MSCs Increase the Expression of Gal-1 in DCs. As shown in Figure 14, the expression of Gal-1 mRNA in the MSC + DC group (0.606 [+ or -] 0.05) was significantly higher than that in DC control group (0.413 [+ or -] 0.042); the expression of Gal-1 mRNA in the Gal-1 + DC group (1.257 [+ or -] 0.122) was significantly higher than that in DC only group.
As shown in Figure 15, expression of Gal-1 protein on DCs in the MSC + DC and Gal-1 + DC groups (1.248 [+ or -] 0.141 and 1.433 [+ or -] 0.085, resp.) was significantly higher than that in DC only group (0.319 [+ or -] 0.057).
The results of RT-PCR and Western blot showed that the expression of Gal-1 mRNA and protein on DCs was enhanced when MSCs were cocultured with DCs but inhibited by the Gal-1 inhibitor TDG.
Immunofluorescence Detected Differential Expression of Gal-1 on the Surface of DCs. As shown in Figure 16, immunofluorescence double staining and DAPI staining were performed in DC control, MSC + DC, Gal-1 + DC, and MSC + DC + TDG groups.
Gal-1 Activated the ERK Pathway and Inhibited the p38 MAPK Pathway in DCs. The MAPK signaling pathway, including ERK1/2, p38 MAPK, and JNK, is known to play an important role in differentiation of DCs and regulation of cytokine secretion .
As shown in Figure 18, the P38 MAPK phosphorylation level of DCs in the MSC + DC and Gal-1 + DC groups (0.246 [+ or -] 0.059 and 0.156 [+ or -] 0.049, resp.) was significantly lower than that in DC only group (0.928 [+ or -] 0.105), indicating that Gal-1 inhibited the p38 MAPK pathway.
In this current study, MSCs and DCs were successfully isolated from mouse bone marrow, and MSCs via an in vitro coculture were found to inhibit the maturation and function of DCs through Gal-1 secretion by regulating the MAPK pathway in DCs.
DCs are known to be the most powerful full-time antigen-presenting cells in vivo.
Previous studies showed that coculture of MSCs and DCs inhibited the differentiation, maturation, and activation of DCs by downregulating expression of costimulatory molecules such as CD80, CD86, and MHC II on the surface of DCs [24, 25].