HSPB1基因对脂多糖诱导的鸡巨噬细胞炎症反应的调控机制

Regulatory mechanism of HSPB1 gene on lipopolysaccharide-induced inflammatory response in chicken macrophages

为了探究热休克因子结合蛋白1(heat shock factor binding protein 1,HSPB1)基因对脂多糖(LPS)诱导的鸡巨噬细胞(HD11细胞)炎症反应的调控机制,试验以不同浓度的LPS刺激HD11细胞不同时间,构建HD11细胞炎症模型,利用HSPB1基因过表达质粒pcDNA3.1-HSPB1瞬时转染HD11细胞后再用最佳浓度的LPS进行刺激,并以仅转染pcDNA3.1和pcDNA3.1-HSPB1的HD11细胞为对照,采用实时荧光定量PCR方法检测转染HSPB1基因过表达质粒pcDNA3.1-HSPB1后对白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)和α干扰素(IFN-α)基因表达的影响。结果表明:在LPS浓度为1μg/mL、刺激时间为9 h时炎症因子IL-1β基因相对表达量极显著升高(P<0.01),成功构建出HD11细胞炎症模型;与转染pcDNA3.1质粒相比,转染HSPB1基因过表达质粒pcDNA3.1-HSPB1后能极显著下调IL-6、IL-1β和IFN-α基因的相对表达量(P<0.01);受LPS刺激且仅转染pcDNA3.1质粒后,能极显著上调IL-6、IL-1β和IFN-α基因的相对表达量(P<0.01),诱发细胞的炎症反应;而与受LPS刺激且仅转染pcDNA3.1质粒相比,转染HSPB1基因过表达质粒pcDNA3.1-HSPB1的HD11细胞受LPS刺激后能极显著下调IL-6、IL-1β和IFN-α基因的相对表达量(P<0.01),降低LPS刺激的炎症反应。说明HSPB1基因具有抗炎作用,与LPS联用有颉颃作用,能抑制由LPS刺激的HD11细胞炎性因子的表达水平,从而缓解细胞炎症反应。

In order to explore the regulatory mechanism of heat shock factor binding protein 1(HSPB1) on lipopolysaccharide(LPS)-induced inflammatory response in chicken macrophages(HD11 cells),in this experiment,different concentrations of LPS were used to stimulate HD11 cells at different times to construct an HD11 cells inflammation model.HSPB1 gene overexpression plasmid pcDNA3.1-HSPB1 was instantaneously transfected in HD11 cells which was then stimulated by the optimal concentration of LPS.HD11 cells transfected only with pcDNA3.1 and pcDNA3.1-HSPB1 were used as the control;real-time PCR method was used to detect the effect of the transfection of HSPB1 gene overexpression plasmid pcDNA3.1-HSPB1 on the expression of interleukin-6(IL-6),interleukin-1 beta(IL-1β) and α interferon(IFN-α) genes.The results showed that the relative expression of inflammatory factor IL-1β was significantly increased(P<0.01) at 1 μg/mL of LPS concentration and the stimulation time was 9 h,and the inflammatory model of HD11 cells was successfully constructed.Compared with the transfected pcDNA3.1 plasmid,the relative expression of IL-6,IL-1β and IFN-α genes could be significantly downregulated after overexpression of the plasmid pcDNA3.1-HSPB1 in HSPB1 genes(P<0.01).Compared with the transfection of pcDNA3.1 plasmid,the relative expression of IL-6,IL-1β and IFN-α genes(P<0.01) could be significantly downregulated after the transfection of HSPB1 gene-overexpression plasmid pcDNA3.1-HSPB1.After being stimulated by LPS and transfecting only pcDNA3.1 plasmid,the relative expression of IL-6,IL-1β and IFN-α genes could be significantly upregulated(P<0.01),inducing the inflammatory response of cells.Compared with LPS-stimulated and only transfected pcDNA3.1 plasmid,HD11 cells that were transfected with HSPB1 gene-overexpressed plasmid pcDNA3.1-HSPB1 were stimulated by LPS,which could significantly downregulate the relative expression of IL-6,IL-1β and IFN-α genes(P<0.01),and reduce the inflammatory response stimulated by LPS.The results indicated that HSPB1 gene had anti-inflammatory effect,and when combined with LPS,it had a fly-down effect,which could inhibit the expression levels of HD11 cell inflammatory factors stimulated by LPS,thereby alleviate the cellular inflammatory response.