ABSTRACT
Ovarian cancer （OC） is one of the three major gynecological malignancies. Due to its insidious onset at the early stage，most of OC patients are diagnosed at the advanced stage， making it become one of the most deadly gynecological tumors and thus a hot topic in the field of gynecology and oncology. Guizhi Fulingwan is a classic Chinese herbal compound derived from Synopsis of Golden Chamber （《金匮要略》） for the treatment of abdominal mass in women on account of its efficacy in resolving stasis， generating new blood， and eliminating mass. The articles concerning the treatment of OC with Guizhi Fulingwan were searched from such databases as China National Knowledge Infrastructure （CNKI）， PubMed，Wanfang Data Knowledge Service Platform， and Chongqing Weipu Database for Chinese Technical Periodicals （VIP） and collated for expounding its action mechanisms， in order to provide ideas for further research on its pharmacological effects，clinical application， and promotion. Clinically，Guizhi Fulingwan has been proved to control the growth of myoma，correct serological indexes，enhance chemotherapy sensitivity and anti-tumor immunity，reduce postoperative recurrence rate， and improve the quality of life of patients. As revealed by experimental research，Guizhi Fulingwan alleviates the pathological state of animal and cell models by promoting mitochondrial apoptosis and tumor immunity，inhibiting vascular factors，inducing cell cycle arrest， and reversing multidrug resistance. Guizhi Fulingwan exerts a certain therapeutic effect on OC through multi-target and multi-channel mechanisms， reflecting the advantages of traditional Chinese medicine in treating OC.
ABSTRACT
Objective:To investigate the effect of Guizhi Fulingwan on ovulation dysfunction in rats with polycystic ovary syndrome with insulin resistance （PCOS-IR） induced by letrazole combined with high fat emulsion. Method:A total of 72 female SD rats were randomly divided into control group， model group， metformin group and Guizhi Fulingwan low， medium and high dose groups， with 12 rats in each group. Except for control group， rats were given letrozole 0.001g·kg<sup>-1</sup> combined with high-fat emulsion 15 mL·kg<sup>-1</sup> for 21 consecutive days to establish model of PCOS-IR. Guizhi Fulingwan low， medium and high-dose groups were administrated with Guizhi Fulingwan 0.31， 0.62， 1.24 g·kg<sup>-1</sup> respectively， metformin group was administrated with metformin 0.27 g·kg<sup>-1</sup>， control group and model group were administrated with 12 mL·kg<sup>-1</sup> of normal saline daily for 30 days. Hematoxylin-eosin（HE） staining was used to observe ovarian tissue pathology morphology， and enzyme-linked immunoassay method （ELISA） was used to detect serum follicle stimulating hormone （FSH）， luteinizing hormone （LH）， testosterone （T）， fasting insulin （FINS） level，and LH/FSH and insulin resistance index （HOMA-IR） were calculated． Western blot was used to detect the expression levels of autophagy key molecular Atg6 yeast homologue （Beclin-1）， autophagy related gene 5（Atg5）， microtubule associated protein light chain 3 （LC3） Ⅱ proteins in the phosphatidylinositol 3-kinase/protein kinase B/rapamycin target protein （PI3K/Akt/mTOR） signaling pathway and autophagy related indicators in rat ovarian tissue. Beclin-1 and LC3Ⅱ protein expressions were detected by immunohistochemistry （IHC）. Result:Compared with control group， the thickness of follicles and follicular granulosa cells in the ovary of the model group also decreased， and the number of corpus luteum significantly decreased， while the white membrane thickness of the ovary increased， and the number of atresia follicles and cystic dilatation follicles increased significantly. Serum T， LH， LH/FSH， FINS， FINS， HOMA-IR were significantly increased （<italic>P</italic><0.01）. Phosphorylated （p） -PI3K， p-Akt， and p-mTOR proteins in ovarian tissue were all decreased （<italic>P</italic><0.05，<italic>P</italic><0.01）. The relative expression levels of autophagy-related protein LC3Ⅱ and Beclin-1 were significantly increased （<italic>P</italic><0.05，<italic>P</italic><0.01）. Compared with model group， the number of follicles in the low， medium and high dose Guizhi Fulingwan group and the metformin group decreased， the number of follicles in atresia and atresia increased， and the follicular granulosa cell layer thickness increased. Serum T， LH， LH/FSH， FINS and HOMA-IR of Guizhi Fulingwan group were significantly decreased （<italic>P</italic><0.05，<italic>P</italic><0.01）， and serum FINS and HOMA-IR of metformin group were significantly decreased （<italic>P</italic><0.01）. The expressions of p-PI3K， p-Akt， and p-mTOR proteins were increased （<italic>P</italic><0.05，<italic>P</italic><0.01）. The expression levels of LC3Ⅱ， Atg5 and Beclin-1 in the medium and high dose groups were significantly decreased （<italic>P</italic><0.01）. Conclusion:Guizhi Fulingwan can activate the PI3K/Akt/mTOR signaling pathway of granular cells， inhibit excessive autophagy of granular cells， improve ovarian function and insulin resistance， and restore ovulation， and the effect is better with high dose.
