| Objective:In this thesis, the effect the liver and spleen core method on ovarian granulosa cell apoptosis by the mitochondrial pathways in mature high prolactin hyperlipidemia animal model is explored to support the mechanism that this disease can be cured from liver and spleen theory targeted in the ovaries. According to the experimental study of mature high prolactin hyperlipidemia rat model, this thesis clarifies the pathway to mature high prolactin hyperlipidemia, further enrich the study of pathogenesis on mature high prolactin hyperlipidemia. At the same time, this thesis provides a new theoretical basis for the mechanism and targets of high prolactin hyperlipidemia, and proposes a new method of treatment and the prescription of safe and effective to provide a scientific basis for Chinese medicine treatment of incurable diseases of the female.Methods:Process of making high prolactin hyperlipidemia animal model:According to preliminary pre-experiment reported in the literature more than ten kinds HPRL animal model of production and comparison of serum PRL levels, SD grade clean sexual maturity female rats were randomly divided into six groups:blank control group, model group, bromocriptine group, the modified Xiaoyaosan low, middle and high dose group, besides, metoclopramide hydrochloride ammonia was injected into mature high prolactin hyperlipidemia animal model in abdominal cavity. This experiment tested of serum PRL levels enzyme-linked immunosorbent assay determination, the expression of be1-2and bax in ovarian granulose cell by immunohistochemical method, mensurated the effect the prescription on the expression of cytopigment C and Caspase-3in ovarian granulosa cell by immunoblotting, determined ultrastructural changes of mitochondria in ovarian granulosa cells by electron microscopy. This thesis researched mitochondrial pathways of apoptosis in ovarian granulosa cell from different levels.Results:By observing the changes of ultrastructure of granulosa cell mitochondria under electron microscope, in model group, cell concaved and had much chipping; vacuolar substance could be seen around cell nucleus; there was nearly not any mitochondrium but a large number of fatty droplets in cytoplasm. In traditional Chinese medicine high dose group, bromocriptine group and blank control group, there were more granulosa cells with morphological rules and some mitochondria and a few fatty droplets in cytoplasm. Traditional Chinese medicine middle dose group took the second place. In Traditional Chinese medicine low dose group, granulosa cells had few quantities and bad shape and fewer organellas.Compared with blank control group, in model group, PRL levels increased(p>0.05), be1-2reduced obviously (P<0.01), bax increased obviously (P <0.01),bc1-2/bax reduced (P<0.01),both caspase-3and cytopigment C increased (P<0.01). Compared with modal group, in bromocriptine group, PRL reduced (p>0.05), be1-2recreased (p<0.05), the expression of bax, bcl-2/bax and cytochrome C had no statistical significance (p>0.05), caspase-3reduced (p<0.05); in traditional Chinese medicine high dose group, PRL reduced (p>0.05), bcl-2increased (p<0.01), bax reduced (p<0.01), bcl-2/bax increased (p<0.01), caspase-3and cytochrome C reduced (p<0.01); in traditional Chinese medicine middle dose group, PRL reduced (p>0.05), bcl-2increased (p<0.05), the expression of bax had no statistical significance (p>0.05), bcl-2/bax increased (p<0.05), caspase-3and cytochrome C had no statistical significance (p>0.05); in traditional Chinese medicine low dose group, PRL reduced (p>0.05), bc1-2increased (p<0.05), bax, bcl-2/bax, caspase-3and cytochrome C had no statistical significance (p>0.05).Conclusion:From the research of ultrastructure in this experimental result with electron microscope technique, this prescription plays important role in improving ultrastructure of granulosa cell mitochondria. This treatment can reduced PRL levels of model animal, increase the expression of bc1-2and bcl-2/bax in ovarian granulosa cell of model animal. On the contrary, it reduced the expression of bax, caspase-3and cytochrome C. The function of ovarian can be improved by holding down ovarian granulosa cell apoptosis with the help of mitochondria. |
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