| Objectives:Polycystic ovary syndrome(PCOS)is the most common metabolic dysfunction and endocrine abnormality in women of reproductive age,its incidence rate is 5%~10%.persistent anovulation,hyperandrogenism and Insulin resistance(IR)are important feature.IR plays an important role in the pathogenesis of PCOS,,about 50-70% patients with PCOS are having varing dedrees of insulin resistance,but the mechanism is not very unclear.The aim of this study was to o examine the relationship between Pi3k/Akt and PCOS-IR by establishing the PCOS-IR rat model with the female SD rats by high-fat diet and cerazole,To further understand the role of metformin on express of Pi3k/Akt in PCOS rats.Methods:1 Experimental animals and groups1.1Experimental animals 6-week-old(185±10g)female Spragu Dawley rats,Purchased from the Guangdong Provincial Animal Center.1.2 Groups The normal control group:8 SD rats;The model group 8,group:8 SD rats;The metformin group 8 SD rats,The metformin + LY294002 group :8 SD rats.2.Methods2.1 The PCOS-IR rats model received with letrozole(1mg/kg/d)and high fat diet for 30 days.The rats were random Iy divided into four groups:Giving metformin 3mg/kg to the metformin group;metformin 3mg/kg and LY294002 totail vein injection was given to Metformin+ blockade group.distilled water was given for an equivalent length of time to rats in the normal control group and model control group.2.2 To observe emotional cycle by vaginal cell smear;Measured the weight of rats and ovaries.2.3 Levels of rat T,E2,LH,FSH and FINS concentrations in serum were measured with rat T,E2,LH,FSH and FINS enzyme-linked immunosorbent assay kits according to the manufacturer’s protocol.FPG and FINS used to evaluate the insulin resistance index(HOMA-IR).2.4 four groups of rats ovarian morphological changes were observed by HE stain2.5 Intraperitoneal glucose tolerance test to to assess insulin resistance.2.6.The transcription of Pi3 k m RNA and Akt m RNA in ovarian and liver tissue w ere measured by semi-quantitative reverse transcription polymerase chain reaction(RT-PC R)2.7The expression of Pi3 k and Akt protein in ovarian and liver tissue was observed and analyzed by Western-blotting.Results:1.the weight of rats and ovaries in model group was significantly higher than that i n the normal group(320.51±19.08 vs 246.60±10.21、88.44±7.01 vs 66.87±7.53,P<0.01).The serum values of T(4.07±3.41 vs 0.65±0.14)、LH(12.88±14.0 vs 0.88±0.35)and LH / FSH were significantly higher in model group than those in normal group,P<0.01.The serum values of FSH were lower than those in the normal group(2.53 ± 1.25 vs 5.46 ± 2.65,P <0.05).The serum level of E2 in the model group was higher than that in the normal group,but no significant statistically difference could be found.The HE sta in of ovarian slices showed that the numbers of follicular cysts and atretic follicles were increased,and the developing follicles,corpus lutea(CL),granulosa cells(GCs)were si gnificantly diminished in the model group;The normal group can be seen more complete corpora lutea and different levels of follicles with thicken layer of GCs.The level FPG in the model group was significantly higher than that of the normal group(6.84 ± 0.17vs4.69 ± 0.36,P <0.01).The level of FPG of two groups reached the peak at 15 min afte r the glucose load test,and the glucose level in the model group was significantly lower than that in the normal group(8.32 ± 0.09 mmol / L vs 7.72 ± 0.22 mmol / L,P<0.05).There was no significant decrease in glucose level in the model group at 120 min after glucose loading,significantly higher than that of the normal group(8.18±2.23mmol/Lvs5.14±0.43mmol/L,P〈0.05).The area under the curve of glucose in the model group wa s significantly increased than that in the normal group(24.37±0.66mmol/Lvs12.94±1.39mmol/L,P<0.05).2.Compared with the model group,The serum values of T,LH,FBG and FINS were significantly decreased after treatment with metformin.Given LY294002 intervention can be blocked the improvement of metformin on reproductive hormone disorders and IR.The expression of Pi3 k m RNA in liver and ovarian tissue of rats was significantly lower in the model group than that in normal group(0.59±0.09 vs 1.03±0.21;0.44±0.07 vs 1.01±0.17,P<0.05).The expression of Akt m RNA was the same as above(0.57±0.21 vs 1.01±0.13;0.57±0.13 vs1.01±0.26,P<0.05).The expression of Pi3 k and Akt m RNA in the liver and ovarian tissues were significantly increased after the rats treated with metformin(0.92 ±0.23 vs 0.59±0.090.82±0.13 vs 0.44±0.07,P<0.05)and(0.74±0.10 vs 0.57±0.21,P<0.05;0.81±0.34 vs 0.57±0.13,P<0.01).LY294002 can block the expression of Pi3 k and Akt m RNA in the liver and ovarian tissue.The protein expression of four groups rat of Pi3 k and Akt in the liver and ovarian tissues was similar to that of gene.Conclusions1.The rat model induced by letrozole and high fat diet was proved to be an ideal PCOS-IR animal model with similar pathological-histology,hormonal changes to those of PCOS patients and IR.2.Metformin can improve ovarian polycystic changes and insulin resistance of pcos.3.Metformin improve the insulin resistance of ovarian and liver tissue may through PI3 K /Akt. |
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