Uzunova, Anelia D.

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Varna Medical University Press: Journals

% и < > ^ С (Л 4 Л 1 /8 ~  '  —  t t y s -WAftAfj 1̂ *YI/1/C£ /OCC'J-̂  '■y*- v/Y /  J'tyt- /  ЛCHANGES IN THE SYSTEM HISTIDINE DECARBOXYLASE-HISTAMINE HISTAMINASE IN NEWBORN THYMECTOMIZED RATSAnelia D. UzunovaIn the past several years the thymus has become the favorite topic of numerous research works and publications (Miller J. F. 1963; *964-ana1965, Azar Henry A. 1963, Policard A. 1963, Good R. A. 19b4,Burnet F. M. 1964, Kemileva Z. — 1965, and others).The incessantly growing interest in this organ is justified on account or the fact that lately, an outstanding place has been ascribed to it in the bio­logical hierarchy (classification) of organs. According to the statement of M. Simmonsen (quoted by 6), „Miller reinstated this half-forgotten organ, specifying its lawful role in ibimunology“.At present, the concept is adopted as definitely established that all immune reactions in the organism are controlled by the thymus (3, 13). At the end of the antenatal period and in newborns it is the chief source for the pro­duction of immunocapable cells for the organism and a center, regulating their distribution.In the opinion of P. Kallos (cited by 16), the studies on the thymus are of utmost importance not merely for immunobiology in the narrow sense of the word, but also for better understanding of all allergic and auto-aller­gic affections; the role played by the thymus in the etiological mechanism of the latter is as yet unknown.Parrot M. V. (20), on the ground of personal experiments, reached the conclusion that the role of the thymus in the development of autoimmune diseases needs further experimental investigations.On the other hand, it is universally accepted that in the course of inflamma­tory and allergic reactions, the histamine a highly active biogenic mono­amine — plays an outstanding role (4, 6, 11, 17, 18, 21, 27). Owing to its pronounced vasoactivity, it participates in vascular reactions in the course of inflammatory phenomena, stimulates phagocytosis and reparative pro­cesses (11, 18, 26). Bath K. G. and Sanyal R. B. presented evidence (6) for enhancement of the allergic component of aseptic and septic inflammation, parallel to the increase of histamine content in tissues.In other experiments (9, 19) it has been demonstrated that antirheumatic drugs inhibit histamine liberation, induced by histamine liberators and b\ the antigen-antibody reaction.The active participation of the thymus in the immune reactivity of the organism and the role played by histamine in the inflammatory and aller­gic responses provided the basis of undertaking a follow-up study on the finfluence of thymectomy upon the system histidine decarboxylase-hista- mine-histaminase all the more, that references on similar investigations are scarcely found in the pertinent literature.16 A. D. Uzunova 2Material and methodA series comprising 120 white, unbreeded male albino rats is employed:' 62 thymectomized and 58 — for control purposes.Thymectomy is performed on newborn rats 0 to 48 hours after birth in accordance with the aspiration method, developed at the Chair. The inter vention is carried out on non-anesthetized animals under strict asepsis. The wound is sprinkled with streptothricin powder and in case of hemorrhage, hemostatic spongy is applied.The investigation of histamine and enzymes of both groups (controls and thymectomized) is carried out at the third month of life. At this age, accor­ding to Dalmasso A. L., Martinez C. and others, the immune capacity of neonatally thymectomized rodents is yet unrecovered.The histamine level is studied in the heart and in the paraarticular connec­tive tissue and muscles in view of the fact that experimental myocarditis and arthritis in intact and thymectomized animals with tracing of systemic histidine decarboxylase-histamine-histaminase, will be subject to subse­quent investigations.Apart of histamine in the tissues studied, the activity is also traced of the histamine-producing enzyme — histidine decarboxylase and of the chief histaminedestroying enzyme — histaminase; the two enzymes just mentio­ned determine the histamine level in the tissues. Investigations on the enzymic activity are also carried out in the small intestine due to the fact that the stated localization is characterized by intense synthesis and deve­lopment of histamine, having an essentia! bearing on the regulation of its total level within the tissues.The animals are killed by striking a blow with a solid object on the head and subsequent decapitation. The material for investigation is obtained promptly after sacrification, ice treated and accordingly elaborated.The histamine is extracted from the tissues after the method of E. Kovach as modified by Shelljazkov and Uzunov (1). Its quantitative determination is performed according to a chemical method, suggested by Tabor and modi­fied by the authors just cited. Histidine decarboxylase and histaminase are determined according to the Telford and West method (8, 23, 24, 25, 26. 