Efficiency of Chinese medicine Bushen Huatan formula for treatment of polycystic ovary syndrome in mice via regulating gut microbiota and PPARγ pathway. / è¡¥è‚¾åŒ–ç—°æ–¹é€šè¿‡è°ƒæŽ§è‚ é“èŒåŠPPARγ通路治疗多囊卵巢综合征的机制.

OBJECTIVES: To explore the effect and mechanism of Chinese medicine Bushen Huatan formula in treatment of polycystic ovary syndrome (PCOS). METHODS: Twenty-four SPF female C57BL/6J mice were randomly divided into 3 groups with 8 animals in each group. Control group was given drinking water ad libitum; PCOS was induced by giving letrozole gavage and high-fat diet in model group and treatment group; treatment group received Bushen Huatan formula suspension for 35 d. The sex hormone levels of mice were detected by enzyme-linked immunosorbent assay. Ovary morphology was observed under light microscope after hematoxylin and eosin staining. The feces in the colon of mice were collected, and the gut microbiota was detected by 16S rRNA sequencing. The short chain fatty acids were detected by gas chromatography-mas spectrometry. The expression of peroxisome proliferator activated receptor (PPARγ) was detected by immunohistochemistry. The mRNA expression of mucin-2, occludin-1, tight junction protein zonula occludens 1 (ZO-1) and PPARγ in intestinal epithelium were detected by realtime RT-PCR. The expression of inducible nitric oxide synthase (iNOS) and PPARγ was detected by Western blotting. RESULTS: Compared with the control group, the body weight, serum levels of follicle stimulating hormone, luteinizing hormone and testosterone in the model group were increased, and serum levels of estradiol were decreased (all P<0.01); the ovarian structure under light microscope was consistent with the characteristics of PCOS. Compared with the model group, the serum levels of sex hormone and ovarian structure in treatment group were improved. The overall structure of gut microbiota in PCOS model mice changed. Compared with control group, there were significantly reduced abundance of Firmicutes, and increased abundance of Verrucomicrobia, Proteobacteria and Actinobacteria inthe model group at phylum level (all P<0.05); there were significantly reduced abundance of Lactobacillus, and increased abundance of Akkermansia, Lachnoclostridium, Lactococcus and Eubacterium_coprostanoligenes at genus level (all P<0.05). The disordered condition of gut microbiota was significantly improved in treatment group. Compared with control group, the contents of acetic acid, propionic acid and butyric acid in feces of model group were significantly decreased (all P<0.05); while the contents of propionic acid and butyric acid in treatment group were significantly increased compared with model control group (both P<0.05). Compared with control group, the mRNA expression of ZO-1 and protein expression of iNOS in model group were significantly increased, and the protein expression of PPARγ and the mRNA expressions of mucin-2 and occludin-1 were significantly decreased (all P<0.05). Compared with model group, the mRNA expression of ZO-1 and protein expression of iNOS in treatment group were decreased, and the protein expression of PPARγ and the mRNA expressions of mucin-2 and occludin-1 were increased. CONCLUSIONS: PCOS induced by letrozole high-fat diet induces microflora imbalance in mice. Chinese medicine Bushen Huatan formula may increase the level of short chain fatty acid by regulating gut microbiota, thereby activating the intestinal PPARγ pathway and improving intestinal barrier function to act as a cure for PCOS.

Short-chain fatty acid-butyric acid ameliorates granulosa cells inflammation through regulating METTL3-mediated N6-methyladenosine modification of FOSL2 in polycystic ovarian syndrome.

Polycystic ovary syndrome (PCOS) is an endocrine and metabolic disorder characterized by chronic low-grade inflammation. Previous studies have demonstrated that the gut microbiome can affect the host tissue cells' mRNA N6-methyladenosine (m6A) modifications. This study aimed to understand the role of intestinal flora in ovarian cells inflammation by regulating mRNA m6A modification particularly the inflammatory state in PCOS. The gut microbiome composition of PCOS and Control groups was analyzed by 16S rRNA sequencing, and the short chain fatty acids were detected in patients' serum by mass spectrometry methods. The level of butyric acid was found to be decreased in the serum of the obese PCOS group (FAT) compared to other groups, and this was correlated with increased Streptococcaceae and decreased Rikenellaceae based on the Spearman's rank test. Additionally, we identified FOSL2 as a potential METTL3 target using RNA-seq and MeRIP-seq methodologies. Cellular experiments demonstrated that the addition of butyric acid led to a decrease in FOSL2 m6A methylation levels and mRNA expression by suppressing the expression of METTL3, an m6A methyltransferase. Additionally, NLRP3 protein expression and the expression of inflammatory cytokines (IL-6 and TNF-α) were downregulated in KGN cells. Butyric acid supplementation in obese PCOS mice improved ovarian function and decreased the expression of local inflammatory factors in the ovary. Taken together, the correlation between the gut microbiome and PCOS may unveil crucial mechanisms for the role of specific gut microbiota in the pathogenesis of PCOS. Furthermore, butyric acid may present new prospects for future PCOS treatments.

Serum metabolomics reveals metabolic profiling for women with hyperandrogenism and insulin resistance in polycystic ovary syndrome.

