Marinov, G.

Trosheva, M.

Yankova, T.

English

Varna Medical University Press: Journals

ADENOSINE TRIPHOSPHATASE ACTIVITY OF T H E VENOUS W A L L IN CAT G. Marinov, M. Trosheva, T . Yankova Key-words: adenosine triphosphatase — vein — smooth muscle cells — artery The knowledge about the act iv i ty of enzyme systems in venous wal ls are rather scanty regardless of the strong interest in vascular wal l metabolism (9, 11, 12). I t can be due along with the purely subjective "step-mother" treat­ment of this problem caused by false concepts that veins are simple rigid tubes (9) to the objective difficulties when investigating the enzyme systems in the venous wal l because of its heterogenous tissue composition (12) and great var ia ­bi l i ty of its structure ( 1 , 3). The aim of the present work is to make a histochemical and biochemical characteristics of adenosine triphosphatase (ATP-ase) act ivi ty of the venous w a l l when veins of different size and structure type are investigated. Literature data about this problem are scanty and contradictory (5, 6, 11, 14, 16). They are not satisfactory when the raising interest in this topic is concerned. Recently, it i s determined by the importance of this enzyme system for the contractile function of the venous wal l (11 , 12). Its disturbances are of essential c l in ical signifi­cance-Material and methods Our study covered 30 sexually mature cats. The following veins were examin ed: v . portae, mesenteric ve in , inferior caval ve in , and femoral ve in . Material-was taken from the abdominal aorta and the femoral artery as controls. Myosin ATP-ase was histochemically ascertained after the calcium-cobalt method of Padykula and Herman in modification (2) at p H 9,2 but non-myosine one — after Wachstein-Meisel 's method in modification (2) at p H 7,2. Sections incu­bated in solution without substrate and sections inactivated by heating in d is t i l ­led water at 80 °C for 10 min (through the latter ones non-enzymatic A T P hyd­rolysis was excluded) were used as controls. The vessels were homogenized in 10 шМ r i s - H C l buffer wi th 0,25 M sucrose at pH 7,4 after preliminary freez­ing at —20 °C for quantitative investigation. The homogenate obtained was centrifugated at 3000 g for 10 min. Supernatant was used to determine K + and ~Ta+ activated Mg 2 + ATP-ase . The determination was carried out after K i d w a y ' s method in a medium containing: MgCl 2 3 mM, K G 10 mM, NaCl 100 mM A T P . mM, r i s - H C l buffer 50 mM at p H 7,4 in a volume of 1,5 ml . Protein of any amples was about 20 ug. Ouabain-sensitive K + and Na+ activated A T P - a s e as determined as a difference between K + and Na+ activated Mg 2+ ATP-ase nd the same one in the presence of 0,2 mM ouabain. P i was determined af-er Tausk i and Shorr (1953) but protein after Lowry et a l . (1951) by using of va lbumin as standard. 'i 32 G . Marinov, M . Trosheva, T. Yankova Results and discussion Myosin and non-myosin ATP-ase histochemical determination after the me­thods described above showed that the reaction was positive in smooth muscle cells (SMC), endothelial cells ( E C ) and in proper vessels (Vasa vasorum). A s S M C were the most considerable component of the venous wal l enzyme ac t iv i ty dis-T i g . 