Negrev, N.

English

Varna Medical University Press: Journals

S T U D Y OF T H E E F F E C T S OF NON-SELECTIVE B E T A - A D R E N E R G I C T R E A T M E N T ON PLASMA THROMBOCYTOPOIETIN ACTIVITY [IN RATS IN CONDITIONS OF AN A C U T E THROMBOCYTOPENIA N. Negrev K e y-words: thrombocytes — 7 5selenomethionine incorpora tion — β-adrenergic a g e n t s — a c u t e thrombocytopenia The relation between the count of peripheral blood thrombocytes and plasma hrombocytopoietin activity is long ago established. It is known that normally, thrombocytopenia is attended by thrombocytopoietin production increase (8, 10) whereas any thrombocytosis — by its decrease (6, 13). Our previous studies show that thrombocytopoietin production in rats in ormal conditions depends also on beta-adrenergic receptors (1 , 3). On the basis f that we make it our object to studying to what extent non-selective (5-adre-ergic effects are related to plasma thrombocytopoietin activity in conditions of n acute thrombocytopenia. This could enable more profound conclusions con-rning the importance of non-selective ^-adrenergic influences and (i-adreno-ptors, respectively, on thrombocytopoietin production. Material and methods Plasma required for thrombocytopoietin activity determination was obtain-d from 53 male rats of Wistar breed with 180—220 g body weight divided into ree groups and injected as followed: I s t group — with Isoprenaline ( IP) hydro-hloride (a non-selective p-adrenostimulator) at dosis 2 X 3 mg/kg b. w.; I I n d group — with Propranolol (PR) hydrochloride (a non-selective P-adrenoblocker) t dosis 2 x 5 mg/kg b. w., and I I I r d group — controls — with saline in the same mount. Injections were made intraperitoneally every 12 hours. 3 h after first pplication all the rats were injected with thromboplastin (TP) after the scheme f Kelemen et al . (7) in our modification, namely: We used T P produced by the esearch Institute of Hematology and Transfusion —_Sofia in our study. The "st injection was done in the tail vein in_yol. 0.5.10 6 m 3 but the second one "ter 40 min intraperitoneally in vol. 1.10 6 m 3 . Thrombocyte count was deter-ined according to the method of Feissly et Ludin (4) at the beginning and the d of the assay. Animals were bleeded by means of aortic cannulation on the 20 t h hour after first T P injection. Separated plasma was stored at —8° С for 10 days, е judged of plasma thrombocytopoietin activity from the changes of thrombo-te count and from 75 s elenomethionine ( 7 5Se-M) incorporation in newly formed rombocytes in recipients-mice under the influence of material tested. We used 1 white male non-thoroughbred mice with 20—30 g b. w. divided into 3 groups, he animals from the first group were injected with plasma from donors treated ith I P + T P , those from the second one — with plasma from donors treated ith P R + T P , and the mice from the third group were controls (injected with plasma from donors treated with saline+TP). We tested the plasma after Pening-ton's method in our modification (12). I t included the following changes: 52 N . Negrev a) In the place of male mice С 57 we used white mice — male non-thorough-bred mice with initial thrombocyte count — 200—380.109/1; b) We injected the specimens in two successive days s. c. at dosis 1.10~6m3 each as contrasted with Penington's method (12) who applied a dosis not higher than 0.5.10~~6m3 but injected the mice with antithrombocyte serum one week before; c) , We injected the isotope ( 7 5Se-M) intraperitoneally at dosis 2 uCi per mouse 1 h after second plasma injection but not on the next day as it was according to Penington. We took 0.5.10~ 6m 3 blood for estimation of the percentage of 7 5Se-M incor­porated in newly formed thrombocytes on the 72 n d hour by using cardiac punc­ture under ether narcosis. We took the determination of isotope incorporation on the 72 n d h (in contrast to Penington who determined it on the 24 t h h) because this period coincided with the peak of thrombocyte count and of percentage of 7 5Se-M incorporated in newly formed thrombocytes. Thrombocyte separation was done according to Penington's method (11) but radiometrics — on the apparatus N / K 350 — Hungary. Data processing was done by the methods of variation statistics. Results and discussion It can be seen on fig. 1 that thrombocyte count decreases at the end of the as­say with 18.96 