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肾脏病与透析肾移植杂志 ›› 2015, Vol. 24 ›› Issue (4): 319-324 .

• 论文 • 上一篇    下一篇

脂肪酸结合蛋白3对足细胞脂肪代谢的影响

  

  • 出版日期:2015-08-28 发布日期:2015-09-01

Effects of fatty acid binding protein 3 on the fat metabolism of podocyte

  • Online:2015-08-28 Published:2015-09-01

摘要:

摘要 目的:观察脂肪酸结合蛋白3(FABP3)过度增加对足细胞脂肪代谢的影响。方法:慢病毒载体转染小鼠永生化足细胞系(HSMPs)以构建稳定过表达FABP3的细胞系。将FABP3-siRNA转染至HSMPs中沉默FABP3,筛选干扰效率最大的FABP3-siRNA片段进行后续干扰实验。实验分组:(1)正常足细胞组(2)NC-siRNA足细胞组(3)FABP3-siRNA足细胞组(4)NC足细胞组(5)FABP3过表达足细胞组(6)正常足细胞+棕榈酸/脂肪酸(PA)组(7)NC-siRNA足细胞+PA组(8)FABP3-siRNA足细胞+PA组(9)NC足细胞+PA组(10)FABP3过表达足细胞+PA组。比色法测定细胞内 TG和游离脂肪酸的含量。定量PCR法检测各组细胞中过氧化物酶增殖激活的受体α(PPARα)、酰基辅酶A氧化酶3(ACOX3)mRNA的表达。Western blot法检测PPARα、ACOX3蛋白表达水平。结果:与正常足细胞组相比,FABP3过表达足细胞组细胞内TG和游离脂肪酸水平增加(P<0.05),而FABP3-siRNA足细胞组TG和游离脂肪酸水平下降(P<0.05)。不论在正常环境还是PA环境下,与正常足细胞组相比,FABP3过表达足细胞组PPARα mRNA和蛋白表达均下调(P<0.05),而ACOX3 mRNA和蛋白表达均上调(P<0.05); FABP3-siRNA足细胞组PPARα mRNA和蛋白表达均上调(P<0.05),而ACOX3 mRNA和蛋白表达均下调(P<0.05)。与FABP3过表达足细胞组相比,FABP3-siRNA足细胞组PPARα mRNA和蛋白表达上调(P<0.05),ACOX3 mRNA和蛋白表达下调(P<0.05);加入PA后,FABP3过表达足细胞+PA组和FABP3-siRNA 足细胞+PA组之间PPARα mRNA和蛋白表达、ACOX3 mRNA和蛋白表达的差异具有显著统计学意义(P<0.01)。结论:FABP3过度增加可导致足细胞脂肪代谢紊乱,抑制FABP3过度增加可纠正足细胞脂肪代谢紊乱状态。该研究结果提示抑制FABP3表达可能对代谢因素如糖尿病、肥胖、高脂血症所致肾脏足细胞损伤有一定防御和治疗作用。

关键词: 脂肪酸结合蛋白3, 过氧化物酶增殖激活的受体α, 酰基辅酶A氧化酶3, 足细胞

Abstract:

ABSTRACT Objective: To investigate the effect and mechanisms of fatty acid binding protein 3 on the fat metabolism of podocyte. Methodology: We transfect the lentivirus vector to heat-sensitive mouse podocytes(HSMPs) to construct the cell line which express FABP3 stabile and excessively. Two siRNAs were desianed and synthesized. One of them that was capable of silencing FABP3 more effectively in HSMPs were chosen to do the next interference study. They were divided into ten cell groups: (1) normal control group; (2) NC-siRNA group; (3) FABP3-siRNA group; (4) NC group; (5) excessive express FABP3 group;(6)normal control+palmic acid(PA) group(7)NC-siRNA+PA group;(8)FABP3-siRNA+PA group;(9)NC+PA group;(10)excessive express FABP3+PA group. Triglycerides(TG) and free fatty acid contents in podocytes were determined by colorimetric method. The mRNA expression of peroxisome proliferator activated receptors α(PPARα) and ACOX3 of each group was determined by using Real-Time PCR. The proteins of PPARα and ACOX3 were examined by using Western blot. Results: Compared with normal control group, the levels of TG and free fatty acid were up-regulated in group5(FABP3 group), while down-regulated in group3(FABP3-siRNA group) (P<0.05).Compared with normal control group, mRNA and protein expressions of PPARα were down-regulated in FABP3 group(P<0.05), while mRNA and protein expressions of ACOX3 were up-regulated in FABP3 group(P<0.05) with or without PA. Compared with normal control group, mRNA and protein expressions of PPARα were up-regulated in FABP3-siRNA group(P<0.05), while mRNA and protein expressions of ACOX3 were down-regulated in FABP3-siRNA group with or without PA(P<0.05). Compared with FABP3 group, mRNA and protein expressions of PPARα were up-regulated in FABP3-siRNA group(P<0.05), while mRNA and protein expressions of ACOX3 were down-regulated in FABP3-siRNA group(P<0.05). There was more conspicuous statistical difference between FABP3+PA group and FABP3-siRNA+PA group in the expression of mRNA and proteins of PPARα and ACOX3(P<0.01). Conclusion: The excessive expression of FABP3 may induce fatty metabolic disturbance in podocytes. Inhibiting the expression of FABP3 may improve the fatty metabolic disturbance in podocytes. These results prompted that inhibiting the expression of FABP3 may ameliorate the metabolic disorder induced podocyte injury, such as diabetes mellitus, obesity and dyslipidemia.

Key words:  Fatty acid binding protein 3, peroxisome proliferator activated receptors α, Acyl-Coenzyme A oxidase 3, podocyte