Background: Endotoxemia, mediated by uncontrolled immunocytes activation toward Lipopolysaccharide, could deteriorate into severe septic shock, but with limited treatment effect. Mesenchymal stem cells (MSCs), with excellent immune regulatory capacities, have displayed potential in multiple inflammatory disease treatment. Gelactin-9 (Gal-9), a newly discovered immune checkpoint, has been demonstrated to mediate immunomodulatory effect of MSCs in vitro. However, its in vivo role in alleviating endotoxemia remains to be elucidated.
Methods: MSCs (2.5×10⁵/ml) were obtained and stimulated with IFN-γ (20ng/ml) for 72 hours. Gal-9 expression on MSCs were measured by ELISA, RT-PCR, flow cytometry and immunofluorescence respectively. Then, experimental endotoxemia was induced by LPS injection (10mg/kg, i.p.), followed by the treatment with Gal-9-MSC (20ng/ml, 72 hours), MSCs and MSC+α-lactose (10.8mg/mL, 500ul, i.v.). Therapeutic effects of MSC-based treatments were assessed by monitoring murine sepsis score, survival rate, splenocyte proportion, phenotype polarization, inflammatory mediator levels and pathological manifestations. Furthermore, Gal-9 expression in multiple organs was also detected after administering the treatments.
Results: It has been found that MSCs expressed Gal-9 and its level was increased in a dose-dependent manner after being stimulated by IFN-γ. Moreover, adoptive transferred of IFN-γ pre-stimulated MSCs into endotoxemia mice was found with relieved symptoms and increased survival rate. Flow cytometry analysis indicated that Gal-9-MSC could promote macrophage polarization to M2-subtype and increase Treg ratios in spleen. Further results also demonstrated that, Gal-9-mediated MSC therapy could assist in attenuating local and circulating pro-inflammatory mediators expression (TNF-α, IL-1β, IFN-γ and iNOS), but increasing anti-inflammatory mediators expression (T-SOD and IL-35). Additionally, after administrating Gal-9-MSC, it was also found there was a significant relief in pathological manifestations, and with a higher expression of Gal-9 in liver, kidney and lung homogenate.
Conclusions: This study revealed that Gal-9 mediated therapeutic effects of MSCs in alleviating endotoxemia injury, which provides a novel idea for supplementing the research of MSC immunoregulatory mechanism, and offers an excellent candidate to be used in treatment of endotoxemia in the clinical settings.