过表达IL-6对山羊毛囊干细胞增殖和迁移的影响

Effects of Overexpression of IL-6 on Proliferation and Migration of Goat Hair Follicle Stem Cells in vitro

旨在通过在山羊毛囊干细胞(HFSCs)中过表达炎症因子白细胞介素6(IL-6),探讨其对山羊HFSCs增殖和迁移的影响,阐明MAPK/ERK信号通路在山羊HFSCs增殖和迁移过程中发挥的作用。本试验通过构建pXJ40-myc-IL-6过表达载体,瞬时转染到山羊HFSCs中,以空载体细胞为空白对照组。利用Western blot定量检测IL-6蛋白的表达,MTT法检测山羊HFSCs的增殖活力,划痕试验判定山羊HFSCs的迁移能力,最后采用Westernblot检测山羊HFSCs中MAPK/ERK和P38信号通路中关键激酶的表达水平。结果表明,将获得的IL-6过表达载体转染至山羊HFSCs3d后,与对照组比较,山羊HFSCs的增殖活力出现抑制,且抑制效果持续至第7天(P<0.05)。IL-6过表达载体转染至山羊HFSCs24h后,处理组划痕部位的愈合率达到84.2%,显著低于对照组的98.5%(P<0.01)。与对照组相比,MAPK/ERK信号通路中的关键激酶ERK1/2的磷酸化水平(p-ERK1/2)显著下调(P<0.01),P38信号通路中的关键激酶p38的磷酸化水平(p-p38)表达量上调,但差异无统计学意义(P>0.05)。综上所述,过表达IL-6基因对山羊HFSCs的增殖和迁移能力具有抑制作用,且过表达IL-6可能是通过MAPK/REK信号通路对山羊HFSCs增殖和迁移发挥负性调控作用,以上结果为揭示山羊HFSCs的迁移及组织修复机制的研究提供依据。

The aim of this study was to investigate the effects of overexpressing inflammatory factor interleukin-6 (IL-6) on the proliferation and migration of goat hair follicle stem cells (HFSCs) and elucidate the role of MAPK/ERK signaling pathway in the proliferation and migration of goat HFSCs. The pXJ40-myc-IL-6 overexpression vector was constructed and transiently transfected into the goat HFSCs,and the empty vector cells were used as the blank control group. Western blot was used to quantitatively detect the expression of IL-6 protein,and MTT assay was used to detect the proliferation of goat HFSCs. The migration ability of HFSCs was detected by wound healing assay. Finally,Western blot was used to detect the expression of key kinases in MAPK/ERK and P38 signaling pathway in goat HFSCs. The results showed that after transfecting the IL -6 overexpression vector into goat HFSCs for 3 days,compared with the control group,the proliferation activity of the HFSCs was inhibited and the sustained inhibition effect was observed until the 7th day( P <0.05). After 24 h of transfection of IL -6 overexpression vector into goat HFSCs,the healing rate of wounding site in the treatment group reached 84.2%,which was significantly lower than that in the control group (98.5%, P <0.01). Compared with the control group,the phosphorylation level of the key kinase ERK1/2 (p-ERK1/2) in the MAPK/ERK signaling pathway was significantly down-regulated ( P <0.01),the phosphorylation level of the key kinase p38 (p-p38) in the P38 signaling pathway was up-regulated,however,the change was not statistically significant ( P >0.05). In summary, the overexpression of IL -6 gene has an inhibitory effect on the proliferation and migration ability of goat HFSCs,and its negative regulatory role on the proliferation and migration of goat HFSCs maybe through the MAPK/REK signaling pathway. The results provide basis for revealing the mechanism of migration and tissue repair of goat HFSCs.