PDF(Pubmed)
Abstract：
Deficient wound healing is frequently observed in patients diagnosed with diabetes, a clinical complication that compromises mobility and leads to limb amputation, decreasing patient autonomy and family lifestyle. Fibroblasts are crucial for secreting the extracellular matrix (ECM) to pave the wound site for endothelial and keratinocyte regeneration. The biosynthetic pathways involved in collagen production and crosslinking are intimately related to fibroblast redox homeostasis. In this study, two sets of human dermic fibroblasts were cultured in normal (5 mM) and high (25 mM)-glucose conditions in the presence of 1 µM selenium, as sodium selenite (inorganic) and the two selenium amino acids (organic), Se-cysteine and Se-methionine, for ten days. We investigated the ultrastructural changes in the secreted ECM induced by these conditions using scanning electron microscopy (SEM). In addition, we evaluated the redox impact of these three compounds by measuring the basal state and real-time responses of the thiol-based HyPer biosensor expressed in the cytoplasm of these fibroblasts. Our results indicate that selenium compound supplementation pushed the redox equilibrium towards a more oxidative tone in both sets of fibroblasts, and this effect was independent of the type of selenium. The kinetic analysis of biosensor responses allowed us to identify Se-cysteine as the only compound that simultaneously improved the sensitivity to oxidative stimuli and augmented the disulfide bond reduction rate in high-glucose-cultured fibroblasts. The redox response profiles showed no clear association with the ultrastructural changes observed in matrix fibers secreted by selenium-treated fibroblasts. However, we found that selenium supplementation improved the ECM secreted by high-glucose-cultured fibroblasts according to endothelial migration assessed with a wound healing assay. Direct application of sodium selenite and Se-cysteine on purified collagen fibers subjected to glycation also improved cellular migration, suggesting that these selenium compounds avoid the undesired effect of glycation.
摘要：
在诊断为糖尿病的患者中经常观察到伤口愈合不足，一种损害活动能力并导致截肢的临床并发症，降低患者自主性和家庭生活方式。成纤维细胞对于分泌细胞外基质（ECM）以修复伤口部位以促进内皮和角质形成细胞再生至关重要。参与胶原蛋白产生和交联的生物合成途径与成纤维细胞氧化还原稳态密切相关。在这项研究中,在1µM硒存在下，在正常（5mM）和高（25mM）葡萄糖条件下培养两组人皮肤成纤维细胞，亚硒酸钠（无机）和两种硒氨基酸（有机），Se-半胱氨酸和Se-蛋氨酸，十天。我们使用扫描电子显微镜（SEM）研究了这些条件诱导的分泌ECM的超微结构变化。此外,我们通过测量在这些成纤维细胞的细胞质中表达的基于硫醇的HyPer生物传感器的基础状态和实时响应来评估这三种化合物的氧化还原影响。我们的结果表明，硒化合物补充将两组成纤维细胞的氧化还原平衡推向更多的氧化色调，这种影响与硒的类型无关。生物传感器响应的动力学分析使我们能够确定Se-半胱氨酸是唯一能同时提高高葡萄糖培养的成纤维细胞对氧化刺激的敏感性并增强二硫键还原速率的化合物。氧化还原反应曲线与硒处理的成纤维细胞分泌的基质纤维中观察到的超微结构变化没有明显关联。然而,我们发现,根据伤口愈合试验评估的内皮迁移,补硒可改善高糖培养的成纤维细胞分泌的ECM.将亚硒酸钠和Se-半胱氨酸直接应用在经过糖化的纯化胶原纤维上也改善了细胞迁移，这表明这些硒化合物避免了糖化的不良影响。
