半乳糖凝集素3调控高糖下肾小球系膜细胞增殖的研究

Galectin-3 regulates proliferation of glomerular mesangial cells under high glucose condition

目的探究半乳糖凝集素3(galectin-3,Gal-3)对高糖培养的肾小球系膜细胞(mesangial cells,MCs)增殖的作用及机制。方法实时定量PCR(qRT-PCR)检测Gal-3在糖尿病肾病(diabetic nephropathy,DN)、正常小鼠肾脏组织和高低糖条件下MCs中的表达;构建、合成Gal-3过表达质粒和siRNA,qRT-PCR和Western blot检测转染效率;在高糖培养的MCs中转染Gal-3-siRNA;在低糖培养的MCs中转染Gal-3过表达质粒,通过5-乙基-2′-脱氧尿嘧啶核苷(5-Ethynyl-2′-deoxyuridine, EdU)检测MCs的增殖能力,通过Western blot检测高低表达Gal-3对Mek和磷酸化Mek表达的影响。结果 qRT-PCR结果显示:与正常小鼠相比,Gal-3在DN小鼠肾脏组织中表达显著升高(P<0.01),且与低糖下的MCs相比,高糖条件下的MCs中Gal-3表达显著升高(P<0.01)。过表达Gal-3后,MCs增殖能力显著提高(P<0.01)且磷酸化Mek表达上调(P<0.01),而在siRNA沉默Gal-3后,MCs增殖能力显著降低(P<0.01)且磷酸化Mek表达下调(P<0.001)。结论 Gal-3在DN中显著上调,并且可能通过调控Mek磷酸化影响MCs的增殖,进而在DN中发挥重要的调节作用。

Objective To investigate the effect and underlying mechanism of galectin-3(Gal-3) on the proliferation of mesangial cells(MCs) cultured under high glucose condition. Methods Real-time quantitative PCR(qRT-PCR) was used to detect the expression of Gal-3 in normal and diabetic nephropathy(DN) mouse kidney tissues and MCs under low and high glucose conditions. Gal-3 overexpression plasmid and siRNA were constructed and synthesized. qRT-PCR and Western blotting were used to detect the transfection efficiency. And Gal-3 overexpression plasmid or siRNA was transfected into MCs under low and high glucose condition, respectively. Further, 5-Ethynyl-2′-deoxyuridine(EdU) was used to detect the cell proliferation ability, and Western blot assay was used to measure the expression of Mek and phosphorylated Mek after overexpression or downexpression of Gal-3 in MCs. Results qRT-PCR results showed that Gal-3 expression was significantly higher in the kidney tissues of DN mice than those of the normal mice(P<0.01), and so was the expression level in MCs under high glucose condition when compared with the cells under low glucose condition(P<0.01). In addition, overexpression of Gal-3 resulted in enhanced proliferation ability of MCs(P<0.01) and the up-regulation of phosphorylated Mek(P<0.01). While, Gal-3 silencing induced the opposite results in the proliferation ability(P<0.01) and phosphorylated Mek expression(P<0.001). Conclusion Gal-3 is significantly up-regulated in DN, and may affect the proliferation of MCs by regulating Mek phosphorylation, and thus plays an important regulatory role in DN.