INTRODUCTION: Polycystic ovary syndrome (PCOS) is a heterogeneous endocrine disorder. Hyperandrogenism (HA) and insulin resistance (IR) are two important pathogenic factors. OBJECTIVE: We aimed to investigate the inherent disturbed metabolic profiles for women with HA or IR in PCOS as well as discover diagnostic biomarkers. METHODS: A total of 286 subjects were recruited for the study. They constituted the following groups: healthy women (C), those with HA (B1), those with IR but not obese (B2) and obese women with IR (B3) in PCOS. Nine cross-comparisons with PCOS were performed to characterize metabolic disturbances. Serum metabolomic profiles were determined by gas chromatography-mass spectrometry. RESULTS AND CONCLUSION: We found a total of 59 differential metabolites. 28 metabolites for B1 vs C, 32 for B2 vs C and 25 for B3 vs C were discovered. Among them, palmitic acid, cholesterol, myo-inositol, D-allose, 1,5-anhydro-D-sorbitol, 1-monopalmitin, 1-monostearin, glycerol 1-phosphate, malic acid and citric acid, were the common differential metabolites among B1 vs C, B2 vs C and B3 vs C, which related to biosynthesis of unsaturated fatty acids, citrate cycle etc. Besides, 9-biomarker panel can diagnose well between HA and IR in PCOS. They provided areas under the receiver operating characteristic curve of 0.8511 to 1.000 in the discovery phase, and predictive values of 90% to 92% in the validation set. The result indicated that the differential metabolites can reflect the underlying mechanism of PCOS and serve as biomarkers for complementary diagnosis of HA and IR in PCOS.

[Effect of Bushen Huatan Recipe on the Akt signal pathway in polycystic ovarian syndrome model rats with insulin resistance: an experimental research].

OBJECTIVE: To observe the effect of Bushen Huatan Recipe (BHR) on the Akt signal pathway in polycystic ovarian syndrome (PCOS) model rats with insulin resistance (IR). METHODS: Fifty Wistar female PCOS rats were randomly and equally divided into 5 groups, i.e., the control group, the model group, the low dose BHR group (5.406 g/kg), the medium dose BHR group (10.812 g/kg), and the high dose BHR group (21.624 g/kg), 10 in each group. Levels of fasting blood glucose (FBG) and insulin were detected to calculate homeostasis model assessment of insulin resistance (HOMA-IR). The expression of insulin receptor substrate 1 (IRS-1), glycogen synthetase kinase-3beta (GSK-3beta), glucose transporter 4 (GLUT-4), and peroxisome proliferator activated receptor (PPAR-gamma) mRNA were detected by RT-PCR. The expression of insulin signal transduction molecular kinase B (Akt) was detected by Western blot. RESULTS: Compared with the control group, HOMA-IR and the mRNA expression of PPAR-gamma mRNA significantly increased, the mRNA expression of GSK-3beta, GLUT-4, and IRS-1, protein expression of Akt and p-Akt significantly decreased in the model group (P < 0.05). Compared with the model group, HOMA-IR significantly decreased, the mRNA expression of GSK-3beta, GLUT-4, IRS-1, and Akt protein significantly increased in the high dose BHR group (P < 0.05, P < 0.01). The mRNA expression of p-Akt protein increased more obviously (P < 0.05, P < 0.01). mRNA expression of GSK- 3beta and GLUT-4 significantly increased (P < 0.05), while the mRNA expression of PPAR-gamma significantly decreased in the low and middle BHR groups (P < 0.05). The expression of p-Akt significantly increased in the low dose BHR group (P < 0.05). CONCLUSIONS: IR and abnormal insulin signal pathway existed in PCOS model rats. BHR could improve IR of PCOS rats, which was correlated with regulating protein expression of insulin signal transduction molecules.

[Effect of electro-acupuncture on clinical outcomes and ovarian hyperstimulation syndrome in in vitro fertilization and embryo transplantation].

OBJECTIVE: To observe the effect of electro-acupuncture (EA) on clinical outcomes and the occurrence of ovarian hyperstimulation syndrome (OHSS) in in vitro fertilization and embryo transplantation. METHODS: Totally 109 patients who routinely received in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) at Reproductive Center were assigned to the control group (56 cases) and the EA group (53 cases) according to even and odd-numbered date. Patients in the control group received controlled ovarian hyperstimulation (COH) referring to GnRH-a long protocol. On the basis of COH, those in the EA group received EA from the day of Gn injection to the day of embryo transfer. Estradiol (E2), vascular endothelial growth factor (VEGF), interleukin-6 (IL-6), and angiotensin (AT) II were measured in all patients on the day of hCG injection, the day of ovum pick up (OPU), and the day of embryo transfer (ET), respectively. The oocyte retrieval rate, good quality embryo rate, clinical pregnancy rate, the abortion rate, and the occurrence of OHSS were compared between the two groups. RESULTS: Compared with the control group, serum E2 levels on the day of OPU and the day of ET were significantly lower in the EA group (P < 0.05). On the day of OPU levels of VEGF and IL-6 also significantly decreased (P < 0.05). Serum levels of VEGF and IL-6 reached the highest line on the day of hCG in the two groups, and then showed a decreasing trend. Compared with the control group at the same time point, serum levels of VEGF and IL-6 obviously decreased on on the day of OPU, hCG, and ET (P < 0.05). The occurrence of OHSS and the canceling rate of transplant cycle were significantly lower in the EA group than in the control group (P < 0.05). CONCLUSIONS: EA, as an adjunctive therapy, could reduce the occurrence of OHSS in IVF. Besides, it did not decrease good embryo rates and pregnancy rates in IVF-ET, which might be associated with lowering local vascular permeability of ovaries.