per cent (p<0.002) as compared with the initial one in mice from the first group but it increases with 95.09 per cent (p<0.001) in mice from the 10*11 10 8 6 В,35±0,25 U-,43*0,17 4,28±OJ7 3,59±0t1l 18,96V* p<0№ 9p<0o9oot W ± 0 > 3 2 in 30,63% p<0,00f JPA(IP+TP 1PA(PR+TP TPA(S+TP • Values in ш бедслп-of the experiment EZ3 Values in the end of {he experiment F i g . I . T h r o m b o c y t o p o i e t i n a c t i ­v i t y of donor's p l a sma — ra ts trea­ted v i t h ^-adrenergic agen ts a n d thromboplastin de te rmined b y i n -• ves t iga t ion of the changes of throm­bocyte coun t i n r e c i p i e n t s - m i c e . Da ta are presen ted as x ± : S x . P e r ­centage difference is" c a l c u l a t e d to-wards i n i t i a l va lues second group. Thrombocytes increase with 30.63 per cent (p<0.00l) in control mice. 75Se-M incorporation in thrombocytes (fig. 2) in experimental and control animals both shows that there is a reduction with 53.48 per cent (p< 0.001) in the first group and an increase with 754.1 per cent (p<0.001) in the second one. Study of the effects o f . . , 53 6,28 ±0,22 It is first of all notable that these two indexes undergo unidirectional and significant changes in the animals from all groups studied. There is a reduction of thrombocyte number and 7 5Se-M incorporation in the first group. This allows us to accept that there is a thrombocyto-poiesis suppression in recipients probably due to the reduced plasma thrombocyto-poietin activity. In our opinion it results from the realized negative feed-back me­chanism (7) in I P + T P treated donors as far as we previously reported that I P alone induced a significant thrombocyte count increase (2). The fact that I P + T P treatment of donors (9) does not cause thrombocyte countan chges probably also contributes to the low plasma thrombo-cytopoietin activity in the practical example. It is possible that the signifi­cantly increased megakaryocyte count in these experimental conditions has a de­finite importance for the inhibition of thrombocytopoietin biogenesis, i . e. in the mechanism of negative feed-back, too (9) thus stimulating the production of thrombocytopoiesis inhibitor. A l l that could reduce total plasma thrombocy­topoietin activity illustrated by the di­minution of thrombocyte count and 7 5Se-M incorporation in newly formed thrombocytes in recipients-mice. The significant increase of thrombo­cyte count and 7 5Se-M incorporation in newly formed thrombocytes in animals tested with plasma from P R + T P trea­ted donors is a sure sign of an enhanced thrombocytopoietin activity. This con­clusion confirms directly our previous investigations that in such donors there is a strongly expressed thrombocyte count reduction, a significant young form count increase, and a considerable decrease of the number of mature forms of the megakaryocyte line (9). We suppose that the increased plasma thrombocyto­poietin activity ^results not only from the thrombocytopenic thromboplastin ac­tion (5, 7) but also mainly from the capacity of the non-selective P-adrenoblo-cker P R to reduce significantly independently thrombocyte count in so far as plasma from donors injected with saline+TP induces a less expressed thrombo­cyte count increase (30.63 per cent). It allows us to suggest that there is a summa­tion of two influences reducing thrombocyte count — that of PR and that of T P . Thus, in the mechanism of negative feed-back thrombocytopoietin biosyn­thesis can be stimulated. Our results obtained allow us to presume that non-selective p-adrenergic influences modify additionally plasma thrombocytopoietin activity in condition of an acute thrombocytopenia, p-adrenostimulation with I P reduces but P-adre IP + TP PR-tJP F i g . 2 . T h r o m b o c y t o p o i e t i n a c t i v i t y of do­nor ' s p lasma — ra ts treated w i t h (3-adrener-gic agen ts and thromboplastin de termined by the percen tage of incorpora ted ? 5 S e M i n newly- fo rmed thrombocytes i n r ec ip ien ts -mice . Da ta are presen ted as x — S x . P e r c e n ­tage difference is c a l c u l a t e d towards the controls noblokade with P R increases considerably plasma thrombocytopoietin activity. 