Abstract：
Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic extracellular matrix disease caused by deficiency in type VII collagen (Col VII). The disease manifests with devastating mucocutaneous fragility leading to progressive fibrosis and metastatic squamous cell carcinomas. Although Col VII abundance is considered the main predictor of symptom course, previous studies have revealed the existence of mutation-independent mechanisms that control disease progression. Here, to investigate and validate new molecular modifiers of wound healing and fibrosis in a natural human setting, and toward development of disease-modulating treatment of RDEB, we performed gene expression profiling of primary fibroblast from RDEB siblings with marked phenotypic variations, despite having equal COL7A1 genotype. Gene enrichment analysis suggested that severe RDEB was associated with enhanced response to TGF-β stimulus, oxidoreductase activity, and cell contraction. Consistently, we found an increased response to TGF-β, higher levels of basal and induced reactive oxygen species (ROS), and greater contractile ability in collagen lattices in RDEB fibroblasts (RDEBFs) from donors with severe RDEB vs mild RDEB. Treatment with antioxidants allowed a reduction of the pro-fibrotic and contractile phenotype. Importantly, our analyses revealed higher expression and deposition in skin of the relatively uncharacterized small leucine-rich extracellular proteoglycan PRELP/prolargin associated with milder RDEB manifestations. Mechanistic investigations showed that PRELP effectively attenuated fibroblasts\' response to TGF-β1 stimulus and cell contractile capacity. Moreover, PRELP overexpression in RDEBFs enhanced RDEB keratinocyte attachment to fibroblast-derived extracellular matrix in the absence of Col VII. Our results highlight the clinical relevance of pro-oxidant status and hyper-responsiveness to TGF-β in RDEB severity and progression. Of note, our study also reveals PRELP as a novel and natural TGF-β antagonist with a likely dermo-epidermal pro-adhesive capacity.
摘要：
隐性营养不良性大疱性表皮松解症（RDEB）是由VII型胶原蛋白（ColVII）缺乏引起的遗传性细胞外基质疾病。该疾病表现为破坏性的粘膜皮肤脆性，导致进行性纤维化和转移性鳞状细胞癌。尽管ColVII丰度被认为是症状过程的主要预测因子，先前的研究揭示了控制疾病进展的突变非依赖性机制的存在.这里,研究和验证在自然人类环境中伤口愈合和纤维化的新分子修饰剂，以及发展RDEB的疾病调节治疗，我们进行了来自RDEB兄弟姐妹的原代成纤维细胞的基因表达谱，具有明显的表型变异，尽管具有相同的COL7A1基因型。基因富集分析表明，严重的RDEB与对TGF-β刺激的反应增强有关，氧化还原酶活性，和细胞收缩。始终如一，我们发现对TGF-β的反应增加，较高水平的基础和诱导的活性氧（ROS），来自重度RDEB与轻度RDEB的供体的RDEB成纤维细胞（RDEBF）的胶原晶格中更大的收缩能力。用抗氧化剂处理可以减少促纤维化和收缩表型。重要的是,我们的分析显示,与轻度RDEB表现相关的相对未表征的富含小亮氨酸的细胞外蛋白聚糖PRELP/prolargin在皮肤中的表达和沉积更高.机制研究表明，PRELP可有效减弱成纤维细胞对TGF-β1刺激和细胞收缩能力的反应。此外，在没有ColVII的情况下，RDEBF中的PRELP过表达增强了RDEB角质形成细胞与成纤维细胞衍生的细胞外基质的附着。我们的结果强调了促氧化剂状态和对TGF-β的高反应性与RDEB严重程度和进展的临床相关性。值得注意的是，我们的研究还揭示了PRELP是一种新型的天然TGF-β拮抗剂，可能具有真皮-表皮前粘附能力。
