参麦开肺散对NIH/3T3成纤维细胞增殖及胶原合成的影响

Effect of Shenmaikaifei Powder on the Proliferation and Collagen Synthesis of NIH /3T3 Fibroblasts

目的探讨参麦开肺散对NIH/3T3成纤维细胞(FB)增殖及体外纤维化细胞模型胶原合成的影响。方法NIH/3T3 FB常规培养后分为正常对照组、模型组和处理组,正常对照组加入含1%胎牛血清(FBS)的细胞培养液500μl,模型组加入使用含1%FBS细胞培养液配制的5 ng/mlβ型转化生长因子(TGF-β)溶液500μl,处理组加入含5 ng/ml的TGF-β+1 mg/ml的参麦开肺散混合溶液500μl,培养24 h。分别采用实时定量聚合酶链反应(PCR)、蛋白免疫印迹(Western blot)和Sircol assay检测细胞内胶原及细胞外基质(ECM)相关基因表达水平、Ⅰ型胶原表达情况以及分泌到细胞上清中胶原蛋白的含量。结果参麦开肺散显著降低胶原及其相关基因、Ⅰ型胶原蛋白的表达以及分泌到细胞上清中胶原蛋白的含量(P<0.05,P<0.01)。结论参麦开肺散可明显抑制TGF-β诱导的NIH/3T3 FB胶原的合成,其发挥抗纤维化作用可能与其抑制FB增殖及胶原合成相关。

Objective To investigate the effect of Shenmaikaifei powder on the proliferation and collagen synthesis of NIH /3T3 fibroblasts models in vitro. Methods NIH /3T3 fibroblasts underwent conventional culture,and then were divided into control group,model group and treatment group. The control group was added with 1% FBS cell culture fluid（ 500 μl）; model group was added with 1% FBS cell culture fluid and 5 ng / ml transforming growth factor-β（ TGF-β） solution（ 500 μl）; treatment group was added with 5 ng /ml TGF-βand 1 mg /ml Shenmaikaifei powder solution（ 500 μl）; the culture time was 24 h. The real-time quantitative PCR（ polymerase chain reaction） was used to examine the transcript expression levels of intra-cellular collagen and extracellular matrix（ ECM） genes,and western blot was conducted to determine the protein expressions of type Ⅰ collagen,and Sircol assay was used to measure the collagen protein contents secreted to the cell supernatant. Results Shenmaikaifei powder significantly decreased the expressions of collagen and related genes,type I collagen and the content secreted to the cell supernatant（ P〈0. 05,P〈0. 01）.Conclusions Shenmaikaifei powder can inhibited the synthesis of collagen in TGF-β-induced NIH /3T3 fibroblasts,and its anti-fibrotic effect may relate to inhibition of fibroblast proliferation and collagen biosynthesis.