Pharmacological Inhibition of Gal-3 in Mesenchymal Stem Cells Enhances Their Capacity to Promote Alternative Activation of Macrophages in Dextran Sulphate Sodium-Induced Colitis

Abstract

Transplantation of mesenchymal stem cells (MSCs) reduces the severity of dextran sulphate sodium- (DSS-) induced colitis. MSCs are able to secrete Galectin-3 (Gal-3), a protein known to affect proliferation, adhesion, and migration of immune cells. We investigate whether newly synthetized inhibitor of Gal-3 (Davanat) will affect production of Gal-3 in MSCs and enhance their potential to attenuate DSS-induced colitis. Pharmacological inhibition of Gal-3 in MSCs enhances their capacity to promote alternative activation of peritoneal macrophages in vitro and in vivo. Injection of MSCs cultured in the presence of Davanat increased concentration of IL-10 in sera of DSS-treated animals and markedly enhanced presence of alternatively activated and IL-10 producing macrophages in the colons of DSS-treated mice. Pharmacological inhibition of Gal-3 in MSCs significantly attenuates concentration of Gal-3 in sera of DSS-treated animals, indicating that MSCs produce Gal-3 in this disease. In conclusion, our findings indicate that Davanat could be used for improvement of MSC-mediated polarization towards immunosuppressive M2 phenotype of macrophages.

Figure 1

MSCs attenuate DSS-induced colitis. Water with 3% DSS was given to mice for 7 days; regular drinking water was fed to control mice. The concentration of Gal-3 in sera of MSCs groups (a). Survival rate of mice with colitis (b). Disease Activity Index (DAI) scored at day 7 using the following parameters: weight loss, stool consistency, and rectal bleeding (c). After DSS treatment length of the entire colon was measured (d). Histological examination was performed with hematoxylin and eosin staining (e). H&E staining images of representative colon tissues are shown at the same magnifications (100x) (e). Data presented as means ± SEM; n = 10 mice per experimental groups. ^∗ P < 0.05, ^∗∗ P < 0.01.

Figure 2

Pharmacological inhibition of Gal-3 in MSCs results in increased concentration of IL-10 in sera of DSS-treated animals. Levels of proinflammatory cytokines in the sera are shown: (a) TNF-α and (b) IL-1β. Levels of anti-inflammatory cytokines in the sera are shown: (c) IL-10 and (d) TGF-β. Values are mean ± SEM (n = 10 per group). ^∗ P < 0.05, ^∗∗ P < 0.01.

Figure 3

Pharmacological inhibition of Gal-3 in MSCs favored alternative activation of macrophages in the colon tissues of DSS-treated mice. The percentage of F4/80+ macrophages in colon tissue (a). Significant decrease in percentage of F4/80+CD206+ and F4/80+ IL-10+ macrophages in MSCs-treated mice (white bars), when compared to MSCs + Davanat-treated mice (black bars), after 7 days on DSS treatment (b and c). The percentage of IL-12- and IL-1β-producing F4/80+ macrophages (d and e). The percentages of inflammatory dendritic cells (f) and FcεRI+CD117+ mast cells (g) as well as CD3+NK1.1+NKT cells (h) are shown. Values are mean ± SEM (n = 10 per group). ^∗ P < 0.05, ^∗∗ P < 0.01.

Figure 4

Pharmacological inhibition of Gal-3 in MSCs enhances their capacity to promote alternative activation of peritoneal macrophages. The significant increase in percentage of F4/80+CD206+ macrophages in MSCs + Davanat-treated Gal-3^−/− macrophages (black bars), when compared to only MSCs-treated Gal-3^−/− macrophages (white bars) (a). The percentage of F4/80+CD11b+ macrophages is shown (a). Representative flow cytometry dot plots are shown. The level of IL-10 and TGF-β in supernatants (b). Values are mean ± SEM (n = 10 per group). ^∗ P < 0.05, ^∗∗ P < 0.01.