Assembly of MSCs into a spheroid configuration increases poly(I:C)-mediated TLR3 activation and the immunomodulatory potential of MSCs for alleviating murine colitis.

BACKGROUND: Inflammatory bowel disease (IBD) is associated with significant clinical challenges due to the limitations of current therapeutic approaches. Mesenchymal stem cell (MSC)-based therapies have shown promise in alleviating IBD owing to their potent immunomodulatory properties. However, the therapeutic efficacy of these cells remains suboptimal, primarily due to the harsh peritoneal microenvironment, which compromises MSC viability and functional capacity after transplantation. METHODS: To address these limitations, this study aimed to improve MSC engraftment and functionality by assembling MSCs into three-dimensional (3D) spheroids and priming them with the Toll-like receptor 3 (TLR3) agonist polyinosinic-polycytidylic acid (poly(I:C)). Their potential for treating IBD was evaluated using male C57BL/6 mice with dextran sulfate sodium-induced colitis. RESULTS: While 3D spheroid formation alone upregulated TLR3 expression and increased MSC survival under oxidative stress, poly(I:C) priming had a pronounced synergistic effect, significantly increasing MSC-mediated splenocyte modulation and oxidative stress resistance. In a murine colitis model, compared with unprimed spheroids or MSC suspensions, poly(I:C)-primed MSC spheroids administered intraperitoneally exhibited increased survival and therapeutic efficacy, effectively alleviating colitis symptoms, reducing colonic inflammation, and promoting tissue recovery. CONCLUSION: Collectively, these findings highlight the synergistic benefits of combining 3D spheroid assembly with TLR3 activation as an innovative strategy to improve the therapeutic efficacy of MSC-based treatments for IBD and other inflammatory diseases by increasing post-engraftment cell survival and immunomodulatory capacity.