Matrix stiffness regulates the immunomodulatory effects of mesenchymal stem cells on macrophages via AP1/TSG-6 signaling pathways

It is well-recognized that the matrix stiffness as an important stem cell niche can mediate stem cell behavior such as attachment, proliferation and differentiation, but how matrix stiffness affects the immunomodulatory efficacy of stem cells has been little explored, which, however, is of significant importance in determining the outcomes of stem cell-based therapies and engineered tissue mimics. We herein studied the immunomodulatory efficacy of mesenchymal stem cells (MSCs) in response to matrix stiffness by the evaluation of macrophage polarization in vitro and inflammatory response in vivo by subcutaneous implantation of MSC-laden hydrogels. Remarkably, we found that soft matrix enabled MSCs to produce significantly higher levels of immunomodulatory factors compared to stiff matrix, and induced the presence of more anti-inflammatory macrophages in vitro and attenuated macrophages-mediated inflammatory response in vivo. More importantly, we revealed stiffness-mediated immunoregulatory effect of MSCs was mainly attributed to tumor necrosis factor-α-stimulated protein 6 (TSG-6), which was mechanosensitively regulated by the MAPK and Hippo signaling pathway and downstream AP1 complex, and which in turn exerted an effect on macrophages through CD44 receptor to inhibit NF-κB pathway. To conclude, our results for the first time identify TSG-6 as the key factor in regulating immunomodulatory efficacy of MSCs in mechanical response, and can be potentially utilized to empower stem cell-based therapy and tissue engineering strategy in regenerative medicine. STATEMENT OF SIGNIFICANCE: Matrix stiffness as an important stem cell niche can mediate stem cell behavior such as attachment and differentiation, but how matrix stiffness affects the immunomodulatory efficacy of stem cells has been little explored, which, however, is of significant importance in determining the outcomes of stem cell-based therapies and engineered tissue mimics. Our results for the first time identify TSG-6 as the key factor in regulating the immunomodulatory efficacy of MSCs in mechanical response, which was regulated by the MAPK and Hippo signaling pathways and downstream AP1 complex, and which in turn exerted an effect on macrophages through CD44 receptor to inhibit NF-κB pathway, and can be potentially utilized to empower stem cell-based therapy and tissue engineering strategy in regenerative medicine.