TNF-α和VEGF协同作用小鼠胚胎干细胞衍生的血管内皮祖细胞促伤口愈合  

Synergistic Effects of TNF-α and VEGF on mESC-Derived EndothelialProgenitor Cells in Promoting Wound Healing

作　　者：杨影 陈东升 赵艾艾 何才蓉 陈凤娇 何运雪 郑梅 陆莹 丁洁 YANG Ying;CHEN Dong-Sheng;ZHAO Ai-Ai;HE Cai-Rong;CHEN Feng-Jiao;HE Yun-Xue;ZHENG Mei;LU Ying;DING Jie(Key Laboratory of Plant Resource Conservation and Germplasm Innovation in Mountainous Region(Ministry of Education),College of Life Sciences/Institute of Agro-bioengineering,Guizhou University,Guiyang 550025,China;Scientific Research Division of Guizhou Provincial People’s Hospital,Guiyang 5500025,China;Department of Deplaning,Basic Medical College,Guizhou University of Traditional Chinese Medicine,Guiyang 550025,China)

机构地区：[1]贵州大学生命科学学院/农业生物工程研究院,山地植物资源保护与保护种质创新教育部重点实验室,贵阳550025 [2]贵州省人民医院科研处,贵阳550025 [3]贵州中医药大学基础医学院解刨学教研室,贵阳550025

出　　处：《中国生物化学与分子生物学报》2024年第2期240-250,共11页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.32260163);贵州省科技计划项目(黔科合基础-ZK[2021]一般108);国家自然科学基金项目(No.81960838)资助。

摘　　要：皮肤伤口愈合是世界范围内的重大临床难题之一,血管生成和炎症反应是影响伤口愈合和组织再生的关键环节。内皮祖细胞(endothelial progenitor cell,EPC)在血管内皮修复和招募炎症细胞促伤口愈合过程中发挥重要作用。然而,体内直接输送EPC低效且会损害细胞存活力和功能进而影响治愈效率。因此,改善EPC的生物学功能以促进伤口愈合很有必要。本研究将小鼠胚胎干细胞(mouse embryonic stem cells,mESC)在10 ng/mL VEGF和5 ng/mL bFGF的作用下诱导获得CD133+CD34+EPC。以mESC衍生的EPC为研究对象,将外源性肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和血管内皮生长因子(vascular endothelial growth factor,VEGF)作用于mESC-EPC,结果表明,10 ng/mL TNF-α和10 ng/mL VEGF协同处理组相比于单因子处理显著促进EPC的黏附、细胞迁移和管腔形成能力(P<0.01)。进一步通过建立小鼠局部皮肤全层创伤模型,在创周处注射10 ng/mL TNF-α和10 ng/mL VEGF协同EPC治疗明显加速伤口愈合(P<0.05),再生皮肤真皮层增厚(P<0.001),促进血管生成素ANG1和ANG2介导CD31+内皮细胞构成的毛细血管网络成熟。随着伤口愈合程度的加深,促炎因子TNF-α在蛋白质水平的表达下调(术后13 d,PBS组、EPC组、VE组、TE组和VTE组的TNF-α相对表达量分别为0.73±0.01、0.60±0.02、0.42±0.02、0.36±0.01和0.34±0.03),创造有利于伤口愈合的炎症微环境。综上所述,10 ng/mL TNF-α和10 ng/mL VEGF的协同作用强化EPC的生物学功能,并通过促进新血管生成和早期炎症反应加速小鼠皮肤伤口闭合和组织重塑,为细胞干预伤口愈合治疗提出新的思路。Skin wound healing is one of the major clinical problems worldwide.Angiogenesis and the inflammation response are critical processes that influence wound healing and tissue regeneration.Endothelial progenitor cells(EPCs)play a pivotal role in vascular endothelial repair and the recruitment of inflammatory cells to promote wound healing.However,direct in vivo delivery of EPC is inefficient and can impair cell viability and function,thereby affecting therapeutic efficacy.Therefore,it is necessary to improve the biological functions of EPC to promote wound healing.This study induced the generation of CD133+CD34+EPC from mouse embryonic stem cells(mESCs)under the influence of 10 ng/mL Vascular Endothelial Growth Factor(VEGF)and 5 ng/mL basic Fibroblast Growth Factor(bFGF).We used mESC-derived EPC as the research object,and applied exogenous tumor necrosis factor-α(TNF-α)and Vascular Endothelial Growth Factor(VEGF)to mESC-EPC.The results showed that the co-treatment with 10 ng/mL TNF-αand 10 ng/mL VEGF significantly promoted the adhesion,cell migration,and luminal formation abilities of EPC compared to the single-factor treatment(P<0.01).Further,using a murine full-thickness skin wound model,local administration of 10 ng/mL TNF-αand 10 ng/mL VEGF in conjunction with EPC treatment markedly accelerated wound closure(P<0.05),thickened the dermal layer of the regenerated skin(P<0.001),and promoted the maturation of the capillary network formed by CD31+endothelial cells mediated by angiopoietin ANG1 and ANG2.As wound healing progressed,the expression of the pro-inflammatory factor TNF-αwas down-regulated(13 days after surgery,the relative expression levels of TNF-αin PBS group,EPC group,VE group,TE group and VTE group were 0.73±0.01,0.60±0.02,0.42±0.02,0.36±0.01,0.34±0.03,respectively),creating a conducive inflammatory microenvironment for wound healing.In summary,the synergistic effects of 10 ng/mL TNF-αand 10 ng/mL VEGF enhance the biological functions of EPC,as well as accelerate wound closure and tissue rem

关 键 词：小鼠胚胎干细胞 血管生成因子 内皮祖细胞 伤口愈合 肿瘤坏死因子-Α 

分 类 号：Q46[生物学—生理学]