GIF preparations, obtained by using Ag-coupled
Sepharose or anti-TCR alpha-chain-coupled Affi-Gel, contained a 55-kDa peptide,
which bound both polyclonal anti-GIF Abs and anti-TCR-alpha mAb in immunoblotting.
The same hybridomas constitutively secrete 13-kDa bioactive GIF peptide that
has no affinity for homologous Ag, but neither the Ag-specific GIF activity
nor 55-kDa GIF peptide was detectable in culture supernatants of unstimulated
cells. Northern blot analysis of mRNA from the anti-CD3-stimulated hybridoma
with 32P-labeled GIF cDNA revealed only 0.6 kb mRNA, which encodes the 13-kDa
nonspecific GIF. No mRNA of the 55-kDa GIF was detectable. A representative
OVA-specific Th hybridoma, DO 11.10 cells contain the 0.6 kb GIF mRNA and
constitutively secrete inactive GIF peptide. However, the Th hybridoma failed
to secrete the 55-kDa peptide or any peptide with the TCR-alpha determinant
upon stimulation with anti-CD3. It appears that the formation of the 55-kDa
peptide with the TCR-alpha determinant is unique for a subset of T cells including
Ts cells that form bioactive GIF.
PMID: 8596020 [PubMed - indexed for MEDLINE]
| 75: Adv Exp Med Biol. 1996;409:295-303. |
Allergen dose dependent cytokine
production regulates specific IgE and IgG antibody production.
Blaser K.
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
The elicitation of a specific immune response against allergens depends on
the recognition of antigenic determinants (epitopes) by specific T and B lymphocytes.
In order to determine the relevant epitopes for human T and B cells and their
features in the regulation and production of specific IgE and/or IgG antibodies,
we have investigated the immune response to bee venom phospholipase A2 (PLA)
in allergic and non-allergic subjects. This enzyme represents the major allergen
in bee sting allergy. It consists of 134 amino acid residues with a carbohydrate
side chain at position 13 and is available as recombinant protein. We have
developed PLA-specific T-cell clones from bee sting allergic and non-allergic
human subjects. Using a panel of dodecapeptides overlapping in 10 residues
and a large set of 18-25 mer overlapping peptides, we detected three epitopes
that were recognized by peripheral blood T-cells and T-cell clones. A fourth
determinant involved the carbohydrate moiety on Asn13 of PLA. Whereas the
CHO-depending epitope seems to be mostly active in allergics, the other three
epitopes are equally recognized by peripheral blood mononuclear cells (PBMC)
of both allergic and non-allergic individuals. In T-cell clones, the ratio
of IL-4/IFN gamma cytokines and the quality of the activating signal depend
on the strength of the binding of the MHC-II/Ag/TcR complex between APC and
T-cells. The number of antigen-specific APC-T-cell contact sites can be varied
in vitro by changing the dose of antigen added to the cell culture. While
isotype switch for both IgE and IgG4 requires IL-4, this cytokine suppresses
antigen-specific IgG4 production by already switched B-cells. Therefore, IL-4
and IFN gamma display counter-regulatory effects on the production of IgE
being responsible for atopic states and IgG4 antibodies which are signs of
a normal immune response to allergen and act as protective antibodies. The
combination of this counter-regulation of IgE and IgG4 antibodies with the
fundamental law of mass action for chemical equilibrium reactions revealed
that the antigen concentration governs to a great part the ratio of IL-4/IFN
gamma secretion and therefore the formation of IgE and IgG and allergy or
protection, together with the equilibrium constant K, which represents immunological
individuality and a measure of Ag presentation.
Publication Types:
· Review
PMID: 9095257 [PubMed - indexed for MEDLINE]
| 76: Clin Exp Allergy. 1995 Dec;25(12):1205-10. |
Ultra rush bee venom immunotherapy
does not reduce cutaneous weal responses to bee venom and codeine phosphate.
Jutel M, Skrbic D, Pichler WJ, Muller UR.
Zieglerspital Bern, Switzerland.
