南方医科大学学报

南方医科大学学报 ›› 2020, Vol. 40 ›› Issue (07): 1001-1007.doi: 10.12122/j.issn.1673-4254.2020.07.13

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胆汁酸通过p-STAT3-SOCS3途径调节下丘脑神经元食欲及抑制信号肽表达

陈春秀，周 永，黄荣凤，王妙然，李 月，李继斌   

  • 出版日期:2020-07-20 发布日期:2020-07-20
  • 基金资助:

Bile acids regulate anorexigenic neuropeptide through p-STAT3-SOCS3 signaling in mouse hypothalamic cells

  

  • Online:2020-07-20 Published:2020-07-20

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摘要： 目的 探索胆汁酸牛磺石胆酸或鹅去氧胆酸对小鼠下丘脑GT1-7细胞食欲抑制信号肽表达的影响及可能的作用途径。方法 将实验分为3组，每组3个平行样本。空白对照组：仅给予含10%胎牛血清高糖培养基培养GT1-7细胞。胆汁酸牛磺石胆酸处理组：分别采用10 nmol/L，100 nmol/L，1 μmol/L和10 μmol/L浓度的胆汁酸牛磺石胆酸处理GT1-7细胞12、24和48 h。鹅去氧胆酸处理组：分别采用10 nmol/L，100 nmol/L，1 μmol/L和10 μmol/L浓度的鹅去氧胆酸处理GT1-7细胞12、24和48 h。采用Real-time PCR 检测GT1-7细胞食欲抑制信号肽阿黑皮素原（POMC）mRNA表达水平；ELISA法检测POMC剪切产物抑制食欲肽α-促黑素细胞激素水平。采用Western blot检测G蛋白偶联胆汁酸受体（TGR5）和法呢醇X受体表达以及磷酸化信号转导转录激活因子3（p-STAT3），磷酸化蛋白激酶（p-AKT），细胞因子信号传导抑制因子3（SOCS3）蛋白表达量。结果 Western blot结果显示，下丘脑神经元GT1-7细胞能表达TGR5和FXR两种胆汁酸受体，且表达量受胆汁酸调节。Real-time PCR检测结果显示，10 μmol/L tLCA或鹅去氧胆酸处理GT1-7细胞24 h后POMC mRNA表达水平上升，ElISA结果也发现10 μmol/L 胆汁酸牛磺石胆酸或鹅去氧胆酸处理GT1-7细胞24 h后具有抑制食欲作用的POMC衍生肽[α]-MSH释放增多。Western blot结果显示，胆汁酸处理可上调GT1-7细胞p-STAT3和p-AKT蛋白表达，且调控STAT3信号通路的SOCS3蛋白表达也有增加。结论 胆汁酸牛磺石胆酸或鹅去氧胆酸通过GT1-7细胞胆汁酸受体TGR5和FXR促进POMC mRNA表达，增加[α]-促黑素细胞激素水平，提示胆汁酸具有通过抑制食欲信号肽表达调节食欲的潜在作用，该作用是通过p-AKT和p-STAT3 -SOCS3 信号通路实现。

关键词: 下丘脑神经元, GT1-7细胞, 胆汁酸, 牛磺石胆酸, 鹅去氧胆酸, 阿黑皮素原

Abstract: Objective To explore the effects of taurolithocholic acid (tLCA) and chenodeoxycholic acid (CDCA) on the expression of aorexigenic neuropeptide in mouse hypothalamus GT1-7 cells. Methods Mouse hypothalamic GT1-7 cells were treated with culture medium containing 10% FBS (control group, n=3) or with 10 nmol/L, 100 nmol/L, 1 μmol/L and 10 μmol/L tLCA (tLCA group, n=3) or CDCA (CDCA group, n=3) for 12, 24 or 48 h. Real-time PCR was performed to determine the expression levels of proopiomelanocortin (POMC) mRNA in the cells, and the production levels of α-melanocyte-stimulating hormone (α-MSH) were assessed using an ELISA kit. Signal transduction and activator of transcription 3 phosphorylation (p-STAT3), threonine kinase phosphorylation (p-AKT), suppressor of cytokine signaling 3 (SOCS3), G protein-coupled bile acid receptor-1 (TGR5) and farnesoid X receptor (FXR) protein were detected by Western blotting. Results Western blotting results showed that mouse hypothalamic GT1-7 cells expressed two bile acid receptors, TGR5 and FXR, whose expressions were regulated by bile acids. Real-time PCR showed that the expression of POMC mRNA was significantly increased in the cells after treatment with 10 μmol/L tLCA or CDCA for 24 h. POMC-derived anorexigenic peptide α-MSH increased significantly in GT1-7 cells after treatment with 10 μmol/L tLCA or CDCA for 24 h. Treatment of the cells with tLCA or CDCA significantly increased the expressions of intracellular signaling proteins including p-STAT3, p-AKT and SOCS3. Conclusion Mouse hypothalamic GT1-7 cells express bile acid receptors TGR5 and FXR. Bile acids tLCA or CDCA can promote the expression of POMC mRNA and increase the production of the anorexigenic peptide α-MSH. The intracellular signaling proteins p-AKT, p-STAT3 and SOCS3 are likely involved in bile acid-induced anorexigenic peptide production.

Key words: hypothalamic cells, GT1-7 cells, bile acid, taurolithocholic acid, chenodeoxycholic acid, proopiomelanocortin