Molecular Mechanism Of Regulation Of Macrophage Immunometabolism By Mesenchymal Stem Cells

Mesenchymal stem cells(MSCs)are multipotent stem cells existing in almost every tissue.Due to their easy access and potent abilities to regulate immune responses,MSC therapies hold great promises to treat inflammatory diseases.It is now recognized that inflammatory cytokines play important roles in the induction of immunoregulatory capabilities of MSCs.Previous works from this laboratory showed that in the presence of IFN?together with TNF?or IL-1?,human MSCs could express high levels of indoleamine 2,3-dioxygenase(IDO)and chemokines.The chemokines would drive immune cells to the neighborhood of human MSCs,where immune cells are suppressed by the enzymatic activities of IDO.Other critical human MSCs immunoregulatory factors such as TNF-stimulated gene-6(TSG-6)and cyclooxygenase-2(COX-2)are also regulated by inflammatory cues.Interestingly,although human cells could not efficiently respond to mouse inflammatory cytokines,human MSCs have been demonstrated to benefit graft vs.host disease(Gv HD)and liver fibrosis in mouse disease models.These observations suggest that besides inflammation induced immunoregulatory capabilities,human MSCs may possess other unappreciated mechanism to regulate immune responses.To investigate the inflammation independent immunomodulatory mechanism of human MSCs,we compared the effects of human umbilical cord-derived MSCs cultured under ambient,normal oxygen condition(NX-MSCs)with those expanded under low oxygen conditions(LO-MSCs)for their ability to treat MOG_35-55-induced experimental autoimmune encephalomyelitis(EAE)in mice.Although NX-MSCs could not effectively alleviate the disease symptoms,LO-MSCs dramatically reduced the clinical scores and inflammatory index of EAE.Moreover,the beneficial effects of LO-MSCs were found to be exerted through their secreted factors since LO-MSCs conditioned medium could mimic the therapeutic effects of LO-MSCs on EAE.To determine the key factors mediating the curative effects LO-MSCs,we compared the gene expression profile of NX-MSCs and LO-MSCs by microarray analysis.Among several genes highly expressed by LO-MSCs,insulin like growth factor 2(igf-2)encodes a soluble factor,IGF-2.Importantly,IGF-2 sh RNA transfection or IGF-2 neutralize antibodies co-administration abolished the beneficial effects of LO-MSCs therapies,demonstrating that IGF-2 is indispensable for the therapeutic effects of LO-MSCs on EAE.IGF-2 is an important factor regulating fetal development.However,the postnatal roles of IGF-2 remain largely elusive.We found that IGF-2 alone,in the absence of any other MSC component,was able to treat EAE,thus adding an immunoregulatory role to IGF-2.IGF-2 treatment significantly reduced the expression of IL-1?and considerably upregulated programmed death ligand 1(PD-L1)expression in central nervous system infiltrated macrophages.Interestingly,IGF-2 could not affect the phenotype of mature macrophages;rather,IGF-2 reprogrammed the responsiveness of macrophages during their maturation and enabled these macrophages to resolve inflammation in the presence of pro-inflammatory cytokines.Upon receiving pro-inflammatory cues,IGF-2-reprogrammed macrophages exhibit a PD-L1^highIL-1?^lowphenotype,and potently promote regulatory T cell differentiation.Furthermore,adoptive transfer of macrophages that were differentiated from IGF-2-treated monocytes significantly boosted regulatory T cell numbers and reduced the severity of EAE.IGF-2 directs the anti-inflammatory responsiveness of macrophages by inducing an irreversible metabolic shift towards oxidative phosphorylation(OXPHOS)through regulating their mitochondrial electron chain complex V.Oligomycin,a specific inhibitor of mitochondrial electron chain complex V,abolish the enhancement of PD-L1 expression and Treg induction abilities of IGF-2-reprogrammed macrophages.In summary,my studies demonstrated that MSCs could modulate immune responses in the absence of inflammatory cytokines and revealed a hitherto unappreciated role of IGF-2 in regulating innate immune memory of macrophages.The curative effects of IGF-2 and IGF-2-preprogrammed macrophages in autoimmune disease demonstrate that manipulating the metabolic phenotype of macrophages by factors like IGF-2 could become a new approach to treat immune disorders.