An in vitro and in vivo Assessment of the Effects of Alpha-MSH on Skeletal Muscle Glucose Uptake in Healthy Human Volunteers

Diabetes mellitus is the fastest growing chronic condition globally. In 2021 the International Diabetes Federation estimated that 1 in 10 (537 million) adults are living with diabetes, with diabetes-related healthcare expenditure increasing by 316% over the last 15 years. Postprandial hyperglycaemic excursions are a key target for most therapeutic strategies, due to glucotoxicity being a major risk factor for the development of diabetic complications. Progression of disease states in both T1DM and T2DM can result in the need for intensive insulin therapy to manage hyperglycaemia. This can result in an increased incidence of hypoglycaemic episodes and chronic weight gain. Consequently, there is a rationale and need for novel pharmacological approaches which may compliment the glucose-lowering response of insulin, thus providing an insulin-sparing effect and may hold promise as use for a future adjunct therapy. Prof. Michael Cowley (Monash University, Melbourne) has published pre-clinical data demonstrating that through melanocortin receptor 5 agonism, pituitary derived alpha-melanocyte stimulatory hormone (a-MSH), improves post-prandial glucose tolerance by inducing glucose uptake into skeletal muscle via an insulin-independent pathway. This thesis aimed to take the first steps in translating these findings to human biology and assess whether targeting of this novel mechanism may hold therapeutic potential in the pharmacological management of glucose homeostasis. Healthy adult volunteers (n=6) were recruited to donate vastus lateralis muscle biopsies, which were processed to isolate myogenic satellite cells that were used to establish primary human myoblast cell lines. These cells lines were characterized and found to be free from fibroblast contamination, capable of differentiating into myotubes, and were insulin responsive. Additionally, an insulin-resistant myotube model was developed in which ectopic lipid accumulation was induced by chronic incubation with palmitic acid. The melanocortin receptor (MCR) profile of the six established cell lines was characterized. MCR1, MCR3, and MCR4 were found to be expressed at stable levels, whilst low levels of MCR5 was also found. Incubation of myotubes with a-MSH significantly increased glucose uptake over basal conditions, with a similar effect size to insulin. In insulin resistant myotubes, the glucose uptake response to a-MSH was ablated. Incubation of a-MSH with insulin resulted in an additive effect on glucose uptake in myotubes. Specific agonism of MCR5 with the peptidomimetic PG-901 increased glucose uptake over control, suggesting that the effect may be in part MCR5 mediated. In commercially obtained primary human adipocytes, MCR1 and MCR3 were both expressed but a-MSH had no impact on glucose uptake. 15 volunteers were recruited into a study investigating whether a-MSH infusion could improve post-prandial glucose tolerance during an oral glucose tolerance test (OGTT) and whole-body glucose uptake during a hyperinsulinaemic-euglycaemic clamp. Participants first underwent four OGTT visits, in which they were infused with placebo or one of three doses of a-MSH in a randomized, double-blind, crossover manner. The highest dose of a-MSH significantly reduced glucose area under the curve (AUC) in concert with a concomitant reduction in plasma insulin AUC. This effect was most pronounced during the hyperglycaemic peak of the OGTT (30-60 minutes). The same cohort of participants then underwent hyperinsulinaemic-euglycaemic clamp studies while being infused with either saline or 150 ng/kg/hr a-MSH in a randomized, double-blind, crossover manner. Glucose infusion rate was consistently increased in the presence of a-MSH. No adverse events were recorded during OGTT or clamp experiments.