糖尿病足溃疡(DFU)是糖尿病的主要并发症,治疗需求未得到满足。本研究应用人脐带间充质干细胞-透明质酸(hucMSCs-HA)凝胶,以无创的方式治疗DFU,并探讨其旁分泌作用和机制。在这项研究中,在分析了HA凝胶的物理和生物学特性后,hucMSCs-HA凝胶应用于2种体内模型(I型和II型DFU),并研究了分子机制。为了评估hucMSCs的旁分泌作用,收集hucMSC条件培养基(MSC-CM)以处理1个体内模型(I型DFU)和2个体外模型(高糖(HG)损伤的人脐静脉内皮细胞(HUVEC)和人皮肤成纤维细胞(HSF))。结果表明,具有多孔微结构的HA凝胶在48h内降解超过90%并膨胀到最大值。hucMSCs-HA凝胶促进DFU大鼠创面愈合,胶原蛋白沉积,和血管生成,其中证实了hucMSCs的旁分泌作用和p38,ERK1/2,JNK的磷酸化,Akt增加了。体外,MSC-CM改善细胞活力,伤口愈合,迁移,管形成,细胞衰老,和异常表达(TNF-α,IL-1β,IL-6,ET-1,p16基因,和HUVECs的PCNA蛋白),也提高了细胞活力,伤口愈合,抗氧化应激,和异常表达(COL1,COL3,COL4,SOD1,SOD2基因,和HSF的PCNA蛋白)。总之,非侵入性外用hucMSCs-HA凝胶对DFU有很好的治疗前景,并通过MAPK和Akt通路介导的旁分泌机制发挥伤口愈合作用。
Diabetic foot ulcer (DFU) is a main diabetic complication with unmet treatment needs. This study applied human umbilical cord-derived mesenchymal stem cells-hyaluronic acid (hucMSCs-HA) gel to treat DFU in a noninvasive external way and investigated its paracrine action and mechanism. In this study, after analyzing the physical and biological properties of HA gel, hucMSCs-HA gel was applied in 2 in vivo models (types I and II DFU), and a molecular mechanism was investigated. To evaluate the paracrine action of hucMSCs, hucMSCs-conditional medium (MSC-CM) was collected to treat 1 in vivo model (type I DFU) and 2 in vitro models (high glucose (HG)-injured human umbilical vein endothelial cells (HUVECs) and human skin fibroblasts (HSFs)). The results indicated that HA gel with a porous microstructure underwent over 90% degradation and swelled to the maximum value within 48 h. In vivo, hucMSCs-HA gel accelerated wound healing of DFU rats by improving re-epithelialization, collagen deposition, and angiogenesis, in which a paracrine action of hucMSCs was confirmed and the phosphorylation of p38, ERK1/2, JNK, and Akt was increased. In vitro, MSC-CM improved cell viability, wound healing, migration, tube formation, cell senescence, and abnormal expressions (TNF-α, IL-1β, IL-6, ET-1, p16 genes, and PCNA protein) of HUVECs, also improved cell viability, wound healing, antioxidant stress, and abnormal expressions (COL1, COL3, COL4, SOD1, SOD2 genes, and PCNA protein) of HSFs. Summarily, noninvasive external application of hucMSCs-HA gel shows great perspective against DFU and exerts wound healing effects through the MAPK and Akt pathways-mediated paracrine mechanism.