Introduction: Ulcerative colitis is an inflammatory disease characterized by immune dysregulation, mucosal damage, and limited efficacy to conventional therapies due to systemic immune dysregulation and the risk of opportunistic infections. However, mesenchymal stem cells (MSC) demonstrated a promising role in immunomodulatory and regenerative properties to overcome the conventional limitation. Herein, we hypothesized a novel platform, namely surface triggered in situ gelation (STIG), for co-encapsulation of MSC spheroids and rapamycin (RAP) depots within alginate to improve therapeutic outcomes.

Methods: MSC spheroids were prepared by using AggreWell plates and RAP microspheres were fabricated via the oil-in-water emulsion method. Co-encapsulation was achieved through the STIG method. In vitro, characteristic analysis such as morphology was observed by scanning electron microscopy, drug-releasing profile by HPLC, cytotoxicity through live/dead imaging, and protein expression studies using western blot and PCR. Co-encapsulated spheroid-Rap depots were administered intraperitoneally to dextran sulfate sodium (DSS) induced chronic mouse model and therapeutic potential was evaluated based on disease activity index (DAI), histopathology, and cytokine profiling.

Results: The in vitro results depicted that RAP depots (1-8 µm) showed a sustained release of RAP for up to 50 days with 3.6 % loading capacity. The depots were uniformly distributed in the microgel of thickness < 100 μm. Notably, co-encapsulated spheroids showed reduced cell apoptosis and improved expression of anti-inflammatory modulators, such as Bcl-2, Cox-2, and IL-10. In vivo, biodistribution studies showed the significant retention of the co-encapsulated group compared to other groups. In DSS induced in vivo model, co encapsulated mice group showed significant improvement in body weight, reduction in DAI, and preservation of colonic histology. Colonic tissues evidenced decreased levels of pro-inflammatory mediators. Moreover, the histological analysis showed a significant reduction in inflammatory infiltration and ulceration which suggested the synergistic role of co-encapsulation.

Conclusion: Thus, the successful preparation of co-encapsulated drug depot and spheroid has shown an improvement with sustained drug delivery, immunomodulatory and regenerative potential which evidenced its significant therapeutic approach in the application of colitis and other inflammatory diseases.