Tumors Of Sanjie Dan ( Serum Containing ) Human Cervical Carcinoma Cell Line Induced By Experimental Study On Apoptosis Of Hela Cells

Objectives:1.To study HuaLiuSanJieDan in inhibiting proliferation on human cervical carcinoma cell line HeLa, inducing apoptosis of human cervical carcinoma cell line HeLa and the influence on cell cycle in virtro.2. To investigate the potential mechanisms of the effects of HuaLiuSanJieDan on human cervical carcinoma cell line HeLa.3.To observe the acute toxicity of HuaLiuSanJieDan.Methods:1. The pharmaceutical serum containing HuaLiuSanJieDan is made from the Traditional Chinese Medicine HuaLiuSanJieDan and the rabbits' serum. In terms of serologic pharmacological test, the Traditional Chinese Medicine is fed to the rabbits, then stripple blood from the rabbits and contracts the serum containing HuaLiuSanJieDan.2. Cell culture: The human cervical carcinoma cell line HeLa is cultured in RPMI-1640 medium supplemented with 10% heat-inactivated fetal calf serum at 37@ in 5% C02 in air.3.Add the different concentration pharmaceutical serum containing the TCM HuaLiuSanJieDan to human cervical carcinoma cell line HeLa. After 24 hours , the light microscopy is used to observe the morphological changes. MTT method is used to detect the OD index and to observe the inhibiting effect on cells, while the effect of HuaLiuSanJieDan on DNA synthesis and cell cycle in human cervical carcinoma cell line HeLa are investigated with the help of Flow Cytometry' (FCM).4.The mice are divided into five groups and the HuaLiuSanJieDan is given orally only once, the method of reformed Karber'method is used to determine the LD50 of the HuaLiuSanJieDan.Results:1.After treated with the pharmaceutical serum containing HuaLiuSanJieDan, the morphologic features of HeLa cells are changed. Their outcoats shrink, they become ball and they detacth from the culture wells. While the other groups hold their nomal shape and attach on the wells tightly.2. The pharmaceutical serum containing HuaLiuSanJieDan can inhibit the growth of HeLa cells. The inhibiting rates of the large medium and small concentration are 56. 10%, 48. 37% and 34. 31% respectively, obviously higher than those of the other groups (P<0. 05). Among the pharmaceutical serum containing HuaLiuSanJieDan groups, the inhibiting rates of the large concentration groups are significantly higher than those of the medium and the small concentration groups (P<0.05).3.Compared to the N.S group and the blank group, the apoptosis ofpercents with HuaLiuSanJieDan at moderate and large concentration are 21.58% and 13.32% respectively (P<0. 05). The percentage of cells at the moderate and large concentration in G0/G1 and S phase decrease, then increase in G2/M phase .At the same time, the cell cycle is arrested in G2/M phase to the different degrees. While the percentage of cells at the small concentration in G2/M and S phase decrease and increase in G0/G1 phase.4. The results of acute toxicity experiment indicate that the LDS0 of HuaLiuSanJieDan on mice is 10. llg/kg orally and the 95% convinced range of LD50 is 9.41 -10. 81g/kg.Conclusions:1.The pharmaceutical serum containing HuaLiuSanJieDan can inhibit the growthof human cervical carcinoma cell line HeLa. 2.The pharmaceutical serum containing HuaLiuSanJieDan at the moderate andlarge concentration can induce apoptosis of the HeLa cells. 3.The pharmaceutical serum containing HuaLiuSanJieDan can influence the cellcycle and the G2/M phase arrest may be one of the mechanisms ofHuaLiuSanJieDan-induced apoptosis.4. The apoptosis of HeLa cells induced by pharmaceutical serum containing HuaLiuSanJieDan are related to the concentration and the time. If the concentration is too low and the time is too short (2.5% 12h), It cannot induce the apoptosis of HeLa cells;If the concentration is too high and the time is too long (15% 36h), It can cause HeLa cells to die.5. The results of acute toxicity experiment indicate that the LD50 of HuaLiuSanJieDan on mice is 10. llg/kg orally and there is some toxic effect in HuaLiuSanJieDan.