Department of Biomedical Engineering, University of Kentucky, Lexington, KY, United States.
Front Immunol. 2022 Aug 18;13:987032. doi: 10.3389/fimmu.2022.987032. eCollection 2022.
Mesenchymal stromal cells (MSC) are sensors of inflammation, and they exert immunomodulatory properties through the secretion of cytokines and exosomes and direct cell-cell interactions. MSC are routinely used in clinical trials and effectively resolve inflammatory conditions. Nevertheless, inconsistent clinical outcomes necessitate the need for more robust therapeutic phenotypes. The immunomodulatory properties of MSC can be enhanced and protracted by priming (aka licensing) them with IFNγ and TNFα. Yet these enhanced properties rapidly diminish, and prolonged stimulation could tolerize their response. Hence a balanced approach is needed to enhance the therapeutic potential of the MSC for consistent clinical performance. Here, we investigated the concentration-dependent effects of IFNγ and TNFα and developed gelatin-based microgels to sustain a licensed MSC phenotype. We show that IFNγ treatment is more beneficial than TNFα in promoting an immunomodulatory MSC phenotype. We also show that the microgels possess integrin-binding sites to support adipose tissue-derived MSC (AD-MSC) attachment and a net positive charge to sequester the licensing cytokines electrostatically. Microgels are enzymatically degradable, and the rate is dependent on the enzyme concentration and matrix density. Our studies show that one milligram of microgels by dry mass can sequester up to 641 ± 81 ng of IFNγ. Upon enzymatic degradation, microgels exhibited a sustained release of IFNγ that linearly correlated with their degradation rate. The AD-MSC cultured on the IFNγ sequestered microgels displayed efficient licensing potential comparable to or exceeding the effects of bolus IFNγ treatment. When cultured with proinflammatory M1-like macrophages, the AD-MSC-seeded on licensing microgel showed an enhanced immunomodulatory potential compared to untreated AD-MSC and AD-MSC treated with bolus IFNγ treatment. Specifically, the AD-MSC seeded on licensing microgels significantly upregulated , , and , and downregulated in M1-like macrophages compared to other treatment conditions. These licensing microgels are a potent immunomodulatory approach that shows substantial promise in elevating the efficacy of current MSC therapies and may find utility in treating chronic inflammatory conditions.
间充质基质细胞 (MSC) 是炎症的传感器，它们通过细胞因子和外泌体的分泌以及直接的细胞-细胞相互作用发挥免疫调节作用。MSC 通常用于临床试验，并有效地解决炎症状态。然而，不一致的临床结果需要更强大的治疗表型。MSC 的免疫调节特性可以通过用 IFNγ 和 TNFα 对其进行预处理（又名许可）来增强和延长。然而，这些增强的特性会迅速减弱，而延长的刺激可能会使它们的反应耐受。因此，需要一种平衡的方法来增强 MSC 的治疗潜力，以实现一致的临床效果。在这里，我们研究了 IFNγ 和 TNFα 的浓度依赖性效应，并开发了基于明胶的微凝胶来维持许可的 MSC 表型。我们表明，IFNγ 处理比 TNFα 更有利于促进免疫调节 MSC 表型。我们还表明，微凝胶具有整合素结合位点，以支持脂肪组织衍生的 MSC (AD-MSC) 附着，并且具有净正电荷以静电方式隔离许可细胞因子。微凝胶可被酶降解，降解速率取决于酶浓度和基质密度。我们的研究表明，每毫克干重的微凝胶最多可以隔离 641 ± 81ng 的 IFNγ。在酶降解后，微凝胶表现出与降解速率成线性相关的 IFNγ 持续释放。在 IFNγ 隔离的微凝胶上培养的 AD-MSC 显示出与 bolus IFNγ 处理相当或超过的高效许可潜力。当与促炎 M1 样巨噬细胞共培养时，与未经处理的 AD-MSC 和用 bolus IFNγ 处理的 AD-MSC 相比，接种在许可微凝胶上的 AD-MSC 显示出增强的免疫调节潜力。具体而言，与其他处理条件相比，接种在许可微凝胶上的 AD-MSC 显著上调了 、 、 和 ，并下调了 M1 样巨噬细胞中的 。这些许可微凝胶是一种有效的免疫调节方法，在提高当前 MSC 治疗的疗效方面具有很大的潜力，并且可能在治疗慢性炎症疾病方面具有实用价值。
