热休克转录因子5在小鼠睾丸不同细胞中的表达及其功能探索

Expression and functional exploration of heat shock transcription factor 5 in mouse testis cells

目的探讨热休克转录因子5(heat shock transcription factor 5,HSF5)在小鼠睾丸不同细胞中的表达情况及其功能。方法培养3种(GC-2、TM3、TM4细胞)小鼠睾丸组织细胞,并通过RT-qPCR绝对定量测定HSF5的mRNA表达水平。利用siRNA干扰技术,针对高表达HSF5细胞进行hsf5基因敲低,通过RT-qPCR和Western blot检测其他几种热休克转录因子以及热休克蛋白表达情况。建立脂多糖感染高表达HSF5细胞模型,分为无脂多糖对照组、脂多糖对照组、脂多糖hsf5敲低组,通过ELISA法测定细胞上清中炎症细胞因子IL-1β、IL-6水平。结果同TM3、TM4细胞相比,GC-2细胞中HSF5的mRNA表达量最高。敲低GC-2中的HSF5表达后,HSF1以及HSP60的mRNA表达下调(P<0.05);HSF1的蛋白表达量也出现下调(P<0.05)。脂多糖感染GC-2细胞模型建模成功,同脂多糖对照组相比,脂多糖hsf5敲低组中的IL-6、IL-1β水平降低(P<0.05)。结论3种小鼠生殖细胞中,GC-2中HSF5的mRNA表达量最高;HSF5在小鼠生殖细胞中极可能充当了免疫保护的角色进而参与精子的发生从而影响男性不育。

Male infertility is a global population health problem,the decline of sperm quality is a main cause of male infertility.Environmental stress on sperm damage has become a research hotspot in recent years.To investigate the expression and function of heat shock transcription factor 5 in different cells of mouse testis.Three types of mouse testis cells were cultured and the mRNA expression level of HSF5 was detected by RT-qPCR.Using siRNA interference technology to knockdown hsf5 gene in GC-2 cell,the effects of other heat shock transcription factors and heat shock proteins related to spermatogenesis were verified by RT-qPCR and western blot.Set the model of lipopolysaccharide infection,GC-2 cells were divided into three groups:control group,lipopolysaccharide control group and lipopolysaccharide hsf5-knockdown group.The levels of IL-1βand IL-6 in cell supernatant was detected by ELISA.Compared with TM3 and TM4 cells,the mRNA expression level of HSF5 in the GC-2 cells is highest.The model of GC-2 cells infected by lipopolysaccharide was successfully established.Compared with lipopolysaccharide control group,the levels of IL-1βand IL-6 in cell supernatant were decreased in the lipopolysaccharide hsf5-knockdown group(P<0.05).ThemRNA expression of HSF5 in GC-2 was the highestamong the three types of mouse testis cells.HSF5 mayact as an immune protector in mouse germ cells toparticipate in spermatogenesis.[Key words]Male infertility;Spermatogenesis.