Laboratory for Fetal and Regenerative Biology, University of Colorado Denver - Anschutz Medical Campus, University of Colorado School of Medicine, Aurora, Colorado.
Department of Surgery, Children's Hospital Colorado, Aurora, Colorado; and.
Physiol Genomics. 2017 Oct 1;49(10):541-548. doi: 10.1152/physiolgenomics.00090.2016. Epub 2017 Aug 25.
Impaired diabetic wound healing is associated with a dermal extracellular matrix protein profile favoring proteolysis; within the healing diabetic wound, this is represented by an increase in activated matrix metalloproteinase (MMPs). Treatment of diabetic wounds with mesenchymal stem cells (MSCs) has been shown to improve wound healing; however, there has not yet been an assessment of their ability to correct dysregulation of MMPs in diabetic wounds. Furthermore, there has been no prior assessment of the role of microRNA29b (miR-29b), an inhibitory regulatory molecule that targets MMP-9 mRNA. Using in vitro models of fibroblast coculture with MSCs and in vivo murine wound healing models, we tested the hypothesis that MSCs correct dysregulation of MMPs in a microRNA-29b-dependent mechanism. In this study, we first demonstrated that collagen I and III protein content is significantly reduced in diabetic wounds, and treatment with MSCs significantly improves collagen I content in both nondiabetic and diabetic wounds. We then found that MMP-9 gene expression and protein content were significantly upregulated in diabetic wounds, indicating elevated proteolysis. Treatment with MSCs resulted in a decrease in MMP-9 gene expression and protein content level in diabetic wounds 3 and 7 days after wounding. Zymographic analysis indicated that MSC treatment also decreased the amount of activated MMP-9 present in diabetic wounds. Furthermore, miR-29b expression was inversely associated with MMP-9 gene expression; miR-29b expression was decreased in diabetic wounds and diabetic fibroblast. Following treatment of diabetic wounds with MSCs, as well as in diabetic fibroblasts cocultured with MSCs, miR-29b was significantly increased. These findings suggest a potential mechanism through which MSCs enhance diabetic wound healing by improving collagen I content in diabetic wounds through decreasing MMP-9 expression and increasing miR-29b expression.
糖尿病创面愈合受损与有利于蛋白水解的真皮细胞外基质蛋白谱有关；在愈合的糖尿病创面中，这表现为激活的基质金属蛋白酶（MMPs）增加。间充质干细胞（MSCs）治疗糖尿病创面已被证明可以改善创面愈合；然而，尚未评估其纠正糖尿病创面 MMPs 失调的能力。此外，以前从未评估过 microRNA29b（miR-29b）的作用，miR-29b 是一种靶向 MMP-9 mRNA 的抑制性调节分子。通过 MSC 与成纤维细胞共培养的体外模型和体内小鼠创面愈合模型，我们检验了这样一个假设，即 MSC 通过 miR-29b 依赖的机制纠正 MMPs 的失调。在这项研究中，我们首先证明糖尿病创面中 I 型和 III 型胶原蛋白含量显著降低，MSC 治疗可显著提高非糖尿病和糖尿病创面中 I 型胶原含量。然后我们发现糖尿病创面中 MMP-9 基因表达和蛋白含量显著上调，表明蛋白水解增加。MSC 治疗可使糖尿病创面 3 天和 7 天后 MMP-9 基因表达和蛋白含量水平降低。酶谱分析表明 MSC 治疗还可减少糖尿病创面中活性 MMP-9 的含量。此外，miR-29b 表达与 MMP-9 基因表达呈负相关；糖尿病创面和糖尿病成纤维细胞中 miR-29b 表达降低。糖尿病创面经 MSC 治疗以及 MSC 与糖尿病成纤维细胞共培养后，miR-29b 表达显著增加。这些发现表明，MSC 通过降低 MMP-9 表达和增加 miR-29b 表达来改善糖尿病创面中 I 型胶原含量，从而增强糖尿病创面愈合，这可能是一种潜在的机制。
