The Risk Factors Of Insulin Resisitance In Polycystic Ovary Syndrome And The Study Of Metformin Intervention

The polycystic ovary syndrome (PCOS) is a heterogeneous disorder with widespread systemic manifestations affecting 5-10% of women of reproductive age. Theories of the pathophysiology of PCOS have implicated primary defects in hypothalamic-pituitary function, ovarian activity, and insulin action, howere, none of these can comfortably accommodate the multiple abnormalities associated with PCOS. PCOS impact on anovulation and infertility, as well as have association with metabolic syndrome, cardiovascular disease and somen cancer. There is no good method to prevent the disease, no direct surrogate to predict the long-term concequence effectively.PCOS is characterized as hyperinsulinemia and hyperandrogenism. The symptoms of PCOS include hirsutism, acne, menstrual disturbance, acanthosis nigricans, evidence of excessive androgen production, polycystic ovary, consequently, anovulation and infertility. The clinical and epidemiological studies have shown that the obese have more sever long-term concequence than non-obese. The heterogeneity and pathogenesis of the disease need further work.The peripheral insulin resistance in PCOS is uniquely due to a defect beyond the activation of the receptor kinase, namely, reduced tyrosine autophosphorylation of the insulin receptor. And inflammatory factors contribute partially to insulin resistance. The biguanide metformin is an insulin sensitizer that can reduce hyperglycemia, improve endocrine disturbance of polycystic ovary syndrome women.In this study we attempt to analysis the association of sex hormone binding globulin and total testosterone with insulin resistance of PCOS women, the value of SHBG and TT to predict the long-term health risk; to assess the circulating level of cytokine and chemokine via enzyme-linked immunosorbent assay(ELISA) in PCOS women; compare clinical characteristic, hormonal assays and the metabolic profiles before and after metformin therapy; to assess paraoxonase 1 activity after metformin therapy; to study autophosphorylation of pY1158 and pYpY1162/1163 as well as insulin receptor-βsubunit in PCOS women before and after metformin therapy.PartⅠThe Risk factors of Insulin Resistance in PCOS一. Inflammatory Factors and Chemokine in Serum of PCOS Women.Objective To assess the circulating levels of cytokines and chemokines in PCOS women. Methods 12 healthy women are served as control group, 24 PCOS women were separated into PCOS_nob and PCOS_ob according BMI. The circulating lever of TNF-α, C-reactive protein(CRP), MCP-1, IL-8 and RANTES were measured via ELISA. The difference among thre group were analysis by ANOVA and independ t test. Results The serum TNF-αlevel is higher between PCOS nob and control (26.36±2.88 vs. 15.08±9.60 pg/ml, P＜0.01), between PCOS_ob and control(33.21±6.59pg/ml vs. 15.08±9.60pg/ml, P＜0.01); No significantly difference CRP were found among control, PCOS_nob and PCOS_ob women(0.38±0.04,0.26±0.06和0.27±0.07μg/ml, P＞0.05); MCP-1 is higher in PCOS_nob women than in control group, but the difference has no statistic means(126.11±56.81 vs. 106.08±30.81 pg/ml, P＞0.05); MCP-1 level is higher in PCOS ob women than control group and the difference has significantly statistic means(183.85±40.02 vs. 106.08±30.81 pg/ml, P＜0.01); IL-8 were higher in PCOS_nob and PCOS_ob than control group and the difference has significantly statistic means(518.27±257.60 vs. 130.93±127.67, 731.52±503.00 vs. 130.93±127.67 pg/ml, P＜0.01); No difference of circulatinglevel of RANTES was found between control, PCOS_nob and PCOS_ob(41.75±10.54, 40.62±15.42, 47.93±15.65pg/ml, P＞0.05)。Conclusion The circulating level of inflammation factors shown on different type in PCOS: Higher level of TNF-αand IL-8 in both PCOS_nob and PCOS_ob, the circulating level of MCP-1 is higher in PCOS_ob women, no higher lever of CRP and RANTES in PCOS women.