| Malignant tumor, a main threat of people's health, has been the second causations of death. The death rate of hepatoma, just bellowing gastric cancer in country and lung cancer in city, has been the second death of cancers in our country. However the current therapies of liver cancer can't serve the purpose very well, it becomes more and more important to research and develop new anti-hepatoma drugs. Recently, the orientation of research of anti-tumor drugs has changed from single cytotoxic drugs to diversified drugs. In this field, Chinese traditional medicine (CTM), as the heritage of thousands years, attracts more and more attention with its remarkable curative effect on tumor. They have predominance in inducing apoptosis, cells communication and regulate immunity. Many CTMS have the ability to inducing apoptosis and improve immunity in tumor-bearing bodies in clinic and laboratory.Planted in Jiangsu ,Baishouwu ,the tuber of Cynanchum auriculatum Royle, is a traditional Chinese herbal medicine. It is sweet and bitter in taste, mildly warm in nature. It possesses the effects of nourishing the liver and kidney, supplementing essence and blood and prolonging life. Present studies suggested the extract of Baishouwu have multiple pharmacological actions, including anti-tumor activity, clearing away free radicals and enhancing immunity. The C21 mono-glucosides sterols (C21) are thought to be the main active component. Study has proved to have cytotoxic activity in vitro and anti-tumor effect on EAC,LEWIS and S180-bearing mice in vivo. In this study, the models of Heps tumor transplanted in mice were established, apoptosis and its immunity mechanism were investigated. The main research methods, contents and conclusions are as follows: 1 The effect on anti-tumor and immune organic indexTo investigate the holistic effect of the C21 sterols on hepatoma bearing organism, we construct mice transplanted tumor models, 24h after the last administration, the mice were put to death. The tumor growth inhibition rate (IR), spleen index (SI) and thymus index (TI) was calculated and tumor tissue samples were taken and examined by light microscopy. After treatment of the C21 steroidal, inhibitory effect on the transplanted Heps was observed on the tumor-bearing mice in a dose-dependent manner. The inhibitory rates are all over 30% in three repeated experiments. We found the morphology of tumor tissues in C21 groups had typical characters of necrosis under the light microscope. Spleen indices decreased and thymus indices rose in C21 groups. This indicated that the C21 sterols could restrain hepatoma and have immune protecting activity.2 Inducing apoptosis and its mechanismApoptosis, a form of cell death, is thought to play a very important role in the happening, development and perdition of tumor. Thus using drugs to induce apoptosis of tumor cell is probably a promising way of tumor therapy and it also provides a new clew to the treatment of hepatoma and other cancers. In this study tumor tissue samples were taken and examined under electron microscopy. The tumor cells were harvested and cell viability or apoptosis was analyzed by Acridine orange and Ethidium bromide (AO/EB) stain. B-cell lymphoma/leukemia-2 gene (bcl-2) in tumor cells was inspected by immunohistochemistry. Apoptosis induced by the C21 was observed under the electron microscope and this was confired with AO/EB. The number of gene protein expression of bcl-2 on tumor cells decreased in the C21 groups compared with model group (P<0.01). This indicated that inhibiting the excessive gene protein expression of bcl-2 for inducing apoptosis maybe one of the mechanisms of anti-tumor for the C21 steroidal.3 Immunity effect on macrophagesMacrophages, cells participating in immuno-modulation, play an important role in host nonspecific and specific immunological functions. The phagocytic ability of peritoneal macrophage to neutral red was detected to reflect holistic cellular immune level. The secretion of NO was detected with method of Nitrate reductase and tumor necrosis factor-alpha (TNF-α) were detected with ABC-ELISA. The result showed that C21 steroidal could significantly augment the peritoneal macrophage phagocytic ability to neutral red. NO and TNF-αwere elevated when compared with normal group. The secretion of NO is higher in 20mg·kg-1 than that in 40mg·kg-1, which differing from macrophage phagocytic ability. This study indicated that the C21 steroidal could improve nonspecific immunological functions on tumor-bearing bodies.4 Immune effect on spleen lymphocytesStudy showed that the immunity decreased in tumor-bearing bodies. Spleen is an important immune organ. In this research, T and B lymphocyte transformation capacity were tested by MTT assay. The secretion of interleukin-2 (IL-2) was detected with ABC-ELISA. Results showed that the proliferation of T, B cells and the secretion of IL-2 decreased after beared with Heps. After treated with C21, T, B cells proliferation was increased to normal level and IL-2 was elevated when compared with model group. This indicated C21can improve the immnunity protecting activity of spleen. |
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