The Expression And Related Mechanism Of Anti-M(u|¨)llerian Hormone In Polycystic Ovary Syndrome

Part1Relationship between Plasma Concentration of Anti-Mullerian Hormone and Clinical Index in Women with Polycystic Ovary SyndromeObjective To analyze interrelation between anti-Mullerian hormone (AMH) levels and body weight(BMI), hormone status and metabolic rate in women with polycystic ovary syndrome (PCOS). Methods Sixty women (30overweight, PCOS1group and30normal weight, PCOS2group) diagnosed as PCOS were enrolled and30women with normal period (15overweight and15normal weight non-PCOS.control1and control2respectively) were as control group. The body weight and height were measured and BMIwas calculated. The serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol, testosterone, adrenal medullary hormone (AMH), androgens, fasting glucose and fasting insulin were detected, and the homeostasis model of assess ment-insulin resistance (HOMA-IR) were caculated and compared between four groups. Results The levels of AMH, LH. LH/FSHP. testosterone, fasting glucose and fasting insulin and HOMA-IR were higher in PCOS group than those of control group(P<0.05). There was no significant difference in AMH level between PCOS1and PCOS2groups (P>0.05). The levels of LH and LH/FSH were lower in PCOS1than those of PCOS2group (P<0.05). The levels of fasting insulin and HOMA-IR were higher in PCOS1group than those of PCOS2group(P<0.05). There were no correlations in AMH and BMI levels in PCOS group (P>0.05). There were positive correlations in AMH and testosterone, fasting glucose and fasting insulin and HOMA-IR in PCOS1group, but a negative correlation between estradiol and FSH. A significant positive correlation was found between LH, LH/FSH, testosterone and AMH in PCOS2group(P<0.05). Conclusion There is a significant increase in plasma AMH in patients with PCOS. The plasma AMH level is not associated with BMI, which may have correlation with hormone imbalance and metabolic disorders.Part2The effect of metformin on AMH-androgen-insulin in women with PCOSObjective Evaluate the levels of AMH in insulin resistance (IR) women with PCOS before and after metformin therapy. To explore the effect of metformin in patients with PCOS, and the possible mechanism on AMH-androgen-insulin. Methods According to the part one, there are26cases with insulin resistance in PCOS1group (obese) and10cases in PCOS2group (non obesity).These patients were enrolled in the second part.All patients received500mg metformin,3times/day,for4months. Serum levels of AMH, sex hormones, insulin, blood glucose, and glucose tolerance test were measured after metformin therapy. The menstrual cycle in two groups were observed. Results The levels of AMH,testosterone,insulin and HOMA-IR were decrease in PCOS1group(obese) after treatment(P<0.05). There are on change in the levels of AMH,testosterone,insulin and HOMA-IR were decrease in PCOS2group(non obesity) after treatment(P>0.05).The recovery rate of menstruation was53.85%in PCOS1and10%in PCOS2, Is similar between the two groups was statistically significant (P<0.05). Conclusion In the presence of insulin resistance, the effect of metformin treatment in patients with obese PCOS is better than the patients non obesity. There are more obvious menstrual cycle improve.the improvement of insulin resistance and hyperandrogenism in Obese PCOS patients after metformin therapy, and the lever of AMH significantly decreased at the same time.Part3The influence of insulin and testosterone on AMH and its receptor protein expression and regulation of AMH promoter activity in ovarian granulosa cellObjective To Explore the influence of insulin and testosterone on AMH and its receptor protein expression and regulation of AMH promoter activity in ovarian granulosa cell. Methods Cell primary culture of SD rat ovarian granulosa, the microscopic morphology identification after HE staining. Cells in processing different can be divided into the following four groups:control group (Blank):pure particle normal training.Insulin(Insulin group):100ng/ml insulin treatment for48hours. Androgen (Androgen group):105mol/L testosterone treatment for48hours. Androgen and Insulin(Insulin+Androgen group):100ng/ml insulin and105mol/L testosterone treatment for48hours together. RT-PCR detection AMH and its receptor mRNA expression and Western blot detection AMH and its receptor protein expression. IHC staining test at the same time. According to the published literature or biology database retrieval AMH promoter sequences, and cloned into the fluorescein report carrier, clone mutant AMH promoter sequences at the same time, in the presence of insulin and testosterone, change detection of fluorescein. Results RT-PCR showed that AMH.AMHR2mRNA expression was increased in Insulin group, Androge group and Androgen+Insulin group compared with the control (P<0.05).Including Insulin+Androgen group increased more obviously, compared with Insulin group and Androge group (P<0.05).Western blot showed that AMH,AMHR2protein expression was enhanced in insulin group, Androge group and Androgen+Insulin group compared with control subjects(P<0.05). AMH protein expression increased more obviously in Insulin+Androgen group,compared with Androge group(P<0.05), AMHR2protein expression increased more obviously in Insulin+Androgen group, compared with Insulin group and Androge group(P<0.05).Immunohistochemical results showed that insulin and testosterone stimulation with Western blot test results are consistent. Dual luciferase report gene detection results show that the sequence of genetic testing in containing the AMH promoter regulation of Insulin group, Androge group and Androgen+Insulin group markedly improved AMH promoter activity, compared with control subjects (P<0.05). Which the AMH promoter activity of Insulin+Androgen group is more obvious, compared with Insulin group and Androge group (P<0.05). Conculsion Insulin and testosterone enhance the mRNA and protein expression of AMH in ovarian granulosa cell obviously, and affect the AMH gene promoter activity, thereby promoting AMH transcriptional expression in ovary granulosa cells,and synergistic effect between insuliln and testosterone were observed.