IRF8基因降表达、PINK1基因过表达对肺泡巨噬细胞NR8383增殖与活性的影响

Effects of down-regulated expression of IRF8 gene and over-expression of PINK1 gene on proliferation and activity of alveolar macrophages NR8383

目的观察干扰素调节因子8(IRF8)基因降表达、PTEN诱导激酶1(PINK1)基因过表达对肺泡巨噬细胞NR8383增殖与活性的影响,探讨IRF8与PINK1基因在该细胞增殖与活性中的作用。方法取对数生长期的小鼠肺泡巨噬细胞NR8383,随机分为7组。IRF8对照组、PINK1对照组分别转染IRF8干扰空质粒、PINK1过表达空质粒,联合对照组共转染用量减半的上述两种质粒;IRF8干扰组、PINK1过表达组分别转染IRF8干扰质粒pG2.2-IRF8-Rat-444、PINK1过表达质粒pCMV3-Rat-PINK1,联合干扰组共转染用量减半的上述两种质粒;正常对照组不做任何处理。转染48 h后收集细胞,采用CCK8法检测细胞增殖的OD值,LDH法检测细胞活性,采用Western blotting法检测细胞中的IRF8、PINK1蛋白。结果与联合对照组、PINK1对照组、IRF8对照组、正常对照组比较,IRF干扰组与PINK1过表达组均IRF8蛋白表达降低而PINK1蛋白表达升高,细胞增殖及细胞活性均降低(P均<0.05);与IRF干扰组、PINK1过表达组比较,联合干扰组IRF8蛋白表达降低而PINK1蛋白表达升高(P均<0.05),细胞增殖及细胞活性均降低,但仅细胞活性差异有统计学意义(P均<0.05)。结论IRF8基因降表达、PINK1基因过表达均可明显抑制肺泡巨噬细胞NR8383增殖及细胞活性,且具有协同作用。

Objective To observe the effects of down-regulated expression of interferon regulatory factor 8(IRF8)gene and over-expression of PTEN induced putative kinase 1(PINK1)gene on the proliferation and activity of alveolar macrophages NR8383,and to explore the role of IRF8 and PINK1 genes in the cell proliferation and activity.Methods Mouse alveolar macrophages NR8383 in the logarithmic growth phase were randomly divided into 7 groups.Alveolar macrophages NR8383 in the IRF8 control group and PINK1 control group were transfected with IRF8 interfering empty plasmid and PINK1 overexpressing empty plasmid,respectively,and the combined control group with half the dosage of the two plasmids.The IRF-8 interference group and the PINK1 overexpression group were respectively transfected with the IRF8 interference plasmid pG2.2-IRF8-Rat-444 and the PINK1 overexpression plasmid pCMVV3-Rat-PINK1,and the combined interference group was co-transfected with half the dosage.The normal control group did not receive any treatment.Cells were collected at 48 h after transfection,OD values of cell proliferation were detected by CCK8,cell activity was detected by LDH,and IRF8 and PINK1 proteins were detected by Western blotting.Results Compared with the combined control group,the PINK1 control group,the IRF8 control group and the normal control group,the IRF8 interference group and the PINK1 overexpression group all showed decreased IRF8 protein expression and increased PINK1 protein expression,and decreased cell proliferation and cell activity(all P<0.05).Compared with the PINK1 overexpression group and the IRF8 interference group,the expression of IRF8 protein decreased and the expression of PINK1 protein increased in the combined interference group(all P<0.05),and both the cell proliferation and cell activity decreased,but only the difference in the cell activity was statistically significant(both P<0.05).Conclusion The down-regulated expression of IRF8 gene and the over-expression of PINK1 gene can significantly inhibit the proliferation and cell activity of alveolar macrophages NR8383 with synergistic effects.