Kovachev, D.

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Varna Medical University Press: Journals

E F F E C T OF PHYTOHAEMAGGLUTININ PHASEOLOSAXIN ON T H E L I F E C Y C L E OF PARAMECIUM CAUDATUM D. Kovachev Key-words: phytohaemagglutinin — Paramecium caudatum — life cycle — generation time The phytohaemagglutinin in ( P H A ) , isolated from the seeds of Phaseolus vulgaris has a well-known mitogenic effect upon lymphocytes in vi t ro (6). I t is established (4) that the reaction of these cells is an answer to an unspecific stimulus. I n fact, the P H A shows a biological ac t iv i ty on other cells, too — epi-thelic (7), Amoebae (3), ciliates (8), certain plants (5). According to the Zech's works (8) the P H A - P (Difco) diminishes the generation time of the ciliates Bur-saria truncatella and activates the growth of Stentor coeruleus in as "stationary phase" by preserving this effect for more than one generation. The object of the present work is to study the effect of the Bulgar ian P H A -Phaseolosaxin ( P H A - F ) on the life cycle of the infusoria Paramecium caudatum. We investigated the effect of P H A as wel l on the generation time of the infuso­ria in a logarithmic phase of growth as on isolated animals in a stationary phase. Material and methods We received the P H A from the Institute for infectious and parasitic disea­ses— Sofia where it is produced after Tznoneva's method (2). We isolated the monoclonal cell culture of Paramecium caudatum in a nutr i t ive medium contain­ing hay extract and inoculated wi th bacterial suspension (Aerobacter aeroge-nes, p H 6,8—7,0). The cul t ivat ion was carried out in a thermostat at 25 °C. The generation time was determined as follows: we isolated in concave glasses one animal from a culture in a logarithmic phase of growth. Immediately after the first cell division we laid one daughter cell in fresh nutr i t ive medium and the other one — in a medium wi th definite P H A concentration. We measured the time for cell division. We determined the generation time for every ind iv i ­dual by using this direct method. The generation time was calculated also accord­ing to the formula: -, where log. 10 A Е —generation time A — number of individuals in a given moment с — 0,3010 (constant of transition into log 10) T — time interval t i l l reaching A — number of individuals. We studied the effect of P H A on the onset of growth in animals from statio­nary culture by leaving the isolated individuals for 10 days and by eliminating t h V d i v i d e d ones. Two groups were formed from the physiologically depressed 38 D. Kovachev infusoria. The controls were transferred in a fresh nutr i t ive medium and the experimental ones — in a medium wi th definite P H A concentration. We noted the number of dividing cells from both groups in intervals of 12 hours. Results and discussion We followed up the generation time of individuals in the logarithmic phase treated wi th 2, 5, 10, 20, 50, 100, 200 and 500 \ig/m\ P H A - F to establish the op­t imal stimulating P H A concentration. The results are shown on table 1 where the generation time ( E ) is an arithmetic mean of a l l direct determinations and it is compared wi th that one of the control series ( E 0 ) . The ration E 0 / E charac­terizes clearly the stimulating effect of the P H A - F . T a b l e 1 Effect of different PHA concentrations on the generation time of infusoria in a logarithmic phase of growth PHA ,и/т1 Number of determinations Mean generation time (hours) Fo/E P control ( 0 ) experiment (E) 2 5 10 20 50 100 200 500 45 46 43 51 52 45 40 J 40 8,72=1=0,32 8,31-0,38 9,05=1=0,29 8,27—=0,37 8,64 =±=0,41 9,12=t=0,32 9,50=1=0,29 8,92=1=0,40 1 8,99=1=0,40 8,06=1=0,31 • 8,38=1=0,47 7,00=-0,42 6,31=1=0,35 7,18=1=0,37 13,01=1=0,42 toxic 0,97>0,5 1,03>0,5 1,08 >0,5 1,18<0,05 1,37<0,001 1,27<0,001 0,73<0,001 These data demonstrate that definite P H A concentrations have a well-ex­pressed stimulating effect on the infusoria by diminishing their generation time. The optimal concentration is 50 fig/ml but the higher concentrations are toxic indeed. Some authors report s imilar data concerning other protozoa (1,8). The generation time of the animals treated wi th 50 ixg/ml P H A shows that the stimu­lating action retains during more than 48 hours. The ratio E 0 / E is mean 1,41. T h i s fact in concordance wi th Zech's experiments (8) and it could be considered as a characteristic peculiarity of P H A biological action. The table 2 illustrates the possibility of P H A to shorten the term in which the stationary culture leaves the physiological depression. I t shows the number of divided cells and their percentage of the total amount. The majority of experimental individuals divide wi th in the interval between 36 and 48 hours. Th i s maximum is reached in the control series wi th in the inter­v a l between 60 and 72 hours. The whole number of treated cells divide t i l l 120 hours while in the control group 15 individuals retain yet non-divided during this period. T h e present experiments demonstrate that the P H A - F concentrations of 50 ixg/ml exert a stimulating effect on the life cycle of Paramecium caudatum by shortening the generation time and diminishing the term in which the culture leaves the stationary phase. Our data support the hypothesis for the unspecific character of P H A on the treated cells. Effect of phytohaemagglutinin phaseolosaxin on the. 39 T a b l e 2 Effect of 50 jig/ml PHA on infusoria from stationary phase culture Hour Control — n = 7l Experiment — n = 74 divided % divided % 96 : 4 3 9 14 10 9 3 3 0 1,41 5,63 4,23 12,68 19,72 14,08 12,68 4,23 4,23 7 4 11 26 9 9 5 1 2 , 9,46 5,41 14,86 35,14 12,16 12,16 6,76 1,35 2,70 Total 56 78,88 74 100,0 R E F E R E N C E S 1. Ц в е т к о в а , A . , H . Я н к о в . Пробл. зарази, u паразит, болести, 1974, 2, 2 4 9 . - 2. Ц о н е в а , М. Авт. свидет. № 12008 от 20. I X . 1964 г. — З. A g r e l l , I . P . S Experim. Cell Res., 42, 1966, 403. — 4. I о а с h i m, H . L . Mature, 210, 1966, 219. — 5. N a g l , W. Planta, 106, 1972, 3, 296. — 6. N o w e ll, P . C . Cancer Res., 20. ВЛИЯНИЕ ФИГОГЕМАГЛЮГИНИНА ФАЗЕОЛОСАКСИНA НА ЖИЗНЕННЫЙ ЦИКЛ PARAMECIUM CAUDATUM Д . Ковачев Р Е З Ю М Е Исследовано стимулирующее действие болгарского препарата фитогемаглютинина Фазеолосаксина на жизненный цикл инфузорий. В концентрации 50 мкг/мл ФГА сокра­щает генерационное время индивидов в логарифмической фазе роста и укорачивает срок выхода культуры из физиологической депресии в стационарной фазе. Высокие концентра­ции ФГА оказывают токсичное действие на инфузории. 

EFFECT OF PHYTOHAEMAGGLUTININ PHASEOLOSAXIN ON THE LIFE CYCLE OF PARAMECIUM CAUDATUM

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EFFECT OF PHYTOHAEMAGGLUTININ PHASEOLOSAXIN ON THE LIFE CYCLE OF PARAMECIUM CAUDATUM

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