(IM07) Improved Anti-inflammatory Effects of Canine Adipose Tissue-Derived Mesenchymal Stem Cells Using 3D Culture

Background Mesenchymal stem cells (MSCs) have been utilized in veterinary medicine for their anti-inflammatory potential. This study explores whether 3D culture improves the anti-inflammatory effects of canine adipose tissue-derived MSCs (cAT-MSCs). Hypothesis/Objectives This study aimed to optimize the culture conditions for cAT-MSC spheroids, assess the stemness of the spheroids, and examine the enhancement of anti-inflammatory factor expression. Animals Canine adipose tissues were collected from healthy donors that had been spayed and neutered at Seoul National University (SNU) Veterinary Medical Teaching Hospital under a protocol approved by the Institutional Animal Care and Use Committee of SNU. Methods cAT-MSC spheroids were generated using ultra-low adhesion plates and their structure and hypoxic conditions were analyzed via confocal imaging. Western blot was used to analyze SOX2 and OCT4 expression. Inflammatory markers were assessed using RT-PCR and ELISA. Conditioned media from cAT-MSC spheroids was applied to canine macrophages (DH82), with pro-inflammatory factor expression and M1 polarization analyzed using RT-PCR and flow cytometry. Results 3D culture induced hypoxic conditions in cAT-MSC spheroids and increased their SOX2 and OCT4 expression (p< 0.05). Anti-inflammatory markers, including TGF-β1, TSG-6, COX-2, PGE2, and IL-10, were elevated in cAT-MSC spheroids (p< 0.05). Conditioned media from cAT-MSC spheroids reduced expression of pro-inflammatory factors TNF-α, IL-1β, and IL-6 (p< 0.01), and decreased M1 polarization of DH82 cells (p< 0.01). Conclusions and Clinical Importance 3D culture enhances the anti-inflammatory properties of cAT-MSCs, suggesting that cAT-MSC spheroids may provide a more effective therapeutic approach for managing canine inflammatory diseases.