Zhang Mingzhao, Liu Hengchen, Shi Manyu, Zhang Tingting, Lu Wenjun, Yang Shulong, Cui Qingbo, Li Zhaozhu
Department of Pediatric Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, China.
Front Cell Dev Biol. 2021 Jun 18;9:659389. doi: 10.3389/fcell.2021.659389. eCollection 2021.
The therapeutic impact of stem cells is potentially largely attributable to secretion of exosomes and soluble factors. The present study evaluates the impact of hepatocyte growth factor (HGF)-expressing tendon stem cells (TSCs) on tendon healing in a rat model. Patellar tendon TSCs were isolated and underwent transfection with lentiviral vectors containing HGF or green fluorescent protein (GFP) genes. , immunohistochemistry of tendons sampled 1 week postsurgery demonstrated that all stem cell-treated groups exhibited higher numbers of CD163 M2 monocytes and IL-10 cells (anti-inflammatory), and lower numbers of CCR7 M1 monocytes and IL-6 as well as COX-2 cells (pro-inflammatory). Effects were most pronounced in the HGF-expressing TSCs (TSCs + HGF) treated group. Histology ± immunohistochemistry of tendons sampled 4 and 8 weeks postsurgery demonstrated that all stem cell-treated groups exhibited more ordered collagen fiber arrangement and lower levels of COLIII, α-SMA, TGF-β1, and fibronectin (proteins relevant to fibroscarring). Effects were most pronounced in the TSCs + HGF-treated group. For the study, isolated tendon fibroblasts pretreated with TGF-β1 to mimic the microenvironment of tendon injury were indirectly cocultured with TSCs, TSCs + GFP, or TSCs + HGF using a transwell system. Western blotting demonstrated that all stem cell types decreased TGF-β1-induced increases in fibroblast levels of COX-2, COLIII, and α-SMA, concomitant with decreased activation of major TGF-β1 signaling pathways (p38 MAPK, ERK1/2, but not Smad2/3). This effect was most pronounced for TSCs + HGF, which also decreased the TGF-β1-induced increase in activation of the Smad2/3 signaling pathway. The presence of specific inhibitors of these pathways during fibroblast TGF-β1 stimulation also attenuated increases in levels of COX-2, COLIII, and α-SMA. In conclusion, TSCs + HGF, which exhibit HGF overexpression, may promoting tendon healing via decreasing inflammation and fibrosis, perhaps partly via inhibiting TGF-β1-induced signaling. These findings identify a novel potential therapeutic strategy for tendon injuries, warranting additional research.
干细胞的治疗作用可能很大程度上归因于外泌体和可溶性因子的分泌。本研究评估了表达肝细胞生长因子（HGF）的肌腱干细胞（TSCs）对大鼠模型中肌腱愈合的影响。分离髌腱TSCs，并用含有HGF或绿色荧光蛋白（GFP）基因的慢病毒载体进行转染。术后1周采集的肌腱免疫组织化学结果显示，所有干细胞治疗组的CD163 M2单核细胞和IL-10细胞（抗炎）数量较多，而CCR7 M1单核细胞、IL-6以及COX-2细胞（促炎）数量较少。在表达HGF的TSCs（TSCs + HGF）治疗组中效果最为明显。术后4周和8周采集的肌腱组织学及免疫组织化学结果显示，所有干细胞治疗组的胶原纤维排列更有序，COLIII、α-SMA、TGF-β1和纤连蛋白（与纤维瘢痕形成相关的蛋白质）水平较低。在TSCs + HGF治疗组中效果最为明显。在本研究中，用TGF-β1预处理以模拟肌腱损伤微环境的分离肌腱成纤维细胞，使用Transwell系统与TSCs、TSCs + GFP或TSCs + HGF间接共培养。蛋白质印迹法显示，所有干细胞类型均降低了TGF-β1诱导的成纤维细胞COX-2、COLIII和α-SMA水平的升高，同时主要TGF-β1信号通路（p38 MAPK、ERK1/2，但不包括Smad2/3）的激活减少。TSCs + HGF的这种效果最为明显，其还降低了TGF-β1诱导的Smad2/3信号通路激活的增加。在成纤维细胞TGF-β1刺激期间，这些信号通路的特异性抑制剂的存在也减弱了COX-2、COLIII和α-SMA水平的升高。总之，表现出HGF过表达的TSCs + HGF可能通过减轻炎症和纤维化来促进肌腱愈合，可能部分是通过抑制TGF-β1诱导的信号传导。这些发现确定了一种针对肌腱损伤的新型潜在治疗策略，值得进一步研究。