BACKGROUND: The rapid administration of bee venom in cumulative doses exceeding
the quantity contained in one bee sting is well tolerated by most of the patients
during 3.5 h of ultra-rush bee venom immunotherapy (VIT). The mechanism of
this tolerance is unknown. OBJECTIVE: The aim of the study was to verify the
hypothesis that either slow mediator depletion of mast cells or blockade of
their surface receptor mechanisms by increasing doses of allergen might be
the major mechanisms of tolerance induced by ultra-rush VIT. METHODS: Nine
bee venom allergic patients with a history of severe systemic reactions after
a bee sting, positive skin tests and bee venom specific serum IgE antibodies
were treated as follows: on the first day a cumulative dose of 111 micrograms
was administered over 3.5 h under intensive care conditions. Further injections
were given on day 7, day 21 and thereafter at 4 week intervals. Intradermal
tests with codeine phosphate (non-specific mast cell degranulation) and bee
venom were performed before the initiation of VIT and 30 min after the last
injection on the same day as well as before the subsequent bee venom injections.
RESULTS: No significant changes of skin reactivity to both codeine phosphate
and bee venom were observed on day 1 (before initiation of VIT and after the
last injection on the same day). CONCLUSIONS: Ultra-rush VIT does not induce
mediator depletion or surface receptor blockade in skin mast cells.
PMID: 8821301 [PubMed - indexed for MEDLINE]
| 77: Arch Biochem Biophys. 1995 Dec 1;324(1):78-84. |
Bee venom phospholipase
A2 is recognized by the macrophage mannose receptor.
Mukhopadhyay A,
Stahl P.
Department of Cell Biology and Physiology, Washington University School of
Medicine, St. Louis, Missouri 63110, USA.
A high affinity and a specific binding site for bee venom PLA2 was found on
the surface of J774E macrophages. The binding sites for bee venom PLA2 are
entirely different from the binding sites for pancreatic and snake venom PLA2
as revealed by competition experiments. Binding and uptake of bee venom PLA2
by J774E macrophages was shown to be competed by mannose-BSA, glucose-BSA,
N-acetylglucosamine-BSA, but not by galactose-BSA, indicating that the binding
of bee venom PLA2 is probably mediated by macrophage mannose receptor. An
affinity labeling experiment revealed that the bee venom PLA2 specifically
binds to a single polypeptide with a mass of approximately 180 kDa. Moreover,
the affinity labeled protein component, i.e., the binding site, was not detected
in the presence of excess mannose-BSA, suggesting that mannose-BSA and the
bee venom PLA2 bind to the same site on macrophages. These observations were
further supported by the binding of bee venom PLA2 to cells which are known
to express the mannose receptor and by specific binding of bee venom PLA2
to the purified mannose receptor. These data confirm that bee venom PLA2 binding
to macrophages is mediated through the mannose receptor.
PMID: 7503563 [PubMed - indexed for MEDLINE]
| 78: Clin Exp Allergy. 1995 Nov;25(11):1108-17. |
Selective restimulation of antigen
or allergen preactivated T cells using OKT3 F(ab)2 results in the secretion
of TH-1 or TH-2-like cytokine patterns.
Jutel M, Wyss-Coray T,
Carballido JM,
Blaser K, Muller UR, Pichler WJ.
Zieglerspital, Bern, Switzerland.
BACKGROUND: The synthesis of IgE is regulated by cytokines secreted from T-helper
cells. The studies on cytokine secretion by peripheral blood mononuclear cells
(PBMC) upon stimulation with antigen or allergen are difficult due to low
levels of cytokines, especially of interleukin-4 (IL-4). OBJECTIVE: In this
study we tried to establish a culture system, which could enable the measurement
of the cytokine profiles in specifically activated cultures. METHODS: Three
methods to potentiate cytokine secretion were evaluated: PBMC from bee venom
or house dust mite (Dermatophagoides pteronyssinus) allergic patients as well
as normal subjects were stimulated either with the major bee venom allergen
phospholipase A2 (PLA) or with the major D. pteronyssinus allergen (Der p
1) or with the control antigens tetanus toxoid (TT) and purified protein derivate
(PPD). After 7 days of culture the cells were restimulated either with plastic
bound OKT3 F(ab)2 monoclonal antibodies (MoAbs), with the appropriate antigen
+ antigen presenting cells or with IL-2. The secretion of cytokines (IL-4,
IFN gamma) was measured after restimulation of the cultures (day 8). RESULTS:
While OKT3 F(ab)2 was unable to activate resting T cells, it could restimulate
preactivated cells. Restimulation with OKT3 F(ab)2 induced higher IL-4 and
IFN gamma secretion than restimulation with IL-2 or antigen. TT and PLA stimulated
a similar cytokine secretion profile in normal and PLA allergic subjects with
substantial levels of both IL-4 and IFN gamma. In contrast, PPD induced virtually
only IFN gamma secretion. Der p 1 stimulated mainly IL-4 secretion but also
IFN gamma production in some mite allergic patients. CONCLUSION: We have established
a cell culture system, which combines antigen specificity with a strong cytokine
inducing signal provided by anti-CD3 MoAbs. TH-1 and TH-2 characteristic cytokine
patterns can be observed in short-term PBMC cultures already after 8 days
of culture.
