IL-1β在细胞坏死过程中对血管平滑肌细胞I型胶原形成的影响及机制研究

Effects of IL-1β Released by Necrotic Cells on Type I Collagen Synthesis in Vascular Smooth Muscle Cells

目的体外实验探讨细胞坏死过程中释放的IL-1β对周围正常血管平滑肌细胞中I型胶原表达的影响变化及内在机制。方法无血清低糖低氧培养条件下建立血管平滑肌细胞坏死模型。收集坏死细胞上清液，干预正常细胞。实验分为对照组（Contr01），坏死上清组（NCS）。IL-1β组，坏死上清＋IL-1拮抗剂组（NCS＋IL-1RA），IL-6组，坏死上清＋L-6拮抗剂组（NCS＋IL-6RA）。MTT检测各组细胞增殖情况：细胞划痕检测各组细胞迁移能力；ELISA检测各组细胞上清中TIMPl含量；RT-PCR检测各组细胞中Timpl、IL-6mRNA表达量；Western印迹检测各组MMP1、I型胶原（CollagenI）、p38、p-p38表达。结果与对照组比较，NCS组、IL-1β组、IL-6组细胞增殖及迁移能力均显著升高，TimplmRNA表达及分泌明显升高，CollagenI表达升高，MMPl表达降低，p38磷酸化程度增强（P〈0．05）；与NCS组比较，NCS＋IL-1RA组及IL-6组中刀-6mRNA表达较低，NCS＋IL-1RA组及NCS＋IL-6RA组增殖及迁移受抑，MMP1表达升高，CollagenI表达降低，p38磷酸化程度受到抑制（P〈0．05）。结论血管平滑肌细胞坏死过程中可能释放IL-1β并刺激正常细胞中IL-6表达，进而调节TIMP1及MMP1的生成以及CollagenI的表达，影响平滑肌细胞细胞增殖及迁移，该过程可能受到p38MAPK信号通路调控。

Objective To investigate the effect of IL-1 β released during cell necrosis on the expressions of type I collagen in normal vascular smooth muscle cells （VSMCs） and the underlying mechanisms. Methods The necrosis model of VSMCs was established under serum-free, hypoxic and glucose-deprived conditions. The supernatants collected were used to trea[ normal cells. Six groups were set up as follows： control group, necrosis cell supernatant group （NCS group） , IL-1β group, NCS ＋ IL-1 antagonist group （ NCS ＋ IL-1RA group）, IL-6 group, NCS ＋ IL-6 antagonist （NCS ＋ IL-6RA group） . The cell proliferation was detected by MTT and the cell migration by wound healing assay. The TIMP1 level in supematants was determined by ELISA. The mRNA expressions of IL-6 and TIMP1 were detected by RT-PCR and the protein expressions of MMP1, collagen I, p38 and p-p38 by Western blotting. Results Compared with the control group, the cell proliferation and migration were significantly increased, the mRNA expression and the secretion of TIMP1 were conspicuously enhanced, the expression of collagen I was markedly elevated, the expression of MMP1 was decreased and the phosphorylation of p38 was strengthened in the NCS group, IL-1β group and IL-6 group （P〈 0.05 ） . NCS ＋ IL-1RA group and IL-6 group than in the NCS group （P 〈 0. 05 ） . The cell proliferation and migration were significatly inhibited,the expresion of MMP1 L L increased, the expression of collagen I decreased and the phosphorylation of p38 suppressed in the NCS＋IL-1 RA group and NCS ＋ IL-6RA group relative to the NCS group （P 〈 0. 05 ） . Conclusion Neerotic VSMCs may release IL-1 β and stimulate the IL-6 expression in nounal cells, thereby regumnng me gen^rut~, MMP1 and the expression of collagen I, affecting the proliferation and migration of VSMG~, This process may be regulated via the P38 MAPK signaling pathwa3t.