1. Histologic cross-section of v. portae. Reaction for non-myosine ATP-ase after Wach-stein-Meisel. Microphotograph, ос. X 10, ob. X 20. Positive reaction in the circular muscular layer of the medial tunic and in the longitudinal muscular layer of the adventitial one tribution in single vascular wa l l tunics was determined mainly by SMC presence in them. W i t h vena portae and mesenteric ve in that possessed a well-developed circular muscular layer in the medial tunic and a massive longitudinal one i n the adventit ial tunic both reactions were positive in both tunics (fig. 1). How­ever, this could be established only in the areas where the adventit ial tunic posses­sed musculature. W i t h the inferior caval ve in that possessed a well-developed circular muscular layer in the medial tunic and practically did not contain any SMC in the adventitial one both reactions were positive in the medial but nega­t ive in the adventit ial tunic excepting singular circumscribed areas containing SMC as well as in the walls of adventit ial intramural vessels. W i t h the femoral ve in wa l l that was considerably thinner and possessed a well-developed circular layer in the medial tunic the reactions were positive mainly in the medial tunic (fig. 2) . The quantitative determination showed that the relative part of ouabain-sensitive K + and Na+ activated ATP-ase in relation to total K + and Na+ acti­vated Mg 2 + ATP-ase remained over 70 per cent in the veins studied. The venous wa l l showed s imilar i ty wi th the arterial one of muscular type as demonstrated by us in the femoral artery (table 1). However, this part was considerably higher than the analogous one when arteries of elastic type were concerned as demonstra­ted in the abdominal aorta (table 1). Adenosine triphosphatase activity.. 33 The histochemical determination of both myosin and non-myosin ATP -ase activities of the venous wal l showed that the presence of these enzymes was first of a l l related to SMC, E C and vasa vasorum. As venous wal l structure and espe­cially SMC distribution demonstrated a great local and regional variability ( 1 , 3) the localization of ATP-ase ac t iv i ty in the three venous wall tunics (inti-mal v medial and adventitial) varied in veins wi th different structural type. This Fig . 2. Histologic cross-section of the femoral vein wal l . Reaction for non-myosine ATP-ase after Wachstein-Meisel. Microphotograph, ос. X 10, ob. X 20. Positive reaction in the circular muscular layer of the media fact could explain some literature controversies concerning the localizat ion of this enzyme in the venous w a l l . These differences were due to insufficiently pre­cise distinction of venous wa l l tunics as wel l as to ignoring of local and regional variations in their structure. - T a b l e 1 Relative part of ouabain-sensttive K"1" and Na^~ activated ATP-ase towards К and N a ^ activated Mg ~ ATP-ase V . portae and v . mesenterica 81,25 % V . cava inferior 69,10 % V . femora lis 79,95 % Aorta abdominalis 42,34 % Arteria femoralis 73,70 % Positive histochemical reactions for both myosin and non-myosin ATP-ases suggested the possibility that SMC of venous wa l l could use high-energy A T P 3 Scripta scientifica medica 34 G . Marinov, M. Trosheva, Т. Yankova (8) while positive reaction for myosine ATP-ase indicated the degree of A T P uti l izat ion for SMC contraction in the veins studied (7). However, the methods applied did not enable the ascertaining of sure gradual differences in the acti­v i t y of both ATP-ases although on the basis of literature data about the presence of differences between arteries and arterioles (7) and between the mesenteric Ar­tery and vein in rat as well (11) such differences could be supposed between1 the venous vessels studied by ourselves as well as between arterial and venous vessels examined in our study. The quantitative examination revealed an interesting fact that the relative part of ouabain-sensitive K + and Na+ activated ATP-ase towards Na+ and K + activated Mg 2 + ATP-ase remained over 70 per cent and was s imilar to that wi th arteries of muscular type while it was considerably lower with arteries of elastic type. Th i s fact requires a further investigation on SMC plasmalemmal membrane fractions