1-磷酸鞘氨醇对肺成纤维细胞纤维化发生的影响和机制

The effect of S1P on HLF cell fibrosis and its mechanism

目的:研究S1P对HLF细胞纤维化的影响及机制。方法:(1)分别用S1P(1μmol/L)、FTY720-P(5μmol/L)、ponesimod(5μmol/L)、SEW2871(5μmol/L)处理HLF细胞24h,使用Western Blot法测ECM相关蛋白表达并与只加培养基空白组对比;(2)分别用S1P1R拮抗剂W146(1μmol/L)、S1P2R拮抗剂JTE-013(0.2μmol/L)和S1P3R拮抗剂TY-52156(1.25μmol/L)预处理HLF细胞1 h,再加入S1P及S1PR激动剂分组孵育24 h后测ECM相关蛋白表达并与无拮抗剂对照组对比;(3)使用S1P2R拮抗剂JTE-013和S1P3R拮抗剂TY-52156共同预处理HLF细胞1h,加入S1P及S1PR激动剂分组孵育24h后测ECM相关蛋白表达并与无拮抗剂对照组及单一拮抗剂组对比。结果:(1)S1P及选择性S1P受体激动剂组均上调ECM表达,但程度不同;(2)S1P1R拮抗剂W146不影响S1P及S1PR激动剂诱导ECM合成,S1P2R拮抗剂JTE-013及S1P3R拮抗剂TY-52156均下调S1P及S1PR激动剂诱导ECM合成;(3)两种拮抗剂同时使用能使S1P及非选择性S1PR激动剂诱导ECM合成进一步降低,但不能使ponesimod组进一步降低。结论:HLF细胞中,S1P通过与S1P2R和S1P3R结合增加ECM相关促纤维化蛋白表达。

Objective To study the effect of S1P on HLF cell fibrosis and its mechanism. Methods （ 1 ） The expression of ECM in HLF ceils was analyzed by using Western Blot after treatment by S1P （1 μmol/L） , FTY720-P（5 μmol/L）, ponesimod （5 p.mol/L） and SEW2871 （5 μmol/L） 24 h; （2）The HLF cells were pre-treated using selective S1PR antagonist W146 （ 1 μ mol/L）, JTE-013 （ 0.2 μmol/L）, and TY-52156 （ 1.25 μmol/L） 1 h before incubation by SIP and S1PR agonists 24 h and then the expression of ECM was analyzed; （3）The HLF cells were pre-incubated using JTE-013 （0.2 μmol/L） and TY-52156（ 1.25 μmol/L） for 1 h and then the expression of ECM was analyzedafter being treated by S1P and S1PR agonists 24 h. Results （ 1 ）SIP and selective SIP receptor agonist increased the expression of ECM to various extents; （2）The S1P1R antagonist W146 did not affectthe expression of ECM induced by S1P and S1PR agonists and S1P2R antagonist JTE-013 and S1P3R antagonist TY-52156 both decreased the expression of ECM induced by SiP and S1PR agonists ; （3）The expression of ECM induced by SIP and SIPR agonists further decreased using both JTE-013 and TY-52156 but not using ponesimod. Conclusion S1P2R and S1P3R are activated under the influence of SiP so as to increase the synthesis of ECM and promote fibrosis gene expression in HLF cells.