Fsp27基因抑制原代肝星状细胞活化与活性的实验研究

Fsp27 gene regulate the activity of primary hepatic stellate cells in vitro

目的探讨脂肪特异性蛋白27（fat—specific protein 27，Fsp27）基因对原代肝星状细胞（ hepatic stellate cells, HSCs）增殖活化的影响和对纤维化相关基因的调节作用。方法实时定量PCR检测原代与活化HSCs中Fsp27基因表达。构建携带Fsp27的慢病毒并转染原代HSCs。CCK-8比色法检测Fsp27基因对HSCs增殖的影响。Western blotting检测HSCsd肌动蛋白（α-SMA）的表达，实时定量PCR检测Fsp27基因对HSCs纤维化相关基因表达的影响。结果原代与活化HSCs的Fsp27表达差异有统计学意义（Fsp27mRNA：2700±500，5±2，t=124．27，P〈0．05）。带Fsp27目的基因的慢病毒转染原代HSCs72h后可明显抑制HSCs的活化（66．2±3．8。37．5±4．3，21．7±1．4，F=43．27，P〈0．05）与增殖（48h：31．5±1．3，6．1±0．1，t=17．35，P〈0．05；72h：36．2±2．2，6．2±0．1，t=21．94，P〈0．05）；Fsp27基因促进基质金属蛋白酶2表达（0．48±0．02，0．85±0．14，1．63±0．21，F=41．56，P〈0．05），降低转化生长因子B1表达（3．2±0．17，2．0±0．3，1．7±0．2，F=21．52，P〈0．05）。结论Fsp27基因抑制原代HSCs活化增殖，调节纤维化相关基因的表达，可能与维持HSCs静息状态细胞表型有关。

Objective To investigate the effect of fat-specific protein 27 （Fsp27） gene on primary hepatic stellate cells （HSCs） in vitro. Methods Rt-PCR was used to detect the expression of Fsp27 gene in primary and activated HSCs. The lentivirus carrying Fsp27 was constructed and transfected into primary HSCs. Effect of Fsp27 gene on proliferation of HSCs was tested by CCK-8 colorimetry. Detection of HSCs α- actin （ a -SMA） expression was tested by Western blotting. Effect of Fsp27 gene on expression of fibrosis-related genes in HSCs were tested by Real-time quantitative PCR. Results The difference of Fsp27 gene between primary HSCs and activiated HSCs was significant （Fsp27 mRNA：2 700 ±500,5±2,t = 124. 27, P 〈 0. 05 ）. After coeuhure for 72h, the proliferation and activation of HSCs was inhibited by Fsp27 （48 h ： 31.5±1.3,6.1±0.1,t=17.35,P〈0.05;72h：36.2±2.2,6.2±0.1,t=21.94,P〈0.05）. Fsp27 can enhance the expression of MMP-2 gene （0. 48±0. 02, 0. 85±0. 14,1.63±0. 21, F = 41.56, P 〈0. 05 ） and down-regulate the expression of TGF-β1 gene in activiated HSCs （ 3.2±. 17,2. 0±0. 3,1.7±0. 2, F = 21.52,P 〈 0. 05 ）. Conclusion Fsp27 gene can inhibit the proliferation and activation of primary HSCs. Fsp27 can regulate the expression of fibrosis related gene. Fsp27 may play an important role in maintenance the quiescent phenotype of HSCs.