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Antigenic properties of allergenic proteins from oriental hornet (Vespa orientalis) venom
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Eleventh World Congress
proteinase a, b and c had estimated mol. wts of 25,000 and 24,000, respectively, as determined by SDS PAGE and a pl of 9.0, 9.3 and 8.5. Incubation of the proteinases with purified human ~2-antiplasmin at a molar ratio of 1:10 resulted in a loss of more than 90% of the initial ~t2-antiplasmin activity within 60 min. In SDS PAGE the ~2-antiplasmin band (68,000) was converted into two products with estimated mol. wts of 56,000 and 11,000. Incubation of human ct2-macroglobulin with the three proteinases at a molar ratio of 1: 1 resulted in the complete loss of the macroglobulin activity (trypsin binding capacity). All three enzymes were inhibited by 20 mM EDTA, they had no thrombin-like effect on fibrinogen and no other procoagulant activities. Induction of vascular protein leakage by the three proteinases and by crude venom, which caused plasma extravasation into the surrounding tissue, could be observed 2 min after intradermal injection of the three proteinases and of crude Crotalus basiliseus venom.
Influence ofdiazepam on hornet fertility. E. Fuksman, I A. Weizman, 1.2 M. Rehavi I and J. S. Ishay I (i Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv 69978, Israel; and 2Research Unit, Geha Psychiatric Hospital). The experiments were done with queenless hornets (Vespa orientalis, Vespinae, Hymenoptera) kept in laboratory conditions in artificial breeding boxes. Feeding of diazepam (Assival) to young hornets completely inhibits or delays for a relatively long period development of their ovaries. In control hornets the ovaries usually develop within 1 or 2 days post-eclosion and comb building commences on the 2nd day of life. The hornets then oviposit into the comb cells and the deposited ova give rise to larvae. Trials were performed on parallel groups of hornets of various ages. When the sedative was administered to hornets aged 0-24 hr the ovaries of these young hornets failed to show any development, so that no oocytes ripened and consequently there was no oviposition whatsoever. Neither were any comb cells built or, at best, only a few were built. When diazepam was given to hornets aged 2 4 4 8 hr, there was only slight development of the ovaries giving rise to only a few eggs and the delayed building of merely a small comb. When the sedative was administered to hornets beyond the age of 48 hr, it exerted no change, i.e. the eggs developed normally and comb building was the same as in the control group. Longevity of hornets was uniform in all the test groups and similar to that in the control. Treatment with flumazenil (Anexate), which is a competitive antagonist to diazepam, did not abrogate effect of the latter on any of the hornet test groups. A few experiments were run to assess, via the radioactive ligand [SH]Ro 15-1788, possible specific binding in neural tissues of hornets of various ages, but no significant binding was detected in any of these. In subsequent experiments we intend to assess [SH]PK 11195 binding to peripheral tissues. Part of M.Sc. thesis to be submitted by EF.
Antigenic properties of allergenic proteins from oriental hornet (Vespa orientalis) venom. E. Levtov, l A. I. Pick, 2 I. Hammel 3 and J. S. Ishay ~ (~ Department of Physiology and Pharmacology, 2Division of Clinical Immunology and Allergy, Beilinson Medical Center, and 3Department of Pathology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, 69978, Israel). In the venom of the Oriental hornet (Vespa orientalis, Hymenoptera, Vespinae), proteins comprise the primary effector of the allergic phenomena induced by its sting. From the venom of the Oriental hornet obtained from adult specimens, we reported the proteins by the techniques of SDS-PAGE and gel filtration. The following proteins were separated out; hyaluronidase having a mol. wt of 45 kDa, two phospholipases of 33 kDa and 30 kDa, respectively, and antigen 5 having a mol. wt of 24 kDa. Polyclonal antibodies to each of these proteins separately were developed in rabbits injected with the isolated venom proteins. Each of the isolated proteins was antigenic and elicited antibody production. The strongest antigen proved to be hyaluronidase while the weakest was antigen 5. The phospholipase elicited a moderate antibody response. Specific assessment of the produced antibodies via immunoblotting revealed the highest specificity of antibody response to be against antigen 5, while the antibodies against the hyaluronidase and the phospholipases displayed very strong cross-reactivity, It is suggested from these findings that significant part of the antibodies elicited are anti-carbohydrate antibodies. Hyaluronidase from Oriental hornet venom cross-reacted strongly with hyaluronidase from the venom of the German wasp (Paravespula germanica). In contrast, the cross-reactivity between the phospholipases from the two mentioned wasp species was relatively weak, while the antigen 5 from the venoms of both did not cross-react at all. In tests for cross-reactivity between the venom proteins of the Oriental hornet and proteins from human body fluids, we found that both hyaluronidase and antigen 5 share antigenic determinants with human seminal fluid, while hyaluronidase also possesses antigenic determinants in common with human serum. An additional finding was that in the serum of rabbits not exposed to hornet venom components, there are antibodies that recognize venom components of both the Oriental hornet and German wasp, although the titre of such antibodies is rather low. This latter finding is probably explainable by the similarity in amino acid sequences within hornet venom components and proteins of plant origin which constitute food of rabbits. Part of Master thesis to be submitted by EL.
