不同剂量加减苍附导痰汤对肥胖型PCOS-IR大鼠胰岛素抵抗的影响

Effects of modified Cangfu Daotan decoction of difference dose on insulin resistance in rats of polycystic ovary syndrome with insulin resistance

目的建立多囊卵巢综合征伴胰岛素抵抗(polycystic ovary syndrome with insulin resistance,PCOS-IR)动物模型,观察加减苍附导痰汤高、中、低剂量组对肥胖型多囊卵巢综合征(polycystic ovary syndrome,PCOS)大鼠胰岛素抵抗和血清性激素水平的影响,以及卵巢组织及骨骼肌组织胰岛素信号通路磷脂酰肌醇-3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin,PI3K/PKB/mTOR)蛋白表达水平的影响。方法选取2 1日龄SPF级SD雌性大鼠6 0只,其中1 2只为空白对照组,剩余48只利用经典Poresky造模方法联合高脂饲料诱导肥胖型PCOS-IR大鼠。造模成功后,随机分为模型对照组、中药低剂量组、中药中剂量组、中药高剂量组各12只,分别实施不同的干预措施。经治疗28天后,用葡萄糖氧化酶法检测空腹血糖(FPG),放射免疫法检测空腹胰岛素(FINS),计算胰岛素抵抗指数(HOMA-IR),酶联免疫法检测大鼠血清中促黄体生成素(LH)、促卵泡生长素(FSH)、睾酮(T);Western Blot法检测大鼠卵巢组织中PI3K、PKB、mTOR蛋白水平的表达。结果 (1)与空白对照比较,模型对照组体重、FPG、FINS、IR、LH、FSH、T、mTOR蛋白表达水平显著升高(P<0.05),PI3K/PKB蛋白表达水平显著降低(P<0.05);(2)与模型对照组比较,中药中剂量组体重显著降低(P<0.05),中药低剂量组、中药中剂量组、中药高剂量组FPG、FINS、IR、T值显著下降(P<0.05);(3)在卵巢组织中,中药高剂量组PI3K/PKB蛋白表达比中药中剂量组、中药低剂量组显著升高(P<0.05);(4)在骨骼肌组织中,中药高剂量组、中药中剂量组PI3K/PKB蛋白表达比中药低剂量组显著升高(P<0.05),中药中剂量组、中药高剂量组mTOR蛋白表达水平比低剂量组显著降低(P<0.05)。结论 (1)中药加减苍附导痰汤能够明显改善PCOS-IR大鼠体重、胰岛素抵抗情况及高雄状态。(2)中药上调PCOS大鼠卵巢组织及骨骼肌组织胰岛素信号通路PI3K/PKB/mTOR的活性,使蛋白PI3K/PKB相对表达量升高,mTOR蛋白相对表达量下降,具有浓度依赖性,推测中药改善肥胖型PCOS-IR大鼠胰岛素抵抗情况可能通过影响胰岛素信号通路PI3K/PKB/mTOR来起作用的。

Objective To establish the animal model of polycystic ovary syndrome with insulin re- sistance （PCOS-IR） to observe the effects of high-dose, medium-dose and low-dose modified Cangfu Daotan decoction （CFDT decoction） on the insulin resistance and sex hormone levels in obese PCOS-IRrats, and their effects on the protein expression levels of insulin signaling pathwws phosphatidylinositol 3- hydroxy kinase/protein kinase B/mammalian target of rapamycin （PI3K/PKB/nITOtl） in ovarian tissues and skeletal muscle tissues. Methods Sixty rats aged 21 days were selected, 12 of which were enrolled into the blank control group, and the remaining 48 rats were treated with classical Poresky modeling method combined with high-tat diet to induce the obese PCOS-IR rat model. After successtul modeling, the rats were randomly divided into control group （ n = 12 ）, low-dose traditional Chinese medicine group （ n = 12 ）, medium-dose traditional Chinese medicine group （n = 12 ） and high-dose traditional Chinese medicine group （ n = 12 ）. Different intermention measures were taken, respectively. After treatment for 28 days, the lasting plasma glucose （FPG） was measured via glucose oxidase method and thsting insulin （FINS） was measured via radioimmunoassay. Moreover, the homeostasis model assessment tor homa insulin-resistance （HOMA- IR） was calculated. Besides, the luteinizing honnone（ LH）, follicle-stimulating honnone （FSH） and tes- tosterone （T） in sermn were measured via enzyme-linked immunosorbent assay. Finally, the protein expression levels of PI3K/PKB/mTOIR in ovarian tissues were detected via Western blot. Results （ 1 ） Compared with those in blank group, the weight and the FPG, FINS, IR, LH, FSH, T and roTOR protein expression level in model control group were significantly increased （P〈0.05） , but the PI3K/PKB protein expression levels were significantly decreased （P 〈0.05 ）. （2） Compared with those in model control group, the weight in medium-dose traditional Chinese medicine group was obviously decreased （ P〈0.05 ） , and the FPG, FINS, IIR and T in low-, medium- and high-dose traditional Chinese medicine groups were obviously increased （P〈0.05）. （3） The PBK/PKB protein expression levels in ovarian tissues in high- dose traditional Chinese medicine group were significantly increased compared with those in medium and low-dose traditional Chinese medicine groups （P〈0.05）. （4） The PI3 K/PKB protein expression levels in skeletal muscle tissues in high-dose and medium-dose traditional Chinese medicine groups were significantly increased compared with those in low-dose traditional Chinese medicine group （ P〈0.05 ） , but the roTOR protein expression level in medium-dose and high-dose traditional Chinese medicine groups were remarkably decreased compared with those in low-dose traditional Chinese medicine group （P〈0.05）. Conclusion （1） Modified CFDT decoction can remarkably improve the weight, insulin resistance and high androgen status of PCOS-IR rats. （2） Traditional Chinese medicine can up-regulate the activity of insulin signaling pathways PI3K/PKB/mTOIR in ovarian tissues and skeletal muscle tissues of PCOS rats, increase the relative protein expression level of PI3K/PKB, decrease the relative protein expression level of roTOR in a concentration-dependent manner. Theretore, it is speculated that the improvement of insulin resistance in obese PCOS-IR rats with traditional Chinese medicine may be realized through attecting the insulin signaling pathways PI3 K/PKB/mTOR.