Exploring the Morphologic Characteristics of Intraperitoneally Injected Three-Dimensional-Cultured Tonsil-Derived Mesenchymal Stem Cells in a Murine Colitis Model Using Electron Microscopy

Three-dimensional (3D)-cultured tonsil-derived mesenchymal stem cells (TMSCs) show high therapeutic efficacy in murine colitis. We evaluated the in vivo localization and formation of 3D-TMSCs using electron microscopy. Mice with dextran sulfate sodium-induced chronic colitis received intraperitoneal injections of 3D-TMSCs. The formation and localization of TMSC aggregates in the peritoneal cavity were assessed by immunofluorescence and electron microscopy. In mice treated with 3D-TMSCs, white spherical aggregates attached to the peritoneal cavity were observed. Immunofluorescence staining revealed the co-localization of human-origin TMSCs and mouse immune cells within these aggregates. The presence of human glyceraldehyde-3-phosphate dehydrogenase (GAPDH) confirmed the presence of human-origin TMSCs in the aggregates. Electron microscopy revealed that 3D-TMSCs were covered with mouse immune cells. The presence of pseudopodia and microvilli facilitated cell-to-cell connections, indicating a complex integrated structure rather than a mere collection of TMSCs. After 15 days, rough endoplasmic reticulum and mitochondria were identified within the cytoplasm of the TMSCs. Prominent autophagosomes and extracellular vesicles were observed within the intraperitoneal 3D-TMSC aggregates. These findings underscore the viability and paracrine effects of 3D-TMSCs and support their potential as an advanced therapeutic option for treating inflammatory bowel disease.