Osteoarthritis (OA) is a painful and debilitating disease affecting over 500 million people worldwide. Intraarticular injection of mesenchymal stromal cells (MSCs) shows promise for the clinical treatment of OA, but the lack of consistency in MSC preparation and application makes it difficult to further optimize MSC therapy and to properly evaluate the clinical outcomes. In this study, we used Sox9 activation and RelA inhibition, both mediated by the CRISPR-dCas9 technology simultaneously, to engineer MSCs with enhanced chondrogenic potential and downregulated inflammatory responses. We found that both Sox9 and RelA could be fine-tuned to the desired levels, which enhances the chondrogenic and immunomodulatory potentials of the cells. Intraarticular injection of modified cells significantly attenuated cartilage degradation and palliated OA pain compared with the injection of cell culture medium or unmodified cells. Mechanistically, the modified cells promoted the expression of factors beneficial to cartilage integrity, inhibited the production of catabolic enzymes in osteoarthritic joints, and suppressed immune cells. Interestingly, a substantial number of modified cells could survive in the cartilaginous tissues including articular cartilage and meniscus. Together, our results suggest that CRISPR-dCas9-based gene regulation is useful for optimizing MSC therapy for OA.

中文翻译：

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CRISPR 介导的 Sox9 激活和 RelA 抑制增强骨关节炎的细胞治疗


骨关节炎 (OA) 是一种痛苦且使人衰弱的疾病，影响着全世界超过 5 亿人。关节内注射间充质基质细胞（MSC）为骨关节炎的临床治疗带来了希望，但间充质干细胞的制备和应用缺乏一致性，使得进一步优化间充质干细胞治疗和正确评估临床结果变得困难。在这项研究中，我们利用 CRISPR-dCas9 技术同时介导的 Sox9 激活和 RelA 抑制来改造 MSC，使其具有增强的软骨形成潜力和下调的炎症反应。我们发现 Sox9 和 RelA 都可以微调到所需的水平，从而增强细胞的软骨形成和免疫调节潜力。与注射细胞培养基或未修饰细胞相比，关节内注射修饰细胞可显着减轻软骨退化并缓解骨关节炎疼痛。从机制上讲，修饰的细胞促进了有利于软骨完整性的因子的表达，抑制了骨关节炎关节中分解代谢酶的产生，并抑制了免疫细胞。有趣的是，大量修饰细胞可以在软骨组织中存活，包括关节软骨和半月板。总之，我们的结果表明基于 CRISPR-dCas9 的基因调控对于优化 OA 的 MSC 治疗非常有用。