活化的CREBH蛋白对脂肪酸诱导的脂肪变性肝细胞中C反应蛋白表达的调控研究  

Effect of activated CREBH protein on the expression of C-reactive protein in fatty-acid-induced steatotic hepatic cells

作　　者：赵雄[1] 艾罗燕[1] 苏大芝[1] 王晓晗[1] 江小柯[1] 许青青[1] 吴昌维[1] 陈志威[1] 范竹萍[1] 

机构地区：[1]上海交通大学医学院附属仁济医院健康保健中心,上海市消化疾病研究所,200127

出　　处：《中华内分泌代谢杂志》2014年第2期141-145,共5页Chinese Journal of Endocrinology and Metabolism

摘　　要：目的建立脂肪酸诱导的脂肪变肝细胞模型，探讨活化的cAMP应答元件结合蛋白H（CREBH）蛋白对该模型中C反应蛋白表达的影响。方法给予L02细胞不同比例和浓度的油酸软脂酸混合液（2：1、1：2和0：3），分别干预12和24h。CCK-8法测定细胞活力。流式测定细胞内脂肪含量。实时PCR方法检测细胞内C反应蛋白、CREBH、C／EBP同源蛋白C／EBP（CHOP）和免疫球蛋白结合蛋白（BIP）mRNA表达水平变化。Western印迹法检测细胞核内活化的CREBH蛋白含量。RNA干扰靶向沉默CREBH蛋白后再分别采用混合脂肪酸干预24h。实时PCR方法检测细胞内C反应蛋白、CREBH、CHOP和BIPmRNA表达水平变化。结果（1）混合脂肪酸诱导肝细胞脂肪变。随着软脂酸比例的升高．细胞内c反应蛋白表达水平升高，并且具有时间依赖性。（2）混合脂肪酸可以诱导CREBH表达升高（P〈0．05）．可增加核内CREBH蛋白水平。（3）混合脂肪酸中软脂酸比例越高，内质网应激标志增加越显著。（4）CREBHsiRNA可有效抑制CREBH表达。敲除CREBH基因后，C反应蛋白表达显著下降（P〈0．05）。结论软脂酸可上调肝细胞C反应蛋白表达水平，但是油酸无此作用。活化的CREBH可调节软脂酸诱导的C反应蛋白表达。Objective To establish a steatosis hepatocyte model in vitro and explore the effect of activated cAMP-responsive element-binding protein H （CREBH） protein on the expression of C-reactive protein （CRP） in this model. Methods L02 cells were cultured with free fatty-acid （FFA） mixtures at different ratios of oleic acid and palmitic acid（2 ： 1, 1 ： 2, and 0 ： 3 ） for 12 or 24 h to induce fat-overloading. Cell viability was investigated by CCK-8, the levels of CRP, CREBH, C/EBP homologous protein （CHOP） , and immunoglobulin-binding protein （BIP） were detected by real-time PCR. The level of CREBH in nuclei was detected by western blot. Then CREBH siRNA or negative control siRNA was transfected into L02 cells, and cultured with FFA mixtures for 24 hours. The levels of CRP, CREBH, CHOP, and BIP were detected by real-time PCR. Results （ 1 ） Cells cultured with FFA mixtures showed significantly increased fat droplets compared with control group. The level of CRP was increased by FFA mixtures, depending on the proportion of palmitic acid. （2）The expression levels of CREBH were increased by FFA mixture（ P〈0.05 ） and its expression in nuclear was also increased by FFA mixture. （ 3 ） Endoplasmic reticulum stress could be induced by FFA mixture, depending on the proportion of palmitic acid. （4） The CREBH siRNA could effectively reduce the expression level of CREBH mRNA. Compared with the negative control group, the expression levels of CRP mRNA were significantly decreased（P〈0.05 ）. Conclusion Palmitic acid, but not oleic acid, could up-regulate the expression of CRP. Palmitic acid-induced expression of CRP was regulated by activated CREBH.

关 键 词：脂肪变性肝细胞 cAMP应答元件结合蛋白H C反应蛋白 内质网应激 饱和脂肪酸 

分 类 号：R575.5[医药卫生—消化系统]