目的 观察过氧化体增殖物激活型受体(peroxisome proliferator-activated receptors,PPARs)激活剂——非诺贝特和吡格列酮对新生大鼠心肌细胞肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)表达水平的影响,探讨可能涉及的机制。方法 体外原代培养新生Wistar大鼠心肌细胞,分别给予不同浓度的非诺贝特(PPARα激活剂)或吡格列酮(PPARγ激活剂)刺激,并辅加脂多糖诱导TNF-α表达。采用半定量RT-PCR法检测TNF-αmNNA的表达,ELISA法检测TNF-α的蛋白水平。心肌细胞瞬时转染TNF-α启动子控制的报告基因载体,测定CAT相对活性以反映TNF-α基因的转录活性。结果 与对照组相比,非诺贝特组和吡格列酮组的TNF-αmRNA及蛋白表达水平均明显降低,且呈剂量依赖性。心肌细胞瞬时转染全长TNF-α启动子控制的报告基因质粒(-721/+17),非诺贝特或吡格列酮可降低脂多糖诱导的CAT相对活性,而转染核因子κB(NF-κB)结合位点缺失的TNF-α启动子控制的报告基因质粒(-182/+17),非诺贝特、吡格列酮或脂多糖刺激均未能改变心肌细胞的CAT相对活性。结论 非诺贝特和吡格列酮可显著抑制新生大鼠心肌细胞中脂多糖诱导的TNF-α表达,具有抗炎作用,其机制可能部分通过抑制NF-κ信息通路发挥作用。
Abstract
OBJECTIVE To investigate the effects of peroxisome proliferator-activated receptors (PPARs)--fenofibrate and pioglitazoneon tumor necrosis factor-α (TNF-α) expression in cardiac myocytes of neonatal rat. The possible mechanism of action was explored. METHODS Primary cultures of cardiac myocytes were prepared from 1 - 3 day old Wistar rats, and then the myocytes were exposed to lipopolysac-charide (LPS) and fenofibrate and pioglitazone at different concentrations. RT-PCR and EL1SA were used to measure TNF-α expression in cultured cardiac myocytes. Transient transfection of TNF-α promoter with or without nuclear factor-kappaB (NF-icB) binding site to cardiac myocytes was performed. RESULTS Pretreatment of cardiac myocytes with fenofibrate or pioglitazone inhibited LPS-induced TNF-α mRNA and protein expression in dose-dependent manner. Proportional supression of TNF-α promoter activity was observed when the myocytes were transiently transfected with whole length of TNF-α promoter ( - 721/ +17) after being stimulated with LPS and fenofibrate or pioglitazone, whereas no change of the promoter activity was observed with the transfection of TNF-α reporter with deletion of NF-icB binding site ( - 182/ + 17) . CONCLUSION Fenofibrate and pioglitazone inhibit cardiac TNF-a expression and appear to play a role in anti-inflammation. The mechanism may be partly involved in suppression of NF-κB pathway.
关键词
非诺贝特 /
吡格列酮 /
肿瘤坏死因子-α /
心肌细胞 /
核因子κB
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Key words
fenofibrate /
pioglitazone /
tumor necrosis factor-α /
cardiac myocytes /
activators /
nuclear factor-kappaB
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参考文献
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脚注
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基金
国家自然科学基金项目(30270551);军队“十五”课题基金(02M012)
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