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A key issue in the field of tissue engineering and stem cell therapy is immunological rejection after the implantation of allogeneic bone marrow-derived mesenchymal stem cells (BMSCs). In addition, maintaining the immunoregulatory function of BMSCs is critical to achieving tissue repair. In recent years, scientists have become interested in fish collagen because of its unique osteoinductive activity. However, it is still unclear whether osteogenically differentiated BMSCs induced by fish collagen maintain their immunoregulatory functions. To address this question, BMSCs were isolated from 8-week-old male BALB/c mice, and a noncontact coculture model was established consisting of macrophages and BMSCs treated with hydrolyzed fish collagen (HFC). Cell proliferation of the macrophages was determined by MTT. The gene and protein expression levels of the M1 and M2 macrophage markers were measured by real-time PCR and enzyme-linked immunosorbent assay (ELISA). To study the role of TNF-α-induced gene/protein 6 (TSG-6), TSG-6 was targeted by short interfering RNA (siRNA) in BMSCs, then the osteogenic differentiation ability of the BMSCs was examined by western blotting. The mRNA expression levels of interleukin-10 (IL-10), CCL22 (a macrophage-derived chemokine), tumor necrosis factor α (TNF-α), and interleukin-12 (IL-12), and the protein expression levels of arginase-1 (Arg-1) and inducible nitric oxide synthase (iNOS) of macrophages cocultured with TSG-6-siRNA-BMSCs+HFC were detected by real-time PCR and western blotting, respectively. The results showed that the osteogenically differentiated BMSCs induced by HFC did not affect the proliferation of macrophages. Osteogenically differentiated BMSCs induced by HFC promoted the expression of M2 macrophage markers IL-10 and CCL22, while HFC inhibited the expression of M1 macrophage markers, including TNF-α and IL-12. The TSG-6 knockdown led to a decrease in the production of TSG-6 without impairing the expression of bone sialoprotein (BSP), osteocalcin (OCN), and Runt-related transcription factor 2 (RUNX2) by BMSCs. TSG-6 silencing significantly counteracted the effect of HFC, and the expression of IL-10, CCL22, and Arg-1 were all decreased in the macrophages cocultured with TSG-6-siRNA-BMSCs+HFC, while that of TNF-α, IL-12, and iNOS were increased relative to the BMSCs+HFC group. The data demonstrated that osteogenically differentiated BMSCs induced by fish collagen retained their immunomodulatory functions. This study provides an additional scientific basis for future applications of fish collagen as an osteogenic component in the fields of tissue engineering and stem cell therapy.

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Impact of Marine-Based Biomaterials on the Immunoregulatory Properties of Bone Marrow-Derived Mesenchymal Stem Cells: Potential Use of Fish Collagen in Bone Tissue Engineering