透明质酸对游离脂肪酸引起的人主动脉血管平滑肌细胞表达IL-6、IL-8和TNF-α的影响

Effect of hyaluronic acid on the expressions of IL-6,IL-8 and TNF-α in human aortic vascular smooth muscle cells induced by free fatty acid

目的探讨透明质酸(HA)对游离脂肪酸(FFA)引起的人主动脉血管平滑肌细胞(HA-VSMC)表达白细胞介素6(IL-6)、IL-8和肿瘤坏死因子α(TNF-α)的影响。方法培养HA-VSMC,用不同浓度(0、100、200、400μmol/L)FFA和不同浓度FFA+高分子量HA(HMW-HA)或低分子量HA(LMW-HA)处理细胞24、48 h。噻唑蓝法检测FFA和FFA+HA对HA-VSMC存活率的影响;Ed U染色检测细胞增殖活性;酶联免疫吸附法检测培养上清IL-6、IL-8和TNF-α含量;实时荧光定量PCR和Western blot检测IL-6、IL-8和TNF-αmRNA及蛋白表达水平。结果(1)FFA在400μmol/L内呈浓度依赖性诱导HA-VSMC增殖;(2)HMW-HA、LMW-HA均对FFA诱导的HA-VSMC增殖有抑制作用,且HMW-HA抑制作用强于LMW-HA;(3)FFA显著促进HA-VSMC IL-6、IL-8和TNF-α的分泌,且有明显的时间和浓度依耐性;HA可以抑制上述作用;(4)实时荧光定量PCR和Western blot结果显示,FFA+HMWHA组、FFA+LMW-HA组IL-6、IL-8、TNF-αmRNA及蛋白表达水平均低于FFA组(均P<0.01),其中FFA+HMW-HA组又低于FFA+LMW-HA组(P<0.01)。结论 (1)FFA诱导HA-VSMC IL-6、IL-8、TNF-αmRNA和蛋白表达升高;(2)HA对FFA引起的HA-VSMC IL-6、IL-8和TNF-α的表达有抑制作用,且HMW-HA抑制效果强于LMW-HA。

Aim To investigate the effect of hyaluronic acid （HA） on the expression of interleukin-6 （IL-6） , IL-8 and tumor necrosis factor-α （TNF-α） in human aortic vascular smooth muscle cell （HA-VSMC） induced by free fatty acid （FFA）. Methods HA-VSMCs were cultured and dealt with different concentrations （0, 100,200, 400 μmol/L） of FFA and FFA plus high molecular weight HA （HMW-HA） or low molecular weight HA （LMW-HA） for 24, 48 hours. The effects of FFA and FFA plus HA on the survival rate of HA-VSMCs were detected by methyl thiazolyl tetrazolium （MIT） method. Cell proliferation activity was detected by EdU staining. The contents of IL-6, IL-8 and TNF-α in culture supernatant were detected by enzyme-linked immunosorbent assay （ELISA）. Real-time fluorescence quantitative PCR and Western blot were used to detect the expression levels of IL-6, IL-8 and TNF-α mRNA and protein. Results （ 1 ） FFA showed a concentration-dependent induction to HA-VSMC proliferation in the 400 μmol/L. （2） HMW-HA and LMW-HA both inhibited the proliferation of HA-VSMC induced by FFA, and the inhibitory effect of HMW-HA was stronger than that of LMW-HA. （3） FFA significantly promoted the secretions of HA-VSMC IL-6, IL-8 and TNF-α in a apparent time- and concentration-dependent manner; HA could inhibit the above effect. （4）Real-time fluorescence quantitative PCR and Western blot results showed that the expression levels of IL-6, IL-8, TNF-α mRNA and protein in FFA＋HMW- HA group and FFA＋LMW-HA group were lower than those in FFA group （all P〈0.01 ）, where the FFA＋HMW-HA group was lower than the FFA＋LMW-HA group （P〈0.01）. Conclusions （ 1 ） FFA can induce the increases of IL-6, IL-8 and TNF-α mRNA and protein expressions in HA-VSMCs. （2） HA can inhibit the expressions of IL-6, IL-8 and TNF-α induced by FFA, and the inhibitory effect of HMW-HA is stronger than that of LMW-HA.