论文部分内容阅读
目的探讨载脂蛋白D(apolipoprotein D,ApoD)在常染色体显性多囊肾病(autosomal dominant polycystic kidneydisease,ADPKD)发病中的作用。方法采用免疫组织化学及蛋白质印迹方法分析ApoD在ADPKD患者肾脏囊肿组织中的表达;蛋白质印迹法和实时荧光定量PCR法分别检测Han:SPRD大鼠肾组织ApoD蛋白和基因表达的变化。应用MTT法检测人重组ApoD蛋白对ADPKD囊肿衬里上皮细胞(WT9-12)增殖的影响;流式细胞术检测经人重组ApoD蛋白作用后WT9-12细胞的凋亡情况及细胞周期改变。结果 ADPKD患者及Han:SPRD杂合型大鼠(cy/+)肾组织中ApoD表达量分别低于正常人及Han:SPRD正常大鼠(+/+)肾组织(P<0.05,P<0.01)。8、12、16、24周龄Han:SPRD杂合型大鼠(cy/+)肾脏中Ap-oD基因表达量均低于同周龄Han:SPRD正常大鼠(+/+)(P<0.05)。人重组ApoD蛋白以100、200、400 ng/ml作用于WT9-12细胞,作用48 h细胞增殖抑制率分别为8.21%、7.59%、8.07%(P<0.05),作用72 h细胞增殖抑制率分别为8.62%、6.43%、9.42%(P<0.05)。人重组表达ApoD蛋白以400 ng/ml浓度刺激细胞72 h,对细胞凋亡无明显影响,但可使G1期细胞增加8.26%,S期细胞减少8.09%(P<0.05)。结论 ApoD蛋白可能通过调节细胞周期减弱WT9-12细胞增殖,其表达减少可能在ADPKD的发病过程中具有一定作用。
Objective To investigate the role of apolipoprotein D (ApoD) in the pathogenesis of autosomal dominant polycystic kidney disease (ADPKD). Methods Immunohistochemistry and Western blotting were used to analyze the expression of ApoD in kidney cysts of ADPKD patients. Western blotting and real-time fluorescence quantitative PCR were used to detect the changes of ApoD protein and gene expression in renal tissues of Han: SPRD rats. The effect of human recombinant ApoD protein on the proliferation of ADPKD cyst-lining epithelial cells (WT9-12) was detected by MTT assay. The apoptosis and cell cycle of WT9-12 cells treated with recombinant ApoD protein were detected by flow cytometry. Results The expression of ApoD in ADPKD patients and Han: SPRD heterozygous rats (cy / +) kidney was lower than that in normal subjects and Han: SPRD normal rats (+ / +) kidney (P <0.05, P <0.01 ). The expression of Ap-oD gene in Han: SPRD heterozygous (cy / +) kidneys at 8, 12, 16 and 24 weeks of age were lower than those in the same age Han: SPRD normal rats (P < 0.05). The inhibitory rates of human recombinant ApoD protein in WT9-12 cells treated with 100, 200 and 400 ng / ml for 48 h were 8.21%, 7.59% and 8.07%, respectively (P <0.05) Respectively, 8.62%, 6.43%, 9.42% (P <0.05). The recombinant ApoD protein was stimulated by 400ng / ml for 72h, but had no effect on apoptosis. However, the ApoD protein increased 8.26% in G1 phase and 8.09% in S phase (P <0.05). Conclusion ApoD protein may reduce the proliferation of WT9-12 cells by regulating the cell cycle. The decreased expression of ApoD may play a role in the pathogenesis of ADPKD.