脂肪间充质干细胞源性外泌体经let-7/TSP-1/TGF-β1轴调控口腔黏膜下纤维化水平的作用机制研究

Oral submucosal fibrosis (OSF) is a serious threat to human health. Submucosal fibrosis is a landmark event. How to reverse the fibrosis phenotype of fibroblast is a key in the treatment of OSF. Studies have confirmed that stem cell-derived exosomes have potential effects on reversing organ fibrosis, but their regulatory effects on oral microenvironment have not been paid attention to and the mechanism has not been discussed also. Our previous studies have confirmed that extracellular particles can remodel the molecular phenotype of fibroblasts. And the expression of thrombospondin-1 (TSP-1) was up-regulated and abundance of has-let-7 was decreased in OSF lesions. It was also demonstrated that let-7 is enriched in human adipose-derived stem cells (hADSCs)-derived exosomes. Based on the potential role of let-7 in the regulation of TSP-1, this study further proposes that hADSCs-derived exosomes could decrease the level of oral submucosal fibrosis, and emphasizes on: (1) the role and mechanism of TSP-1 in the regulation of oral fibroblast fibrosis; (2) the role and mechanism of let-7 in the regulation of TSP-1/TGF-β1 axis by hADSCs-derived exosomes during reprogramming of oral fibroblast fibrosis phenotype. This study is expected to reveal the potential value and molecular mechanism of hADSCs-derived exosomes in the treatment of OSF.

口腔黏膜下纤维性变（OSF）严重危害人类健康，黏膜下基质纤维化是其标志性事件，如何逆转成纤维细胞纤维化分子表型是治疗本病的关键问题。研究证实干细胞来源的外泌体对改善器官纤维化有潜在作用，而其对口腔微环境的调控作用尚未被关注，机制亦未被探讨。申请人前期研究证实：细胞外微粒可重塑成纤维细胞分子表型；在OSF病变中凝血酶敏感蛋白1（TSP-1）表达上调而has-let-7丰度降低；且证明了人脂肪间充质干细胞（hADSCs）外泌体富集let-7。基于let-7调控TSP-1的潜在作用，本课题拟进一步提出hADSCs源性外泌体可改善口腔黏膜下纤维化水平，并集中探讨：①TSP-1调控口腔成纤维细胞纤维化的作用和机制；②hADSCs源性外泌体经let-7调控成纤维细胞TSP-1/TGF-β1轴并重编程口腔成纤维细胞纤维化表型的作用和机制。本研究有望揭示hADSCs外泌体在治疗OSF的潜在价值和分子机制。

口腔粘膜下纤维性变（Oral Submucous Fibrosis, OSF）是一种慢性、隐匿性疾病，被认为是口腔癌前状态，严重损害人类健康带来较大的经济社会负担。当前对OSF研究涉及临床病理、分子机制的多个方面，近年来对OSF疾病微环境以及其所导致的口腔癌微环境特点已为当下的热点之一。本课题利用诸多分子生物学技术，在体内外多维度验证和探究TSP-1在OSF发生发展中的表达和作用。创新性采用大鼠和人类OSF器官模型验证THBS1在OSF发生发展中的作用。在组织和细胞学层面上，证实了THBS1在人和大鼠的OSF相关成纤维细胞中表达升高；进一步测定活化成纤维细胞标记物，包括THBS1，α-SMA, FSP-1和COL1A1等相关成纤维相关分子标记物在成纤维细胞附着的OSF类器官(FAOs)系统中被重现。在FAOs中，槟榔刺激状态下可促进了成纤维细胞的形态激活、激活标记物表达和胶原沉积。实验验证了槟榔碱刺激可促进口腔黏膜上皮细胞分泌TGF-β1，并参与成纤维细胞中TSP-1依赖的TGF-β1/Smad信号转导。在体内模型中，发现THBS1/TGF-β1信号轴特异性抑制剂LSKL可减轻OSF模型大鼠口腔黏膜白色病变和张嘴受限，抑制口腔黏膜下组织胶原沉积，本研究为靶向THBS1治疗OSF患者提供了分子生物学基础。截止目前，本课题组较好地完成了对应的研究目标。