PMID: 8581844 [PubMed - indexed for MEDLINE]
| 79: Eur J Pharmacol. 1995 Oct 24;285(3):281-8. |
Effects of marine 2-polyprenyl-1,4-hydroquinones
on phospholipase A2 activity and some inflammatory responses.
Gil B, Sanz MJ, Terencio MC,
De Giulio A,
De Rosa S, Alcaraz MJ, Paya M.
Department of Pharmacology, University of Valencia, Faculty of Pharmacy, Spain.
Three 2-polyprenyl-1,4-hydroquinone derivatives (2-heptaprenyl-1,4-hydroquinone:
IS1, 2-octaprenyl-1,4-hydroquinone: IS2 and 2-[24-hydroxy]-octaprenyl-1,4-hydroquinone:
IS3) isolated from the Mediterranean sponge Ircinia spinosula, were evaluated
for effects on phospholipase A2 activity of different origin (Naja naja venom,
human recombinant synovial fluid and bee venom), as well as on human neutrophil
function and mouse ear oedema induced by 12-O-tetradecanoylphorbol 13-acetate
(TPA). IS1 interacted minimally with these responses. In contrast, IS2 and
IS3 inhibited human recombinant synovial phospholipase A2 in a concentration-dependent
manner, with minor effects on the rest of the enzymes. Both compounds slightly
affected superoxide generation and degranulation in human neutrophils, whereas
they decreased thromboxane B2 and leukotriene B4 synthesis and release in
a mixed suspension of human platelets and neutrophils stimulated by ionophore
A23187, with IC50 values in the microM range. IS3 was the most effective inhibitor
of the synthesis of thromboxane B2 by human platelet microsomes and of leukotriene
B4 by high speed supernatants from human neutrophils. IS2 and IS3 showed topical
anti-inflammatory activity against the TPA-induced ear inflammation in mice,
with similar effects on oedema and a higher inhibition of IS3 on leukocyte
migration, estimated as myeloperoxidase activity in supernatants of ear homogenates.
Some structure-activity relationships were established since differences in
the prenylated chain attached to the hydroquinone moiety result in important
modifications of these inflammatory responses.
PMID: 8575515 [PubMed - indexed for MEDLINE]
| 80: J Immunol Methods. 1995 Oct 12;186(1):27-36. |
Preparation of soluble
recombinant T cell receptor alpha chain by using a calmodulin fusion expression
system.
Ishii Y, Nakano T, Honma N, Yuyama N, Yamada Y, Watarai H, Tomura T, Sato M, Tsumura H, Ozawa T, et al.
Division of Immunobiology,
We have isolated a full length T cell receptor alpha chain (TCR alpha) cDNA
derived from a bee venom phospholipase A2-specific mouse suppressor T cell
hybridoma. A bacterial fusion expression system was constructed using rat
calmodulin as a fusion partner for production of soluble TCR alpha. In this
system, calmodulin-TCR alpha fusion protein was expressed at a high level
in the soluble fraction of bacterial cell lysate, and could be purified by
binding of calmodulin portion of the protein to phenyl-Sepharose. Using this
system, fusion proteins containing a TCR alpha peptide corresponding to the
complete extracellular region, V alpha-J alpha region or C alpha extracellular
region were isolated. TCR alpha peptides were then released from the fusion
proteins by digestion with thrombin which recognizes a linker sequence between
calmodulin portion and TCR alpha segment. Polyclonal antibodies against constant
region of TCR alpha chain (C alpha) were obtained by immunization of rabbits
with the recombinant C alpha peptide. ELISA for TCR protein was established
by using the polyclonal antibodies and the monoclonal antibody specific for
C alpha region.
