Department of Orthopaedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, USA.
Diabetes Research Institute and Cell Transplant Center, Miller School of Medicine, University of Miami, Miami, FL, USA.
Sci Rep. 2024 Nov 27;14(1):29438. doi: 10.1038/s41598-024-80050-9.
Osteoarthritis (OA) represents a significant global health burden without a known disease modifying agent thereby necessitating pursuit of innovative therapeutic approaches. The infrapatellar fat pad (IFP) serves as a reservoir of mesenchymal stem/stromal cells (MSC), and with adjacent synovium plays key roles in joint disease affecting local inflammatory responses. Therapeutically, IFP-MSC have garnered attention for their potential in OA treatment due to their immunomodulatory and regenerative properties. However, optimizing their therapeutic efficacy necessitates a comprehensive understanding of how growth medium and inflammatory/hormonal priming influence their behavior. In this study, we isolated and expanded IFP-MSC in three different growth media: DMEM + 10% fetal bovine serum (FBS), DMEM + 10% human platelet lysate (HPL), and xeno-/serum-free synthetic (XFSF) medium. Subsequently, cells were induced with an inflammatory/fibrotic cocktail (TIC) with or without oxytocin (OXT). We evaluated various parameters including growth kinetics, phenotype, immunomodulatory capacity, gene expression, and macrophage polarization capacity. Our results revealed significant differences in the behavior of MSC cultured in different media. IFP-MSC cultured in HPL and XFSF exhibited superior growth kinetics and colony-forming abilities compared to those cultured in FBS. Furthermore, both HPL and XFSF media enhanced the expression of MSC markers (> 90%) and potentiated their immunomodulatory properties. Notably, XFSF-conditioned IFP-MSC demonstrated the highest attenuation of peripheral blood mononuclear cell (PBMC) proliferation, indicating their robust immunosuppressive capacity. Additionally, TIC priming further augmented the immunomodulatory functionality of MSC, with IFP-MSC exhibiting enhanced suppression of PBMC proliferation upon TIC priming. Of particular interest, gene expression analysis revealed distinct patterns in TIC + OXT induced MSC compared to TIC only induced, with upregulation of genes associated with immunomodulatory and regenerative functions. Furthermore, TIC + OXT priming promoted M2 polarization in macrophages, suggesting a potential therapeutic strategy for immune-mediated inflammatory joint conditions including OA. Our findings highlight the critical influence of growth medium and inflammatory/hormonal priming on MSC behavior and therapeutic potential. XFSF and HPL media offer promising alternatives to FBS, enhancing MSC growth and immunomodulatory properties. Moreover, TIC + OXT priming represents a novel approach to augment MSC immunomodulation and promote M2 polarization, providing insights into potential therapeutic strategies for OA and other immune-mediated inflammatory conditions.
骨关节炎（OA）是一种严重的全球健康负担，目前尚无已知的疾病修饰药物，因此需要寻求创新的治疗方法。髌下脂肪垫（IFP）是间充质干细胞/基质细胞（MSC）的储存库，与相邻的滑膜一起在影响局部炎症反应的关节疾病中发挥关键作用。治疗方面，IFP-MSC 因其具有免疫调节和再生特性而备受关注，有潜力用于 OA 治疗。然而，要优化其治疗效果，就需要全面了解生长培养基和炎症/激素启动如何影响其行为。在这项研究中，我们在三种不同的生长培养基中分离和扩增了 IFP-MSC：DMEM+10%胎牛血清（FBS）、DMEM+10%人血小板裂解液（HPL）和无血清/无动物源合成（XFSF）培养基。随后，用炎症/纤维化鸡尾酒（TIC）和/或催产素（OXT）诱导细胞。我们评估了各种参数，包括生长动力学、表型、免疫调节能力、基因表达和巨噬细胞极化能力。我们的结果表明，在不同培养基中培养的 MSC 行为存在显著差异。在 HPL 和 XFSF 中培养的 IFP-MSC 的生长动力学和集落形成能力优于在 FBS 中培养的 MSC。此外，HPL 和 XFSF 培养基均增强了 MSC 标志物的表达（>90%），并增强了其免疫调节特性。值得注意的是，XFSF 条件培养基对外周血单核细胞（PBMC）增殖的抑制作用最强，表明其具有强大的免疫抑制能力。此外，TIC 启动进一步增强了 MSC 的免疫调节功能，TIC 启动后 IFP-MSC 对 PBMC 增殖的抑制作用增强。特别有趣的是，基因表达分析显示，TIC+OXT 诱导的 MSC 与仅 TIC 诱导的 MSC 之间存在明显不同的模式，与免疫调节和再生功能相关的基因上调。此外，TIC+OXT 启动促进了巨噬细胞向 M2 极化，这表明这可能是一种治疗免疫介导的炎症性关节疾病（包括 OA）的潜在策略。我们的研究结果强调了生长培养基和炎症/激素启动对 MSC 行为和治疗潜力的关键影响。XFSF 和 HPL 培养基是 FBS 的有前途的替代品，可增强 MSC 的生长和免疫调节特性。此外，TIC+OXT 启动代表了增强 MSC 免疫调节和促进 M2 极化的新方法，为 OA 和其他免疫介导的炎症性疾病的潜在治疗策略提供了新的思路。
