BackgroundGanglioside GM3 mediates adipocyte insulin resistance, but the role of GM3 in diabetic wound healing, a major cause of morbidity, is unclear.PurposeDetermine whether GM3 depletion promotes diabetic wound healing and directly activates keratinocyte insulin pathway signaling.ResultsGM3 synthase (GM3S) expression is increased in human diabetic foot skin, ob/ob and diet-induced obese diabetic mouse skin, and mouse keratinocytes exposed to increased glucose. GM3S knockout in diet-induced obese mice prevents the diabetic wound healing defect. Keratinocyte proliferation, migration, and activation of insulin receptor (IR) and insulin growth factor-1 receptor (IGF-1R) are suppressed by excess glucose in wild type cells, but increased in GM3S −/− keratinocytes with supplemental glucose. Co-immunoprecipitation of IR, IR substrate-1 (IRS-1), and IGF-1R, and increased IRS-1 and Akt phosphorylation accompany receptor activation. GM3 supplementation or inhibition of IGF-1R or PI3K reverses the increased migration of GM3S−/− keratinocytes, whereas IR knockdown only partially suppresses migration.ConclusionsCutaneous GM3 accumulation may participate in the impaired wound healing of diet-induced diabetes by suppressing keratinocyte insulin/IGF-1 axis signaling. Strategies to deplete GM3S/GM3 may improve diabetic wound healing.

GM3, the simplest ganglioside, regulates cell proliferation, migration, and invasion by influencing cell signaling at the membrane level. Although the classic N-acetylated form of GM3 (NeuAcLacCer) is commonly expressed and has been well studied, deacetylated GM3 (NeuNH 2 LacCer, d-GM3) has been poorly investigated, despite its presence in metastatic tumors but not in noninvasive melanomas or benign nevi. We have recently found that d-GM3 stimulates cell migration and invasion by activating urokinase plasminogen activator receptor (uPAR) signaling to augment matrix metalloproteinase-2 (MMP-2) function. However, the mechanisms by which d-GM3/ uPAR increase MMP-2 expression and activation are not clear. By modifying the expression of d-GM3 genetically and biochemically, we found that decreasing d-GM3 expression inhibits, whereas overexpressing d-GM3 stimulates, p38 mitogen-activated protein kinase (MAPK) activity to influence MMP-2 expression and activation. p38 MAPK (p38) activation requires the formation of a membrane complex that contains uPAR, caveolin-1, and integrin a5b1 in membrane lipid rafts. In addition, knocking down or inhibiting focal adhesion kinase (FAK), phosphoinositide 3-kinase (PI3K), or Src kinase significantly reduces d-GM3-induced p38 phosphorylation and activation. Taken together, these results suggest that d-GM3 enhances the metastatic phenotype by activating p38 signaling through uPAR/integrin signaling with FAK, PI3K, and Src kinase as intermediates. Elucidation of the mechanisms by which d-GM3, a newly discovered, potential biomarker of metastatic melanomas, promotes cell metastasis will help us to understand the function of d-GM3 in metastatic melanomas and may lead to novel GM3-based cancer therapies. Mol Cancer Res; 11(6); 665-75. Ó2013 AACR. IntroductionAlthough recent therapeutic progress for metastatic melanomas has significantly improved, including the more recent availability of small-molecule inhibitors targeting BRAF mutation or both BRAF mutation and MEK, or anti-CTLA4 antibody, survival is minimally prolonged and mortality rate of metastatic melanomas remains high (1-3). It is of utmost importance to discover specific molecular biomarkers that can distinguish melanomas with metastatic propensity from more indolent ones, which will improve prognostication, allow for earlier and more efficacious treatments, and provide additional targets for novel therapy.