PMID: 7561145 [PubMed - indexed for MEDLINE]
| 81: Int Immunol. 1995 Oct;7(10):1649-57. |
Administration of IL-12 during
ongoing immune responses fails to permanently suppress and can even enhance
the synthesis of antigen-specific IgE.
Germann T, Guckes S, Bongartz M, Dlugonska H,
Schmitt E, Kolbe L, Kolsch E, Podlaski FJ,
Gately MK, Rude E.
Institut fur Immunologie, Mainz, Germany.
The synthesis of antibodies of the IgE isotype in mice largely depends on
IL-
PMID: 8562510 [PubMed - indexed for MEDLINE]
| 82: Clin Exp Allergy. 1995 Sep;25(9):828-38. |
Bee venom immunotherapy induces
a shift in cytokine responses from a TH-2 to a TH-1 dominant pattern: comparison
of rush and conventional immunotherapy.
McHugh SM, Deighton J, Stewart AG, Lachmann PJ,
Ewan PW.
Molecular Immunopathology Unit, Medical Research Centre, Cambridge, UK.
BACKGROUND: The mechanism of immunotherapy is unclear. Allergic disease is
known to involve enhanced TH-2 cytokine responses to allergen. OBJECTIVE:
In order to investigate the mechanisms of immunotherapy, we have examined
changes in cytokine secretion before (13 patients) and during (nine patients)
both rush and conventional venom immunotherapy (VIT) in bee venom allergic
patients. METHODS: Peripheral blood mononuclear cells were stimulated in vitro
with bee venom, non-specific antigen or mitogen and secretion of IL-4 (TH-2)
and IFN gamma (TH-1) over the culture period measured. RESULTS: Untreated
patients had TH-2 responses to venom and TH-1 responses to antigen and strong
proliferative responses to venom. Controls showed no response (proliferation
or cytokines) to venom and the normal TH-1 response to antigen. VIT resulted
in marked changes in cytokine secretion to venom, with reduction of the abnormal
TH-2 response and induction of a TH-1 response. The pattern differed in rush
and conventional VIT. One day after rush VIT there was a significant fall
in IL-4 secretion (P < 0.01), which rose by 3 weeks then declined. In conventional
VIT there was a gradual reduction of IL-4 production significant after 2 months
and undetectable by 6 months. IFN gamma secretion was induced by VIT. Proliferative
responses mirrored the IL-4 changes. One day after rush VIT there was a loss
of T cells, monocytes and NK cells from peripheral blood. CONCLUSION: This
study shows that immunotherapy shifted cytokine responses to allergen from
a TH-2 to a TH-1 dominant pattern, suggesting direct effects on T cells. How
these cytokine changes relate to clinical desensitization is not clear. In
the longer term they would result in an isotype switch from IgE to IgG. Early
changes in cytokine or chemokine production might downregulate mast cell or
basophil reactivity and explain the rapid desensitization in rush VIT.
Publication Types:
PMID: 8564721 [PubMed - indexed for MEDLINE]
| 83: J Immunol. 1995 Sep 1;155(5):2605-13. |
A link between catalytic
activity, IgE-independent mast cell activation, and allergenicity of bee venom
phospholipase A2.
Dudler T, Machado DC, Kolbe L, Annand RR, Rhodes N, Gelb MH, Koelsch K, Suter M, Helm BA.
Swiss Institute of Allergy and Asthma Research, Davos, Switzerland.
The molecular and cellular mechanisms controlling Ab isotype selection following
encounter of a given Ag are still unclear, although the regulatory role of
cytokines is established. In the present study we explored the possibility
that the nonimmunologic interaction of an allergen with cells of the innate
immune system might result in a release of mediators that promote IgE isotype
selection in adaptive responses. Using the bee venom allergen phospholipase
A2 (PLA2) and a mutant variant lacking enzymatic function, we show that PLA2,
but not its catalytically inactive variant, is able to induce IgE-independent
mediator release, including IL-4, from rodent mast cells. Assessing the in
vivo relevance of these observations, we find that repeated injections of
low doses of active enzyme into mice induce the synthesis of high levels of
PLA2-specific IgE, while immunization with the inactive form yields no detectable
IgE response. Both Ags were similarly immunogenic when high doses of Ag were
used for immunization. These findings suggest that mast cells might be a source
of IL-4 at the onset of specific immunity against sources of allergens such
as bee venom that contain PLA2 and support the concept that the biologic action
of an Ag on cells of the innate immune system can play a role in determining
adaptive immune responses.
