摘要
目的晚期糖基化终产物受体(the receptor of advanced glycation end products,RAGE)特异性小干扰RNA(small in-terfering RNA,siRNA)能在肝星状细胞(hepatic stellate cell,HSCs)T6内抑制RAGE、α-平滑肌肌动蛋白(alpha-smooth muscle ac-tin,α-SMA)和Ⅰ型胶原mRNA和蛋白的表达,其在HSCs的激活和胶原的形成中发挥着重要的作用。探讨RAGE特异性siRNA在原代大鼠HSCs中对致纤维化细胞因子生成的影响。方法构建RAGE特异性siRNA表达载体,分离培养原代大鼠HSCs,将重组载体转染入原代大鼠HSCs,以空白组和转染非特异性siRNA表达载体pAKD-NC组为对照,分别用PCR法和Western blot检测各组原代HSCs RAGE、β1转化生长因子(transforming growth factor-β1,TGF-β1)和结缔组织生长因子(connective tissuegrowth factor,CTGF)mRNA及蛋白的表达。结果转染特异性siRNA表达载体pAKD-GR126的原代HSCs RAGE mRNA和相对分子质量为42000的RAGE蛋白的表达分别为空白组和pAKD-NC组的(42.32±6.16)%、(43.24±7.50)%和(51.06±13.79)%、(47.94±5.36)%,差异有统计学意义(P<0.05)。同时TGF-β1 mRNA和蛋白的表达率分别占空白组和pAKD-NC组的(43.72±0.76)%、(44.75±3.78)%和(45.35±2.03)%、(47.71±3.32)%(P<0.05),CTGF mRNA和蛋白的表达分别是空白组和pAKD-NC组的(39.32±7.22)%、(39.50±4.58)%和(42.58±5.95)%、(43.74±2.16)%,差异有统计学意义(P<0.05)。结论 RAGE特异性siRNA可抑制原代大鼠HSCs RAGE基因的表达,并能有效抑制致纤维化细胞因子的生成。
Objective Specific small interfering RNA （siRNA） targeting receptor for advanced glycation end products （RAGE） can inhibit the expressions of RAGE, alpha-smooth muscle actin （ct-SMA） and collagen type I in hepatic stellate cell （HSC） T6, which indicates the key role of RAGE in the activation of HSCs and expression of collagen. This study aims to investigate the effect of RAGE specific siRNA on the expressions of profibrogenic cytokines in primary rat HSCs. Methods Primary rat HSCs were isolated and cultured in vitro. The expression vectors of specific siRNA targeting RAGE were constructed and transfected to the primary rat HSCs, and untreated and unspecific siRNA-transfected primary rat HSCs served as controls. The expressions of RAGE, transforming growth factor-β1 （TGF-β1） and connective tissue growth factor （CTGF） in the primary HSCs were detected by real-time PCR and Western blotting respectively. Results The mRNA and 42KD protein expressions of RAGE in the pAKD-GR126-transfected primary HSCs were （42.32 ±6. 16） %, （43.24 ± 7.50） %, （51.06 ± 13.79） % and （47.94 ± 5.36） % of those in the untreated and pAKD-NC-transfected control groups （ all P 〈 0.05 ）. The mRNA and protein expressions of TGF-β1 and CTGF in the pAKD- GR126-transfected primary HSCs were （43.72 ± 0.76） %, （44.75 ± 3.78） %, （45.35 ± 2.03 ） %, （47.71 ± 3.32 ） %, （ 39.32 ± 7.22 ）%, （39.50 ± 4.58 ）%, （42.58 ± 5.95 ）% and （43.74 ± 2.16）% of those in the untreated and pAKD-NC-transfected control groups （ all P 〈 0.05 ）. Conclusion RAGE specific siRNA can inhibit the RAGE gene expression in primary rat HSCs, and effectively suppress the production of profibrogenic cytokines.
出处
《医学研究生学报》
CAS
北大核心
2013年第5期465-468,共4页
Journal of Medical Postgraduates
基金
江苏省自然科学基金(BK2009284)
关键词
小干扰RNA
糖基化终产物受体
原代肝星状细胞
致纤维化细胞因子
Small interfering RNA
Receptor for glyeation end product
Primary hepatic stellate cell
Profibrogenic cytokine
