单细胞解析骨性关节炎分子机制及其潜在干细胞治疗

      骨性关节炎（Osteoarthritis , OA）是一种中老年人群中常见的慢性、退行性的关节疾病，主要表现为软骨细胞胞外基质降解、软骨组织退化、软骨下骨异常和滑膜出现炎症等病理特征。在临床上，骨性关节炎患者主要表现为关节疼痛、关节肿胀、关节活动受限和关节畸变等症状，严重时需要进行费用昂贵的关节置换手术，后期甚至会导致残疾，严重影响着患者的生活质量。全球受骨性关节炎疾病影响的人口总数约有5.28亿，整体呈现随年龄增长而增高的趋势。骨性关节炎的致病因素包括年龄、性别、肥胖和遗传等。然而，骨性关节炎发生的具体分子机制尚不完全清楚。软骨细胞被认为是关节软骨中的唯一细胞类型，但人们对软骨细胞异质性的认识不清晰，阻碍了人们对骨性关节炎发病机制的理解。

      本研究首先使用人健康膝关节软骨组织中软骨细胞的单细胞转录组，鉴定了9个软骨细胞亚群，分别是稳态软骨细胞（homeostatic chondrocyte，HomC）、增殖性软骨细胞（proliferate chondrocyte，ProC）、肥大软骨细胞前体细胞（prehypertrophic chondrocyte，PreHTC）、肥大软骨细胞-1（hypertrophic chondrocyte-1，HTC-1）、调节性软骨细胞（regulatory chondrocyte，RegC）、肥大软骨细胞-2（hypertrophic chondrocyte-1，HTC-2）、纤维化软骨细胞前体细胞（prefibrochondrocyte，PreFC）、纤维化软骨细胞（fibrochondrocyte，FC）和增殖性软骨细胞（proliferate fibrochondrocyte，ProFC）。本研究发现了两个不同的HTC亚群，其中HTC-1特异性表达与细胞凋亡和程序性细胞死亡相关的基因。另外，本研究鉴定了软骨细胞的两个分化轨迹，软骨细胞纤维化轨迹（ProC→PreHTC→HTC-2→PreFC→FC）和软骨细胞凋亡轨迹（ProC→PreHTC→HTC-1）。本研究还比较了健康软骨和骨性关节炎软骨中不同软骨细胞亚群表达谱的差异，新鉴定出了增殖性软骨细胞-2（proliferate fibrochondrocyte-2，ProFC-2）亚群，它可能通过炎症促进骨性关节炎的发展。本研究还发现在膝盖软骨组织骨性关节炎病变后，ProC和PreHTC等处于软骨细胞分化轨迹起始位置的细胞亚群群体出现一定程度的萎缩，而PreFC、FC和HTC-1等处于软骨细胞分化轨迹末端的细胞亚群出现了一定程度的扩增，说明软骨细胞分化轨迹起点位置细胞的减少和末端位置细胞的增多，会促进骨性关节炎的发展。

      关节软骨结构和功能的异常是骨性关节炎的主要病理特征，骨性关节炎的有效治疗需要修复损伤的软骨组织，使其恢复完整的结构，因而使用干细胞辅助修复损伤软骨是治疗骨性关节炎的有效方法。间充质干细胞（mesenchymal stem cells，MSCs）是起源于中胚层的多能干细胞，具有自我更新能力、高度增殖能力和多向分化能力。它的来源十分广泛，可以从骨髓、脐带、脂肪、牙髓和滑膜等多种组织中分离得到，是再生医学和自身免疫性疾病治疗中理想的细胞来源。因此比较不同组织来源的间充质干细胞的特征是优化间充质干细胞治疗骨性关节炎临床应用的基础。本研究分析了来自脐带、骨髓、滑膜组织和脂肪组织的间充质干细胞的单细胞RNA-seq数据，发现所有间充质干细胞样本中均存在成软骨间充质干细胞亚群，因而具有骨性关节炎软骨修复的治疗潜能。另外，本研究发现脐带间充质干细胞的细胞干性和免疫调节特性高于其他组织来源的间充质干细胞，说明它是避免免疫应答的理想细胞来源。本研究还发现脐带间充质干细胞中成软骨间充质干细胞亚群数量占比高于其他组织来源的间充质干细胞，且脐带间充质干细胞成软骨分化诱导后，成软骨间充质干细胞亚群比例增加，同时表现出更明显的软骨特征，说明脐带间充质干细胞是软骨修复和骨性关节炎治疗的理想干细胞。另外本研究还鉴定了诱导成软骨调控的基因和转录因子，为提高诱导效率提供了更好的靶点。

      综上所述，本研究构建了关节软骨的细胞图谱，增强了人们对关节软骨中软骨细胞异质性的认识，研究了软骨细胞在骨性关节炎中的变化，发现了骨性关节炎的分子机制，并为间充质干细胞治疗骨性关节炎提供了理论基础。

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