Eleventh World Congress
proteinase a, b and c had estimated mol. wts of 25,000 and 24,000, respectively, as determined by SDS PAGE and a pl of 9.0, 9.3 and 8.5. Incubation of the proteinases with purified human ~2-antiplasmin at a molar ratio of 1:10 resulted in a loss of more than 90% of the initial ~t2-antiplasmin activity within 60 min. In SDS PAGE the ~2-antiplasmin band (68,000) was converted into two products with estimated mol. wts of 56,000 and 11,000. Incubation of human ct2-macroglobulin with the three proteinases at a molar ratio of 1: 1 resulted in the complete loss of the macroglobulin activity (trypsin binding capacity). All three enzymes were inhibited by 20 mM EDTA, they had no thrombin-like effect on fibrinogen and no other procoagulant activities. Induction of vascular protein leakage by the three proteinases and by crude venom, which caused plasma extravasation into the surrounding tissue, could be observed 2 min after intradermal injection of the three proteinases and of crude Crotalus basiliseus venom.
Influence ofdiazepam on hornet fertility. E. Fuksman, I A. Weizman, 1.2 M. Rehavi I and J. S. Ishay I (i Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv 69978, Israel; and 2Research Unit, Geha Psychiatric Hospital). The experiments were done with queenless hornets (Vespa orientalis, Vespinae, Hymenoptera) kept in laboratory conditions in artificial breeding boxes. Feeding of diazepam (Assival) to young hornets completely inhibits or delays for a relatively long period development of their ovaries. In control hornets the ovaries usually develop within 1 or 2 days post-eclosion and comb building commences on the 2nd day of life. The hornets then oviposit into the comb cells and the deposited ova give rise to larvae. Trials were performed on parallel groups of hornets of various ages. When the sedative was administered to hornets aged 0-24 hr the ovaries of these young hornets failed to show any development, so that no oocytes ripened and consequently there was no oviposition whatsoever. Neither were any comb cells built or, at best, only a few were built. When diazepam was given to hornets aged 2 4 4 8 hr, there was only slight development of the ovaries giving rise to only a few eggs and the delayed building of merely a small comb. When the sedative was administered to hornets beyond the age of 48 hr, it exerted no change, i.e. the eggs developed normally and comb building was the same as in the control group. Longevity of hornets was uniform in all the test groups and similar to that in the control. Treatment with flumazenil (Anexate), which is a competitive antagonist to diazepam, did not abrogate effect of the latter on any of the hornet test groups. A few experiments were run to assess, via the radioactive ligand [SH]Ro 15-1788, possible specific binding in neural tissues of hornets of various ages, but no significant binding was detected in any of these. In subsequent experiments we intend to assess [SH]PK 11195 binding to peripheral tissues. Part of M.Sc. thesis to be submitted by EF.
Antigenic properties of allergenic proteins from oriental hornet (Vespa orientalis) venom. E. Levtov, l A. I. Pick, 2 I. Hammel 3 and J. S. Ishay ~ (~ Department of Physiology and Pharmacology, 2Division of Clinical Immunology and Allergy, Beilinson Medical Center, and 3Department of Pathology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, 69978, Israel). In the venom of the Oriental hornet (Vespa orientalis, Hymenoptera, Vespinae), proteins comprise the primary effector of the allergic phenomena induced by its sting. From the venom of the Oriental hornet obtained from adult specimens, we reported the proteins by the techniques of SDS-PAGE and gel filtration. The following proteins were separated out; hyaluronidase having a mol. wt of 45 kDa, two phospholipases of 33 kDa and 30 kDa, respectively, and antigen 5 having a mol. wt of 24 kDa. Polyclonal antibodies to each of these proteins separately were developed in rabbits injected with the isolated venom proteins. Each of the isolated proteins was antigenic and elicited antibody production. The strongest antigen proved to be hyaluronidase while the weakest was antigen 5. The phospholipase elicited a moderate antibody response. Specific assessment of the produced antibodies via immunoblotting revealed the highest specificity of antibody response to be against antigen 5, while the antibodies against the hyaluronidase and the phospholipases displayed very strong cross-reactivity, It is suggested from these findings that significant part of the antibodies elicited are anti-carbohydrate antibodies. Hyaluronidase from Oriental hornet venom cross-reacted strongly with hyaluronidase from the venom of the German wasp (Paravespula germanica). In contrast, the cross-reactivity between the phospholipases from the two mentioned wasp species was relatively weak, while the antigen 5 from the venoms of both did not cross-react at all. In tests for cross-reactivity between the venom proteins of the Oriental hornet and proteins from human body fluids, we found that both hyaluronidase and antigen 5 share antigenic determinants with human seminal fluid, while hyaluronidase also possesses antigenic determinants in common with human serum. An additional finding was that in the serum of rabbits not exposed to hornet venom components, there are antibodies that recognize venom components of both the Oriental hornet and German wasp, although the titre of such antibodies is rather low. This latter finding is probably explainable by the similarity in amino acid sequences within hornet venom components and proteins of plant origin which constitute food of rabbits. Part of Master thesis to be submitted by EL.