363-OR: Matrix Metalloprotease-2 as a Novel Regulator of Glucose Utilization by Adipocytes and High-Fat Diet-Induced Glucose Intolerance

Obesity and its related comorbidities, such as glucose intolerance and hyperglycemia, pose risks for type 2 diabetes. Studies on the murine models of high-fat diet (HFD)-induced obesity revealed that glucose transporter 4 (GLUT4) expression on white adipocytes is critical for absorption of excess glucose and maintenance of whole-body glucose homeostasis. Matrix metalloproteases (MMPs) play a crucial role in remodeling the white adipose tissue (WAT) during obesity. MMPs have multiple protein substrates, surprisingly, it is unknown if MMPs can directly cleave GLUT4 on adipocyte surface and impair glucose absorption. To determine the highly active MMP expressed in the WAT during obesity, we placed male C57BL/6J mice on an HFD for 5, 9, and 11 weeks or on a standard chow diet, examined the development of hyperglycemia, inflammation, and MMP activity in the gonadal WAT using gelatin zymography. Flow cytometry studies on gonadal WAT confirmed an increase in inflammatory immune cell infiltration, and RT-PCR analysis confirmed elevated TNF-α in late stages of obesity. Zymography identified MMP2 as the highly active MMP in the WAT after HFD. Next, we used the Seahorse extracellular flux analyzer to determine the metabolic fitness of differentiating 3T3-L1 adipocytes after treatment with MMP2 for 2 hours. Glycolysis stress tests revealed MMP2, even at 200 ng/ml, attenuated glucose utilization, as indicated by 30% decreased basal glycolysis, 50% decrease in glycolytic capacity, and blunted glycolytic reserve. Further, in silico structural analysis using AlphaFold identified a potential MMP2 cleavage site on the extracellular domain of both murine and human GLUT4. This encouraged us to hypothesize that MMP2 reduces glucose utilization in differentiating adipocytes by cleaving GLUT4 extracellular domains. This is a novel mechanism that may add to insulin resistance to promote hyperglycemia and suggests inhibition of MMP2 would promote healthy obesity.