二,The predict effect of SHBG and TT in insulin resistance in PCOSObjective To analysis the correlation between SHBG, TT and insulin resistance and metabolic disturbance in PCOS. Methods 344 cases of PCOS and 100 cases of normal women were including in the study, SHBG and TT level were compared between PCOS and normol contrl, the association between SHBG, TT and human parameters, endocrine hormone, lipds and glucose profile are analysis. Taking a ROC curve between SHBG and IR and acquired a cut-off point, compare difference between group with a higher SHBG/TT and group with lower SHBG/TT. Results SHBG is lower in PCOS than normal control(114.03±88.45 vs 201.05±105.76mmol/L, P＜0.01) and TT is higher in PCOS than in normol control(0.81±0.28 vs 0.49±0.18ng/dl, P＜0.01). SHBG, BMI, WHR and TG have a inverse correlation with insulin resisitance, endocrine and metabolic abnormal is more sever in group with low SHBG, and TT only has a positive correlation with insulin resisitance. Conclusions Both TT and SHBG has a correlation with higher level of insulin in PCOS. SHBG is an only independent risk factor to predict insulin resistance in PCOS women.PartⅡThe Clinical Effection and Mechanism of Metformin on PCOS一. Clinical Study of Metformin Therapy on PCOSObjective This is a prospective, randomized control study to evaluate clinical effect of metformin in PCOS women. Methods 30 health women with regular cycles are served as control group(group 1), 140 PCOS women were separated in to 4 groups randomized after categorized as nob-PCOS and ob-PCOS according BMI: GROUP 2 (PCOS nob, n=30) and grop 3 (ob-PCOS_ob, n=40) were treatmented with medroxyprogesterone acetate, group 4 (PCOS_nob, n=30) and group (PCOS_ob, n=40) were treatmented with metformin. Hormonal, lipid and glucose profiles were study before and on the sixth month of therapy. Results No change were found of human parameter, endocrine hormone, lipid and glucose profile in group 2 and 3 after therapy. BMI was decreased in group 5 after therapy(28.35±4.11 vs. 26.65±3.99 kg/m~2, 0.90±0.04 vs. 0.88±0.04 kg/m~2, respectively, P＜0.01), F-G score decreased (7.46±3.42 vs. 5.41±2.43, P＜0.01); LH/FSH ratio decreased (1.78±0.67 vs. 1.39±0.58, P＜0.01); In both group 4 and group 5, TT is decrease (0.87±0.24 vs. 0.70±0.21ng/dl, 0.88±0.25 vs. 0.67±0.22ng/dl, P＜0.01), SHBG increase (97.61±61.74 vs. 151.38 ±78.74mmol/L, 71.26±48.11 vs. 104.22±66.81mmol/L, P＜0.01), and FAI decrease accordingly(5.99±6.66 vs. 2.25±1.61, 6.03±5.29 vs. 3.04±2.64, P＜0.01), also FINS decreased (7.66±3.51 vs. 5.06±2.10mmol/L, P＜0.01; 19.01±21.32 vs. 10.00±6.09mmol/L, P＜0.01), as well as IAUC decreased (173.49±84.01 vs. 145.57±60.35mmol. h/L, 338.33±199.53 vs. 201.74±106. 19mmol. h/L, P＜0.01), GAUC in group 5 are decreased (18.96±3.48 vs. 17.64±3.36mmol. h/L, P＜0.01), HOMA-IR are decreased in both group 4 and group 5 (1.40±0.69 vs 0.91±0.38, P＜0.01; 3.78±4.70 vs. 1.88±1.29, P＜0.05); Cholesterol decreased in group 5 (4.58±0.90 vs. 4.36±0.84mmol/L, P＜0.01) and triglycerides were decreased in group 4 (1.57±0.89 vs. 1.40±0.71mmol/L, P＜0.05), low-density lipoprotein decreased in group 4 and group 5 (2.73±0.57 vs. 2.33±0.43mmol/L, 2.95±0.57 vs. 2.60±0.52mmol/L, P＜0.01)。The spontaneously ovulation rate in group 4 is higher than group 5(66.67% vs. 40%), the difference has statistic means (P＜0.05)。Conclusion 6 months of metformin therapy can improve the disturbance of PCOS women in endocrine and metabolic profiles effectively.二. The Effect of Metformin on CytokinesObjective To assess the effect of metformin on circulating level of cytokines in PCOS women. Methods 12 healthy women with regular menstrual cycles were take as control group (group 1), 48 women with PCOS were divided into 4 groups according BMI: group 2 (nob—PCOS, n=12) and group 3 (ob-PCOS, n=12) were treatment with medroxyprogesterone acetate, group 4 (nob—PCOS, n=12) and group 5 (ob-PCOS, n=12) were treatment with metformin. The serum TNF-α,IL-8 and MCP-lwere measured before the treatment and on the sixth month of therapy. Result No significantly change of TNF-αwere found in group2 and group 3 (26.36±2.88 vs. 23.82±4.22pg/ml, 33.21±6.59 vs. 30.20±5.15pg/ml,respectively, P＞0.05), while a significantly change in group 4 and group 5 (29.01±6.67 vs. 23.58±4.17pg/ml, 35.58±8.03 vs. 28.61±6.42pg/ml, respectively, P＜0.05). The change of IL-8 in group 2 and group 3 have no significantly mean (518.27±257 .59 vs. 479.49±241.01pg/ml, 731.52±502.97 vs. 669.75±456.41pg/ml, respectively, P＞0.05), while decrease in group 4 and group 5 have significantly mean (541.49±218.30 vs. 266.32±87.14pg/ml, 720.65±485.84 vs. 197.06±88.71pg/ml, respectively, P＜0.01); Also no significantly of MCP-1 were found in group 2 and group 3(126.11±56.81 vs. 121.76±53.40, 183.85±40.02 vs. 157.68±60.59pg/ml, P＞0.05), while significantly change were found in group 4 and group 5 (128.28±22.93 vs. 105.02±30.06,197.42±80.39 vs. 122.60±58.13pg/ml, P＜0.01)。