54 N . Negrev It seems most probably that beta-adrenoceptors play a regulatory role in throm­bocytopoietin production. Therefore, modulation effect of β-adrenoceptors on plasma thrombocytopoietin activity is present in conditions of an acute throm­bocytopenia, too. R E F E R E N C E S 1. H е г p е в , H . , Т . Г а н ч е в . В : Н а ц . к о н г р . х е м а т о л . и к р ъ в о п р е л и в . , I I I . С о ф и я , 1983 г . Р е з ю м е т а , 78 . — 2. Н е г р е в , Н . , Т . Г а н ч е в . Г е м а т о л . и т р а н с ф у -зиол . , 1986 (под п е ч а т ) . — 3. Н е г р е в , Н . , Т . Г а н ч е в . Г е м а т о л . и т р а н с ф у з и о л . , 1986 (под п е ч а т ) . — 4 . Т о д о р о в , Й . К л и н и ч н и л а б о р а т о р н и и з с л е д в а н и я в д е т с к а т а в ъ з р а с т . С . , М е д . и ф и з к . , 1962. — 5. G i г a u d о, L . А . , А . P . D а 1 m a s s о . T h r o m b . Haemost. , 39, 1978, No 3, 7 3 3 - 7 4 2 . - 6. H a r k е r , L . A . A m e r . J . P h y s i o l . , 218, 1970, 1376—1380. — 7. К е 1 е m е n , Е . , D . L е h о с z k у , I . С s е r h a t i , F . K r i z s a . A c t a Haemato l . , 29, 1963, No 1, 16—26. — 8. N а k е f f, A . , K . R о о z е n d а a l . A c t a Haemato l . , 54, 1975, 340—344. — 9. N е g r е v , N . S c r . . S c i . Med. , 22, 1985, 44—48. — 10 . О d e l l , J . Т . , I . R . M u r p h y . Blood, 44, 1974, 147—156. — 1 1 . P е n i n g t о n , , D . G . B r i t . M e d . J . , 4, 1969, 782—784. — 12. P е n i n g t о n , D . G . B r i t . Med . J . , 1 , 1970, 606—608. — 13. S h r е i n е r, D . , J . L e v i n . J . C l i n . Inves t . , 49 , 1970, 1 7 0 9 -1713. И С С Л Е Д О В А Н И Е Э Ф Ф Е К Т О В Н Е С Е Л Е К Т И В Н О Г О Б Е Т А - А Д Р Е Н Е Р Г И Ч Е С К О Г О В О З Д Е Й С Т В И Я Н А П Л А З М Е Н Н У Ю Т Р О М Б О Ц И Г О П О Э Т И Н О В У Ю А К Т И В Н О С Т Ц У К Р Ы С В У С Л О В И Я Х ОСТРОЙ Т Р О М Б О Ц И Т О П Е Н И И Н . Негрев Р Е 3 Ю М Е И с с л е д о в а н ы и з м е н е н и я ч и с л а т р о м б о ц и т о в и в к л ю ч е н и е 7 5 С е л е н о м е т и о н и н а ( 7 5 S e - M ) в н о в о о б р а з о в а н н ы е "тромбоциты у р е ц и п и е н т о в - м ы ш е й под в л и я н и е м п л а з м ы д о н о р о в - к р ы с . У с т а н а в л и в а е т с я , что у р е ц и п и е н т о в , к о т о р ы м в в о д и л а с ь п л а з м а д о н о р о в , и н ъ е ц и р о ­в а н н ы х и з о п р е н а л и н г и д р о х л о р и д о м ( н е с е л е к т и в н ы м б е т а - с т и м у л я т о р о м ) в с о ч е т а н и и с т р о м б о п л а с т и н о м , ч и с л о т р о м б о ц и т о в у м е н ь ш а е т с я на 18,96 % ( р < 0 , 0 0 2 ) , а в к л ю ч е н и е 7 5 с е -л е н о м е т и о н и н а п р и в о д и т к у м е н ь ш е н и ю ч и с л а т р о м б о ц и т о в на 53,48 % ( р < 0 , 0 0 1 ) . Р е ц и п и ­е н т ы , т е с т и р о в а н н ы е п л а з м о й д о н о р о в , и н ъ е ц и р о в а н н ы х п р о п р а н о л о л г и д р о х л о р и д о м (не ­с е л е к т и в н ы м б е т а - а д р е н о б л о к е р о м ) в с о ч е т а н и и с т р о м б о п л а с т и н о м , х а р а к т е р и з у ю т с я н а ­л и ч и е м з н а ч и м о г о у в е л и ч е н и я к а к ч и с л а т р о м б о ц и т о в (95,09 % , ( р < 0 , 0 0 1 ) , т а к и в к л ю ч е ­н и я 7 5 S e - M — 75,41 % ( р < 0 , 0 0 1 ) . К о н т р о л ь н а я г р у п п а р е ц и п и е н т о в , т е с т и р о в а н н ы х п л а з ­мой д о н о р о в , и н ъ е ц и р о в а н н ы х ф и з и о л о г и ч е с к о й с ы в о р о т к о й в с о ч е т а н и и с т р о м б о п л а с т и ­ном , п о к а з ы в а е т у в е л и ч е н и е ч и с л а т р о м б о ц и т о в на 30,63 % ( р < 0 , 0 0 1 ) . А в т о р с ч и т а е т , что б е т а - а д р е н е р г и ч е с к и е в о з д е й с т в и я д о п о л н и т е л ь н о и з м е н я ю т т р о м б о -Ц и т о п о э т и н о в у ю а к т и в н о с т ь п л а з м ы в у с л о в и я х о с т р о й т р о м б о ц и т о п е н и и , п р и ч е м б е т а - а д р е -н о с т и м у л я ц и я и з о п р е н а л и н о м в ы з ы в а е т п о н и ж е н и е , а б е т а - а д р е н о б л о к а д а п р о п р а н о л о л о м — с и л ь н о е п о в ы ш е н и е п л а з м е н н о й т р о м б о ц и т о п о э т и н о в о й а к т и в н о с т и . 

STUDY OF THE EFFECTS OF NON-SELECTIVE BETA-ADRENERGIC TREATMENT ON PLASMA THROMBOCYTOPOIETIN ACTIVITY [IN RATS IN CONDITIONS OF AN ACUTE THROMBOCYTOPENIA

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STUDY OF THE EFFECTS OF NON-SELECTIVE BETA-ADRENERGIC TREATMENT ON PLASMA THROMBOCYTOPOIETIN ACTIVITY [IN RATS IN CONDITIONS OF AN ACUTE THROMBOCYTOPENIA

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