睾酮的快速非基因组效应对3T3-L1脂肪细胞胰岛素敏感性的影响

Testosterone impairs insulin sensitivity in 3T3-L1 adipocytes and the related nongenomic mechanisms

目的探讨睾酮的快速非基因组效应对3T3-L1成熟脂肪细胞胰岛素敏感性的影响及分子机制。方法将3T3-L1前脂肪细胞诱导成熟，用10^-9-10^-5moL／L睾酮分别预处理0～30min及24h，[^3H]-2-脱氧葡萄糖掺入法检测葡萄糖摄取率；通过免疫印迹检测胰岛素受体（InsR）／蛋白激酶（Akt）／糖原合成激酶（GSK3β）的活性及表达。结果有胰岛素刺激时，睾酮预处理30min的胰岛素促葡萄糖摄取率随睾酮浓度增高而下降，10^-5mol／L睾酮组的摄取率的升高倍数（4．2±0．4）显著低于无睾酮组（5．5±0．4），P〈0．05；10^-6mol／L睾酮预处理3～30min的InsR／Akt／GSK3β的磷酸化水平较无睾酮组均明显下降（均P〈0．05）。有胰岛素刺激时，睾酮预处理24h的胰岛素促葡萄糖摄取率也随睾酮浓度增高而下降，10^-5mol／L睾酮组的摄取率的升高倍数（4．0±1．0）显著低于无睾酮组（4．5±1．0），P〈0．05；10^-7-10^-6mol／L睾酮预处理24h的InsR／Akt／GSK3β的活性均明显下降（均P〈0．05），但InsR／Akt／GSK3β的表达水平均不受睾酮影响（均P〉0．05）。结论高浓度睾酮的快速非基因组效应可能通过抑制胰岛素信号分子InsR／Akt／GSK3β的活性从而在诱导成熟脂肪细胞产生胰岛素抵抗中起重要作用。

Objective To investigate the influence of rapid nongenomic effect of androgen on the insulin sensitivity of mature adipocytes and the molecular mechanism thereof. Methods Fetal rat preadipocytes of the line 3T3-L1 were cultured to develop into mature adipocytes. 3T3-L1 adipocytes were pretreated with testosterone of the concentration of 10 ^-9 - 10 ^-5 mol/L for a short-time （0 - 30 minutes） or a long-time （24 hours）. Insulin （Ins） 200 μl at the concentration 100 μmol/L and 2-deoxy [^3H] glucose were added to examine the glucose uptake. Phosphorylation and protein expression of Ins receptor （InsR） and its downstream signaling molecules （ Akt and GSK3β） were analyzed by Western blotting. Results The Ins-stimulated glucose uptake after the pretreatment of testosterone for 30 min and 24 h decreased gradually in response to the increasing of the concentration of testosterone with the nadir both at the testosterone concentration of 10^-5 mol/L （both P 〈 0.05）. The phosphorylation levels of InsR, Akt, and GSK3β were significantly down-regulated by adding of testosterone at the concentration of 10 ^-6 mol/L for 3 - 30 minutes （ all P 〈 0.05 ） or by adding of testosterone at the concentrations of 10 ^-7 - 10 ^-6 mol/L for 24 hours （ all P 〈 0.05）. The protein expression levels of InsR, Akt, and GSK3β however, were not significantly affected by testosterone treatment. Conclusion Rapid nongenomic effect of androgen may contribute to the insulin resistance in adipocytes.