Fei Xinfeng, Cai Yuying, Lin Feng, Huang Yongyi, Liu Te, Liu Yan
Department of Ophthalmology, Shanghai Fourth People's Hospital Affiliated to Tongji University School of Medicine, 100 Haining Road, Shanghai, 200080, China.
Department of Ophthalmology, Shanghai General Hospital, School of Medicine, Shanghai JiaoTong University, Shanghai, 200080, China.
Hum Cell. 2021 Jan;34(1):86-98. doi: 10.1007/s13577-020-00442-7. Epub 2020 Oct 3.
Severe corneal injury is one of the main causes of loss of visual function. Mesenchymal stem cells (MSCs) have the ability to repair damaged cells in vivo. The present study aimed to explore whether MSCs could function as a cell therapy tool to replace traditional methods to treat corneal injury. CD44 + /CD105 + mesenchymal stem cells isolated from mouse amniotic fluid (mAF-MSCs) were injected into mice after cryoinjury to induce corneal endothelial cell injury. Histopathological assays indicated that mAF-MSCs could promote the growth of corneal epithelial cells, reduce keratitis, and repair the corneal damage caused by low temperature. cDNA microarray analysis revealed that the mAF-MSCs affected the expression patterns of mRNAs related to cell proliferation and differentiation pathways in the mice after transplantation. The results of quantitative real-time PCR and western blotting revealed that NAT12, NAT10, and the ETV4/JUN/CCND2 signaling axis were elevated significantly in the mAF-MSC-transplantation group, compared with those in the phosphate-buffered saline-treated groups. High performance liquid chromatography-mass spectroscopy results revealed that mAF-MSCs could promote mRNA N4-acetylcytidine (ac4C) modification and high expression of N-acetyltransferase in the eyeballs. RNA immunoprecipitation-PCR results showed that a specific product comprising Vegfa, Klf4, Ccnd2, Jun, and Etv4 mRNA specific coding region sites could be amplified using PCR from complexes formed in mAF-MSC-transplanted samples cross-linked with anti-ac4C antibodies. Thus, mouse amniotic fluid MSCs could repair the mouse corneal cold injury by promoting the ETV4/JUN/CCND2 signal axis activation and improving its stability by stimulating N4-acetylcytidine modification of their mRNAs.
严重角膜损伤是视力功能丧失的主要原因之一。间充质干细胞（MSCs）具有在体内修复受损细胞的能力。本研究旨在探讨MSCs是否可作为一种细胞治疗工具来替代传统方法治疗角膜损伤。将从小鼠羊水分离的CD44 + /CD105 +间充质干细胞（mAF-MSCs）在冷冻损伤后注入小鼠体内以诱导角膜内皮细胞损伤。组织病理学分析表明，mAF-MSCs可促进角膜上皮细胞生长，减轻角膜炎，并修复低温引起的角膜损伤。cDNA微阵列分析显示，mAF-MSCs影响移植后小鼠中与细胞增殖和分化途径相关的mRNA表达模式。定量实时PCR和蛋白质印迹结果显示，与磷酸盐缓冲盐水处理组相比，mAF-MSC移植组中NAT12、NAT10和ETV4/JUN/CCND2信号轴显著升高。高效液相色谱-质谱结果显示，mAF-MSCs可促进眼球中mRNA N4-乙酰胞苷（ac4C）修饰和N-乙酰转移酶的高表达。RNA免疫沉淀-PCR结果表明，使用PCR可从与抗ac4C抗体交联的mAF-MSC移植样品中形成的复合物中扩增出包含Vegfa、Klf4、Ccnd2、Jun和Etv4 mRNA特异性编码区位点的特异性产物。因此，小鼠羊水MSCs可通过促进ETV4/JUN/CCND2信号轴激活并通过刺激其mRNA的N4-乙酰胞苷修饰来提高其稳定性，从而修复小鼠角膜冷损伤。