PMID: 7544378 [PubMed - indexed for MEDLINE]
| 84: Clin Exp Allergy. 1995 Aug;25(8):704-12. |
Tryptase and histamine release
due to a sting challenge in bee venom allergic patients treated successfully
or unsuccessfully with hyposensitization.
Eberlein-Konig B,
Ullmann S, Thomas P, Przybilla B.
Dermatologische Klinik und Poliklinik, Ludwig-Maximilians-Universitat, Munich,
Germany.
BACKGROUND: Hyposensitization with been venom leads to full protection in
most, but not all patients with IgE-mediated systemic reactions to bee stings.
OBJECTIVE: To determine the relationship of clinical reactivity to the release
of mediators and to changes of antibody concentrations in the peripheral circulation
at a bee sting challenge test. METHODS: Blood was sampled before (1 min) and
at 15, 60 and 180 min after a sting challenge from 19 patients on hyposensitization.
Of these six still reacted and 13 were protected. Histamine, mast cell tryptase,
bee venom-specific IgE and IgG in the serum, and histamine release from peripheral
blood leucocytes (PBL) upon exposure to bee venom were determined. RESULTS:
Tryptase above the detection level was found only at 15 (60) min in 4/6 (1/6)
patients who reacted. After the sting challenge there was a significant increase
of the histamine levels in patients who reacted at 15 min (P < 0.05) and
in patients who did react at 60 and 180 min (P < 0.01). The total histamine
content of PBL was significantly decreased after 15 and 60 min in patients
who reacted (P < 0.01) and in those that did not (P < 0.05). Bee venom-induced
histamine release was significantly reduced in patients reacting and those
that did not at 15 min (P < 0.05), and was significantly decreased in reactors
also at 60 and 180 min (P < 0.05/0.01). Specific IgG antibodies showed
a minor decrease (P < 0.05) after the sting challenge in both groups, whereas
specific IgE did not change significantly. CONCLUSION: These results indicate
that bee venom anaphylaxis is associated with the release of mediators from
both mast cells as well as basophils. Successful hyposensitization does not
induce a state of immunological non-reactivity, but rather alters the magnitude
and the pattern of mediator release.
PMID: 7584681 [PubMed - indexed for MEDLINE]
| 85: J Immunol. 1995 Apr 15;154(8):4187-94. |
Bee venom immunotherapy
results in decrease of IL-4 and IL-5 and increase of IFN-gamma secretion in
specific allergen-stimulated T cell cultures.
Jutel M, Pichler WJ, Skrbic D, Urwyler A, Dahinden C, Muller UR.
Ziegler Hospital, Bern, Switzerland.
The mechanisms of bee venom immunotherapy (VIT) are largely unknown. The aim
of this study was to follow the changes of T cell cytokine secretion during
the course of VIT. Ten bee venom-allergic patients with a history of severe
systemic reactions, positive skin tests, and bee venom (BV)-specific serum
IgE Abs were treated as follows: on the first day, a cumulative dose of 111
micrograms, starting with 0.1 microgram, was administered s.c. under intensive
care conditions. Further injections of 100 micrograms BV were given on day
7, day 21, and thereafter at intervals of 4 wk. Blood samples were obtained
just before the initiation of VIT, after the last injection on the same day,
and before the subsequent BV injections on days 7, 21, and 50 of VIT. Peripheral
blood mononuclear cells (PBMC) were stimulated with phospholipase A (PLA),
the major BV allergen, or with a control Ag tetanus toxoid (TT). Cytokine
secretion was measured 24 h after restimulation of the cultures with solid-phase
bound OKT3 F(ab')2 mAbs after 7 days of culture. In PLA-stimulated cultures,
VIT resulted in decreased IL-4 and IL-5 and increased IFN-gamma secretion.
In TT-stimulated cultures, we observed similar levels of cytokines before
and during VIT. We conclude that ultra-rush VIT changes allergen-specific
T cell reactivity.
PMID: 7706753 [PubMed - indexed for MEDLINE]
| 86: J Immunol. 1995 Apr 15;154(8):4027-31. |
|