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目的研究富含半胱氨酸的酸性分泌糖蛋白(SPARC)对人常染色体显性多囊肾病(ADPKD)囊肿衬里上皮细胞(CLECs)细胞周期及其调控基因表达的影响,初步探讨SPARC对CLECs的作用。方法体外条件下用不同浓度SPARC处理CLECs,5鄄鄄2脱溴氧尿苷酶联免疫吸附测定法(BrdUELISA)测定CLECs增殖;流式细胞术检测细胞周期;实时荧光定量RT鄄PCR方法检测CLECs细胞周期调控基因ClnD1、P21Waf1表达水平的变化。结果μg水平的SPARC能有效抑制ADPKDCLECs增殖(P<0.01),使细胞周期停滞于G0/G1期。10μg/mlSPARC刺激后,ClnD1mRNA水平(3.56±0.54)×104拷贝/百万GAPDH较对照组(7.50±0.99)×104显著减弱,P21WmRNAaf1水平(7.72±0.85)×103较对照组(4.25±1.38)×103显著增强。结论SPARC能够有效抑制CLECs细胞周期的进展。其机制可能通过抑制ClnD1、促进P21Waf1的表达,抑制细胞通过G1鄄S期限制点,从而对其增殖产生显著的抑制作用。
Objective To investigate the effect of cysteine-rich acidic secretory glycoprotein (SPARC) on the cell cycle and the expression of regulatory genes in cyst-lining epithelial cells (CLECs) of human autosomal dominant polycystic kidney disease (ADPKD). To investigate the effects of SPARC on CLECs Role. Methods CLECs were treated with different concentrations of SPARC in vitro, the proliferation of CLECs was detected by 5-deoxy-2-deoxyuridine (BrdUELISA), the cell cycle was detected by flow cytometry, the expression of CLECs was detected by real-time fluorescence quantitative RT-PCR Changes of cell cycle regulatory genes ClnD1, P21Waf1 expression. Results μg of SPARC could effectively inhibit the proliferation of ADPKDCLECs (P <0.01), and arrested the cell cycle in G0 / G1 phase. Compared with the control group (7.50 ± 0.99) × 104, the ClnD1 mRNA level (3.56 ± 0.54) × 104 copies / million GAPDH was significantly decreased after 10μg / ml SPARC stimulation. The level of P21W mRNAaf1 (7.72 ± 0.85) × 103 was 4.25 ± 1.38 × 103 is significantly enhanced. Conclusion SPARC can effectively inhibit the cell cycle progression of CLECs. The mechanism may inhibit ClnD1, promote the expression of P21Waf1, and inhibit the cells through the G1-S phase restriction point, thereby significantly inhibiting its proliferation.