of veins wi th different structure and size (13). Th i s can enab­le a more precise interpretation and reflect on the restricted progress of studying K + and Na+ ATP-ase as an enzyme expression of Na+ pump in SMC (12) as wel l as on s t i l l unclarified question of ATP-dependent Ca 2 + transport in venous w a l l SMC (11). These processes are most probably of essential importance for the early adaptation of the venous wal l in case of such widespread disease, namely the varicose veins of the lower limb (4). R E F E R E N C E S 1. Б а н к о в , В. Строеж и васкуларизация на вените. Докт. дис. Варна—Ленин­град, 1968. — 2. Л о й д а, З. , Р . Г о с с р а у, Т. Ш и б л е р . Гистохимия ферментов. М., Мир, 1982. — 3. М а р и н о в , Г. Строеж и васкуларизация на вените на подбедри-цата и задколянната яма. Канд. дис. Варна, 1969. — 4. М а р и н о в, Г., В. Б а н к о в . Ранни промени в гладкомускулните клетки на венозната стена при развитието на вари-козен процес. — В: Биология и патология на съдовата система. С , 1984, с. 34. — 5. A d a m s , С. W. М. Arterial histochemistry in relation to structure, function and di­sease. — I n : Structure and function of the circulation. New York, etc., Plenum Press, 1982, 3, 1 — 157. — 6. M с С a b е, M. G . J . , A. P . С i n n i n g d a m, N . G . W у a t t, C. W. R o t h i n e . Brit. J . Surg., 54, 1967, 147. — 7. С a n n о n, M. S., C. J o n e s , Т. V . P e t e r s o n . Blood Vessels, 19, 1982, 186—196. — 8. С a n n о n, M. S. , E . D . К а г е s, J . W. W a g n e r , G . C. G o d w i n , J . B . G e l d e r d . Acta Anat., 113, 1982,296— 312. — 9. H a s h i m o t o , J . , S. S h i k a t a t a , Т. H a r a . Acta Histochem. Cyto-chem., 9, 1976, 197—202. — 10. K i d w a y , A . M. R a d c l i f f е , Е. E . D a n i e l . Bio-chim. Biophys. Acta, 2ЗЗ, 1971, 38—43.— 11. К w a n, C. Y . Enzyme, 28, 1982, 317— 327. — 12. K w a n , C. Y . , P. K o s t k a . MoL Physiol., 3, 1983, 265—275. — 13. K w a n , C . Y . , R . M. K- W. L e e , Е. E . D a n i e l . Blood Vessels, 18, 1981, 171 —186. — 14. L о r е n z, D . , A . D o r n , R . R e d i n g . Arch. Histochem., 41 , 1971, 1—6. — 15. L о w r y, D. H . , A . R o s e n b r o u g , A . L . F a r r, R . J . R a n d a l l . J . Biol. Chem., 193, 1951, 265. — 16. P r e r o v s k i , I . , J . S v e j с a r, D. U r b a n o ­v a , E . K r u s z e w s k a , J . L i n h a r t. I n : Congr. Internat. de Phlebologie, I I I . ' Rap­port, 691—694. — 17. T a u s k y , N . H . , E . S h o r r . J . Biol. Chem., 202, 1953, 675— 685. Adenosine triphosphatase activity. 35 АДЕНОЗИНТРИФОСФАТАЗНАЯ АКТИВНОСТЬ ВЕНОЗНОЙ СТЕНКИ У КОШКИ Г. Маринов, М . Трошева, Т. Янкова Р Е З Ю М Е Исследовалась аденозинтрифосфатазная активность венозной стенки у кошки. Из­учались вены с различным просветом и типом строения. Применялись гистохимические ме­тоды Padykula, Herman (1955) и Wachstein Meisel (1957) в модификации Лойда, Госрау, Шиблер (1982), а также биохимический метод Klaway (1971). Отделяемый фосфат опреде­лялся ме одом Tausky, Shorr (1953). Локализация аденозинтрифосфатазной активности венозной стенки обусловлена ее структурными особенностями. Энзимная активность вы­является в медии или в медии и адвентиции в зависимости от наличия гладкой мускулату­ры и собственных сосудов в них. Количественное определение пока ывает, что удельный вес оубаин-сенситивной Na^~ и К~̂~ — активированной аденозинтрифосфатазы в изучаемых нами венах задерживается выше 70 %. При этой стоимости венозная стенка похожа на стен­ки артерий мышечного типа. 

ADENOSINE TRIPHOSPHATASE ACTIVITY OF THE VENOUS WALL IN CAT

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ADENOSINE TRIPHOSPHATASE ACTIVITY OF THE VENOUS WALL IN CAT

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