Effects Of Bu-Shen-Yi-Qi Formula On The Growth And Apoptosis Of Human First Trimester Trophoblasts

Spontaneous abortion is one of the most common complications of pregnancy. Between 15 and 25% of clinically recognized pregnancies culminate in spontaneous abortions. According to the theory of "Kidney is the commander of reproduction", we established the Yi-Qi formula of replenishing kidney. Bu-Shen-Yi-Qi formula was proved to be an effective recipe in abortion cases. Serial clinical investigations and animal studies showed that Bu-Shen-Yi-Qi formula protected pregnancy by regulating endocrine-immune disturbance in vivo. As a special endocrine and immunological organ in pregnancy, placenta is essential in pregnancy and for fetal growth. However, the effect of Bu-Shen-Yi-Qi formula on placenta is not wellkown. The functional and structural unit of the human palcenta is chorionic villi. Cytotrophoblasts (CTs) are key stem cells in villi development. Herbs have complicate components, and only the elements in blood perform herbs' funtion, so blood serum administered with herbs is the real "preparation" which really works. The present study will investigate the regulation of Bu-Shen-Yi-Qi formula on the growth and apoptosis of human first trimester trophoblasts by seropharmacology.Section I Effects of Bu-Shen-Yi-Qi Formula on the Growth of Human First Trimester TrophoblastsI The Isolation and Differentiation of Human First Trimester Trophoblasts[Purpose] To isolate human first trimester cytotrophoblasts and discuss their differentiation in vitro.[Methods] We isolated human first trimester cytotrophoblasts using different digestive conditions. After a 24h culture in vitro, cells morphology was observed by discrepancy microscopy. The shapes of cells stained by Giemsa dye were also analyzed under the light microscopy. Immunocytochemistry methods were used to determine the expression of the specialized protein. Cells surface microstructures were observed byscanning electron microscopy and the invasive ability of extravillous cytotrophoblasts were evaluated by Transwell invasive assay.[Results] (1) The cell populations were obtained by low and once enzymatic digestion without vibration. These cells were characterized by large, round nuleus and mutiple nucleolus with less cytoplasm. Cells aggregated but never fused in matrigel or rat-tail gel. Immunocytochemistry proved that this type of cells expressed cytokeratin 7 but not vimentin. Cells migrated and invaded in matrigel, penetrated through transwell PET membrane. The results showed that this kind of cells had the proliferative and invasive properties of EVCT. (2) The remaining villi were digested by sequential high enzymatic digestion. The cells obtained by this way were VCT characterized by small nucleus and extending cytoplasm, in contrast to EVCT, these cells surface exhibited numerous microvillous projections. VCTs fuse to form mutiplenucleated syncytiotrophoblasts which expressed hCG- but not vimentin. (3) Cells obtained by low and sequential enzymatic digestion were implantated in the plate covered by matrigel or rat-tail gel. These cells contained two types of cytotrophoblasts which expressed cytokeratin but not vimentin.[Conclusions] Different isolation methods might obtain different cytotrophoblasts with different characteristics and functions. Both EVCT and VCT play crucial role in the development of placenta and execution of its functions.II Effects of Bu-Shen-Yi-Qi Formula on the Growth of Human First Trimester Trophoblasts[Purpose] To investigate the effect of Bu-Shen-Yi-Qi formula on the growth of human first trimester trophoblasts by seropharmachology.[Methods] Drug-containing serum were collected by sacrificing the rats after a three-day administeration with herbs intragastricly. Using same approach, the blank control were preparated from rats administered with saline. Add 5%, 10%, 20% drug-contained serum respectively into cultured trophoblasts. Cells surface microstructure was observe by scanning electron microscopy after a 48h or 72h culture with different media. After a culture in media with drug-containing serum for 24h, 4...