28), modified by Shelljazkov and Uzunov (1). Histidine decarboxylase acti­vity is expressed in mg histamine, produced by mg protein for 3-hour incu bation. The histaminase activity is likewise expressed in mg histamine, inac­tivated by mg protein in 2-hour incubation.The protein in the enzymic material is determined after the Lowry method.The listed below reagents are used for the enzymatic reactions: histidine, histamine hydrochloride, amino guanidine and pyridoxale-6-phosphate commercially available by the firm Merck.The data recovered are statistically elaborated according to the method of Student.Results and discussionThe results of the experiments performed are illustrated in tables 1, 2 and 3 and in figures 1 and 2.Investigation of histamine content in the control animals disclosed in the average 9,1 mg histamine per gram fresh cardiac tissue and 9,7 mg for3 Changes in the System Histidine Decarboxylase—Histamine 17Table 1Hislainine per gram Fresh TissutTissues nIntact animals Thymectomized animalsp inX <Jx n X <JxJoints 36 9,7 0.3 ■10 N,7 0,4 0,001Heart 36 9.1 0,02 40 11.7 0,5 0,001the paraarticular connective tissue and muscles. These values conform with the data published for histamine level by W. Feldberg in 1955 (International Histamine Symposium) (4). The normal values found out are similar to those reported by other authors (18, 28), employing the biological method for histamine testing.The histamine level in the tissues investigated of thymectomized animals is rised: ll,7pg/gram tissue in the heart and 14,7 pg/gram tissue in the joints, which, expressed in percentage, are accordingly 128% and 159%. The difference in the values of histamine in intact and thymectomized ani­mals is statistically reliable [p (t) <  0,001] (Fig. 1).Mg histaminne per gram fresh tissue□  in tact animals 1%1 thymectomized animalsFig. IHistidinedecartoxylasof histamine per mg protein tor $  hours)□  intact animalsthymectomized animalsFig. 2Table 2 illustrates the data obtained for histidine decarboxylase activity in mg histamine per mg protein, over a 3-hour incubation. They appear to be quite significant insofar changes in histamine level are concerned. In all thymectomized animals, the average increase of histidine decarbo­xylase activity amounts to 151% for the small intestine, 137% for the cardiac muscle and 122% for the joints. The differences in enzymatic acti­vity are statistically reliable [ p (t) <  0,999] (Fig. 2).It is evident from table 3 that histaminase activity in thymectomized animals does not differ within statistically reliable range from the activity2 Scriptn scientifica inedicu18 Л. D. UsunovaTabl eHistidine Decarboxylase Activity (ц£ Histamine per |xg Protein for 3 hours)TissuesIntact animals Thymectomized animalsp (1)ti X <Tx n X dxSmall intestine 12 262 10 12 396 18 <  0,00Joints 12 119 17 12 146 5,7 <  0,00Heart 12 69 3,4 12 95 3,4 <  0,00of the enzyme in intact animals l p (t) >  0,50]. The latter fact is corrobo rated by the results obtained, concerning the histamine and histidine decar boxylase activity level, since there would be no increase of histamine content in the tissues investigated in spite of enhanced histidine decarboxylase activity, if it were accompanied by an increase of the chief histamine-inacti­vating enzyme as well — the histaminase. It is beyond doubt that the rise in tissue histamine is brought about by intensified synthesis.Tabl e  3Histaminase activity ([eg Histamine per \xg Protein for 2 hours)TissuesIntact  animal Thymectomized a nimelsP (*ln X dx n X dxSmall intestine 10 231 5,5 10 232 16 >  0,50Joints 10 1 16 8,0 10 121 13 0,50Heart 10 29 9.2 t o 21 8,4 0,50Moreover, it is worthwhile mentioning the fact that histaminase activity in the heart of both groups of animals is rather low. In half of the cases, the amino guanidine, added to the enzymic sample (a substance accounting for complete histaminase inhibition), instead of precluding histamine destruc­tion, activies, although insignificantly, its separation (development). It is rather difficult to find an explanation of this phenomenon merely on the ground of the experiments conducted. In all likelihood, other enzymes as well (methyl, acetylene etc.) exert a controlling effect on the histamine me­tabolism in the heart.The paucity of literature references dealing with changes in the system histidine decarboxylase histamine — histaminase in thymectomized animals deprives the authors of the possibility to make a comparative assess­ment of the results in the experiment herein described.5 Change* in the System Histidine Decarboxylase—Histamine. . 19At this stage of the work, it is rather difficult to establish the contribut­ing cause for the alterations in the system histidine—decarboxylase — hista­mine — histaminase, but anyway, they could presumably be related to the complex metabolic and morphologic changes, occurring in thymectomi- zed animals (3).Acknowledgement: We wish to express our deepest gratitude to Prof. Shelljazkov, Head of the Chair of Pharmacology, for his valuable aid and guidance during the application of the methods for histamine investigation.R E F E R E N C E S1. Ж е л я з к о в ,  Д.  K-, П. Д.  У з у н о в .  I национален конгрес но биохимия ибнофизика, 25—27 май, 1966.2. К е м н л е в а ,  3. — Експериментална медицина и морфология, 1966 , 2, 69—70. 3- П ол нка р ,  А. Физиология и патология лимфоидноЛсистемн Медта, М., 1963.4. У с п е н с к и й ,  В. И. Гистамнн. Медгиз, М., 1963.5. A z a r H e n r y ,  А. — Arch. Pathol., 1963 , 76 , 653 —658.6. В ha th , К. G-, R- K- S a n y a l .  — 7. pliarrn. and pharmacol., 1964, 16, 385.7. Co ms a, .1. Physiologic et physiopathologie du thymus. Paris, 1959.8. H a e g e r ,  K., G. Kal i  I son.  — Acta phys.scand., 1962, 25, 230—242.9. H a i n i n g ,  C. G. -  Brit. J. Pharmacol., 1965, 11, 35710. G i e r t, R. J., G. dc P a s q u a 1 c. — J. Endocrinol , 1963, 26, 2. 209 213.11. G o z s y ,  B., L., K a t o. — Arch. int. pharmacodyn., 1961, 148, 1—2.12. G u n t h e r ,  O. -  Dtscli. med. Woclwnschr., 1964, 89, 987—99013. M i l l e r ,  J. F. Lancet, 1, 43, 1963.14. M i l l e r ,  J. F. -  Ann. hist. Pasteur, 1963, 105, 1007-100915. Mi l l e r ,  J. F. — Science, 1964, 144 1544—155116. Mi l l  er, .1. F., Du k n k .  Die Biologiedes Thymus, S. Karger, Basel—New York,196417. M i s h r а. В. P., R. K- S a n y a l .  — J. Pliarm. and pharmacol., 1955, 11, 127.18. N o r п-S v e n d. Aitu pharm. et toxicol., 1965, 22, 369—378.19. N o r t h o v e r ,  B. .1.. G S u h r n m a n i a n .  Brit. J. pharmacol., 1961, 16,163—169.20. P a r r o t t .  M. V. — Acta alter polo pica, 1965, 20, 227—234.21. S t e r n ,  P.. A. N i c u l i n .  — Intern, arch, allergy, 1960, 16, 157—162.22. S iK h o r I a n d D a v i d ,  Е. K- O 1 g a, K. A r c h e r, D. A. P e t e r s e n  Ra y-m o n d et al. Lancet, 1965, 7377, 130—133.23. T e l f o r d ,  J. M. — Nature, 1963, 197, 701.24. T e l f o r d ,  .1. M., G. B. We s t .  J. pharm. and pharmacol., 1961, 18, 75.25. T e l f o r d ,  .1 M. — Biochemical pharmacology. 1965, 14, 1713—173026. We s t ,  G. B. — J. pharm. and pharmacol., 1952, 14, 61827. We s t ,  G. B. — J. Pharm. and pharmacol., 1959, 11, 513.28. W a t o n, N. G. — Brit. J. pharmacol., 1956. 11, 119.ИЗМЕНЕНИЯ В СИСГЕМЕ ГИСТИДИНДЕКАРБОК.СИЛ A3A — ГИСТАМИН —Г ИСТ АМИНАЗА У НЕОНАТАЛЬНО ТИМЗКТОМИ РОВ AH HblX KPMCАннелия Д. Узунова РЕЗЮМЕАвтор изучает количественно изменения в системе гистидиндекарбо- ксилаза — гистамин гистаминаза в мьннце сердца, околосуставной соединительной тканн и в тонких кишках у неонатально тимзктомиро- ванннх бель(х крме.20 A. D. UzunovaУстанавливается статистически достоверное повишение уровня гиста- мина в изучаемнх тканях на третьем месяце после тимзктомии.Гистаминообразующий знзим гистидиндекарбоксилаза, также увеличи- вается в тканях тимзктомированннх крие в статистически достоверннх размерах.Количествени не изменения в активности гистаминази не уетанавлш ваютея.Демается попитка обсуждения результатов проведенннх зке пер и ментов.

CHANGES IN THE SYSTEM HISTIDINE DECARBOXYLASE-HISTAMINE HISTAMINASE IN NEWBORN THYMECTOMIZED RATS

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CHANGES IN THE SYSTEM HISTIDINE DECARBOXYLASE-HISTAMINE HISTAMINASE IN NEWBORN THYMECTOMIZED RATS

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