Conclusion 6 cycles of metformin therapy can decrease circulating level of TNF-α, IL-8 and MCP-1 in PCOS women.三. Tyrosine Autophosphorylation of Insulin Receptor of Red Blood Cell in PCOS Women and the Effect of MetforminObjective To detect autophosphorylation of pY1158, pYpY1162/1163 in PCOS women and the effect of metformin. Methods 8 healthy women with regular cycles served as control group, 8 women with PCOS whose BMI is less than 25kg/m~2 categorized as nob-PCOS group, 12 women with PCOS whose BMI is more than 25kg/m~2 categorized as ob-PCOS group. Both nob and ob group were treatmented with metformin for 3 months. Autophosphorylation of pY1158,pYpY1162/1163 and insulin receptor-βwere detected before and on the thirds months. Results The concentration of pY1158 in control group is higher than nob-PCOS group and ob-PCOS group (40.97±8.55 vs 28.53±9.24, 40.97±8.55 vs 25.70±13.81Units/ml, p＜0.05); The concentration of pY1158 is increased after insulin stimulation in three groups(40.97±8.55 vs 47.85±14.07 Units/ml, 28.53±9.24 vs 33.03±7.90 Units/ml, 25.70±13.81 vs 31.29±14.94 Units/ml, p＜0.05); Both of the concentration of pY1158 in nob-PCOS and ob-PCOS were increased after metformin therapy (28.53±9.25 vs 40.86±10.52 Units/ml, 25.70±13.81 vs 31.67±18.06 Units/ml,p＜0.01) and after metformin therapy in vivo and insulin stimulation in vitro (28.53±9.25 vs 50.07±12.74Units/ml, 25.70±13.81 vs 31.92±18.13 Units/ml, p＜0.01))。No difference of concentration of pY116/1163 were found between control group and nob-PCOS group, ob-PCOS group (13.45±1.33 vs. 14.38±1.72, 13.45±1.33 vs. 14.34±3.66Units/ml, respectively p＞0.05), No change were found in nob-PCOS and ob-PCOS group after insulin stimulation (13.45±1.33 vs 13.89± 1.30, 14.37±1.71 vs 15.25±2.08, 14.34±3.66 vs 15.05±1.94 Units/ml, p＞0.05); The concentration of pY116/1163 was increased in nob-PCOS women after metformin therapy, the difference has significantly statistic mean (14.37±1.71 vs 17.94±3.34, p＜0.01), it is also increased in ob-PCOS group and the difference has statistic means (14.34±3.66 vs 15.63±3.21 Units/ml, p＜0.05); The concentration of pYpY1162/1163 were further increased after metformin therapy in vivo and insulin stimulation invitro (14.37±1.71 vs 19.62±2.90, 14.34±3.66 vs 18.91±5.45 Units/ml, p＜0.01). No difference of insulin receptor-βwere found between control group and nob-PCOS, ob-PCOS group (1.89±0.33 vs 1.58±0.28, 1.89±0.33 vs 1.69±0.26ng/ml, p＞0.05); There is no change in concentration of IR-βin three groups after insulin stimulation (1.88±0.33 vs 1.75±0.31, 1.58±0.28 vs 1.43±0.28, 1.69±0.25 vs 1.68±0.71ng/ml, p＞0.05); Also no change are found in nob-PCOS and ob-PCOS group after metformin (1.58±0.28 vs 1.34±0.25, 1.69±0.25 vs 1.48±0.31ng/ml, p＞0.05) and metformin therapy in vivo and insulin stimulation in vitro (1.58±0.28 vs 1.60±0.25, 1.69±0.25 vs 1.52±0.51ng/ml, p＞0.05)。Conclusion Impaired tyrosine autophosphorylation in pY1158 perhapous contribute to insulin resistance in PCOS. The autophosphorylation of pY1158,pYpY116/1163 are increase after metformin therapy, no difference were found of IR-βbetween PCOS and control and no change were found after metformin therapy.四. The PON-1 Activity in Adolescence Girl with PCOS and the Effect of MetforminObjective To assess PON 1 activity in adolescence with PCOS and the therapy effect of metformin. Methords 15 non PCOS girl wjth regular cycles served as control(group 1), 30 purberty girls who diagnosis as PCOS were separated into 2 groups: group 2 (n=15) were treatmented with medroxyprogesterone, group 3 (n=15) were treatmented with metformin; The PON 1 activity were measured before the treatment and on the sixth months of the therapy. Result PON 1 activity in group 1 is higher than in group 2 (2.60±0.82 vs. 1.62±1.38 Units, P＜0.05) and group 3 (2.60±0.82 vs. 1.60±0.77 Units, P＜0.05). No significant change were found in group 2 after therapy (1.62±1.38 vs. 1.39±0.51 Units, P＞0.05), while in group 3, PON 1 were increased (1.60±0.77 vs. 2.52±0.92 Units, P＜0.01), the change have significantly statistic means. Result Serum PON 1 activity were decreased in puberty girl with PCOS and can be improved by metformin therapy.