ABSTRACT
Objective:To investigate the effect of Guizhi Fulingwan on autophagy of ovarian granulosa cells in mice with polycystic ovary syndrome （PCOS）. Method:Twenty SD mice were randomized into a normal group （<italic>n</italic>=10） and a PCOS model group （<italic>n</italic>=10）， followed by PCOS modeling and <italic>in vitro</italic> culture of extracted ovarian granulosa cells. The ovarian granulosa cells of normal mice were classified into the control group and treated with 10% blank serum while those of PCOS mice into the experimental groups and with 10% Guizhi Fulingwan-containing serum at different concentrations （17.6， 35.1， 70.2 mg·kg<sup>-1</sup>） and 10% metformin-containing serum （25 mg·kg<sup>-1</sup>）， respectively， for 72 h. During the modeling， the changes in mouse body weight were measured. After modeling， the ovarian morphology was observed by microscopy， and the fasting blood glucose （FBG） was measured by Roche glucometer. Following the detection of fasting insulin （FI） and testosterone （T） levels by radioimmunoassay， the proliferation of ovarian granulosa cells was determined using cell counting kit-8 （CCK-8） to figure out the maximal dose of drug-containing serum that did not obviously affect the cell viability for subsequent assay. The autophagy of ovarian granulosa cells was examined by flow cytometry， and the protein expression levels of intracellular microtubule-associated protein 1 light chain 3Ⅰ （LC3Ⅰ）， LC3Ⅱ， Beclin1， and p62 were assayed by Western blott. Result:Compared with the blank group， the model group showed increased body weight and elevated FI， FBG， and T levels （<italic>P</italic><0.05，<italic>P</italic><0.01）， indicating the successful modeling of PCOS mice. Flow cytometric assay proved that the incubation with 10% Guizhi Fulingwan serum-containing medium resulted in a decline of autophagy （<italic>P</italic><0.05）. As demonstrated by Western blot assay results， the protein expression levels of Beclin1 and LC3 Ⅱ/Ⅰ in the model group increased significantly as compared with those of the blank group， whereas the expression level of p62 decreased significantly （<italic>P</italic><0.05，<italic>P</italic><0.01）. Compared with the model group， the medium- and high-dose Guizhi Fulingwan groups exhibited significantly down-regulated Beclin1 and LC3 Ⅱ/Ⅰ levels but remarkably up-regulated p62 （<italic>P</italic><0.05，<italic>P</italic><0.01）. Conclusion:Guizhi Fulingwan inhibits the autophagy of ovarian granulosa cells by down-regulating the protein expression levels of Beclin1 and LC3 Ⅱ/Ⅰ.
ABSTRACT
Objective:To explore the molecular mechanism of modified Guizhi Fulingwan in rats with uterine fibroids. Method:Seventy-two female adult SD rats of SPF grade were randomly divided into a model group， a normal group， and a preventive administration group. The model group and preventive administration group were established by estrogen and progestin loading method. After successful modeling， the rats in the model group were randomly divided into a western medicine group （mifepristone）， the high-dose traditional Chinese medicine（TCM） group， and a low-dose TCM group. All the rats were dosing as required once a day for 28 consecutive days. Hematoxylin-eosin（HE）staining was used to observe the morphological changes of the uterus. The micRNA gene chip was used to detect the expression profile of uterine micRNA gene. Differential expressions of micRNA were screened by bioinformatics methods. Gene function enrichment was used to predict the possible signaling pathways in rats with uterine fibroids by modified Guizhi Fulingwan. Result:Compared with the normal group， microRNA of the model group was 1 up-regulated and 9 down-regulated. Compared with the model group， microRNA of the high-dose group of TCM group was 2 up-regulated and 1 down-regulated， in the preventive administration group， 9 was up-regulated and 2 was down-regulated. Gene function enrichment analysis indicated that four signaling pathways were closely related to uterine fibroids. They were mitogen-activated protein kinase （MAPK） signaling pathway， Wnt signaling pathway， mammalian rapamycin target protein （mTOR） signaling pathway and vascular endothelial cell growth factor （VEGF） signaling pathway. Conclusion:Modified Guizhi Fulingwan affected the expression profile of micRNA in rat model of uterine fibroids induced by estrogen and progesterone， suggesting that modified Guizhi Fulingwan may involve in a variety of biological processes such as signal transduction and gene regulation in the treatment of uterine fibroids.