目的：探讨TNF-α对兔胆管成纤维细胞P311/TGF-β1/α-SMA通路的影响及川芎嗪的干预作用。方法：分离、培养正常家兔胆管成纤维细胞并鉴定，将胆管成纤维细胞分别给予TNF-α、TNF-α联合不同浓度的TMP（0.08、0.4、2.0 mg/mL）干预48 h，以无处理的胆管成纤维细胞为空白对照，用CCK-8法检测细胞增殖水平；real-time PCR检测细胞P311、TGF-β1、α-SMA mRNA表达；Western blot检测细胞TGF-β1、α-SMA蛋白表达。结果：与空白对照比较，胆管成纤维细胞经TNF-α处理后，增殖明显增强，P311、TGF-β1、α-SMA mRNA以及TGF-β1、α-SMA蛋白表达明显上调（均P<0.05）；TMP对TNF-α的上述效应有抑制作用，并且呈现浓度依耐趋势，其中0.4、2.0 mg/mL的TMP有明显作用（均P<0.05）。结论：TNF-α可能通过调控P311/TGF-β1/α-SMA信号通路促进胆管成纤维细胞增殖，TMP能抑TNF-α对该通路的活化，故可能对良性胆道狭窄有防治作用。
Influence of TNF-α on activity of P311/TGF-β1/α-SMA signaling pathway in rabbit bile duct fibroblasts and the interventional effect of tetramethylpyrazine
Objective: To investigate the influence of TNF-α on activity of P311/TGF-β1/α-SMA signaling pathway in rabbit bile duct fibroblasts and the interventional effect of tetramethylpyrazine (TMP). Methods: Bile duct fibroblasts of rabbits were isolated and cultured, and then identified. The bile duct fibroblasts were exposed to TNF-α or TNF-α plus different concentrations (0.08, 0.4 and 2.0 mg/mL) of TMP respectively for 48 h, using untreated bile duct fibroblasts as blank control. Afterwards, the cell proliferation was determined by CCK-8 assay, mRNA expressions of P311, TGF-β1 and α-SMA were measured by real-time PCR, and protein expressions of TGF-β1 and α-SMA were examined by Western blot analysis. Results: In bile duct fibroblasts after TNF-α treatment, the cell proliferation significantly accelerated, and the mRNA expressions of P311, TGF-β1 and α-SMA as well as the protein expressions of TGF-β1 and α-SMA were significantly up-regulated compared with blank control cells (all P<0.05); TMP inhibited the above effects of TNF-α with a concentration-dependent tendency, and the inhibitions exerted by TMP at 0.4 and 2.0 mg/mL were significant (all P<0.05). Conclusion: TNF-α can promote proliferation of bile duct fibroblasts possibly via regulating P311/TGF-β1/α-SMA signaling pathway in fibroblasts, and TMP can inhibit the activation of this pathway induced by TNF-α, so it may have preventive and therapeutic effect on benign biliary stricture.