Mechanisms of adipocyte dysfunction in metabolic disease and cancer

Adipose tissue (AT) dysfunction is a hallmark of unhealthy obesity and links it to metabolic comorbidities. Dysfunctional AT exhibits inhibited adipogenesis, impaired lipid metabolism, and increased inflammation. However, we are still lacking a sufficient understanding of the mechanisms of adipocyte dysfunction and how they contribute to conditions such as insulin resistance, type 2 diabetes, non-alcoholic fatty liver disease, and cancers. MiR- 107, miR-221-3p, and thyroid hormone responsive protein (THRSP) have been shown to play a role in murine models of metabolic disease in AT or other tissues, but the data in human adipocytes was limited or function unknown. Increasing knowledge of adipocyte dysfunction is critical for the development of new approaches to treat obesity and prevent its metabolic comorbidities. This thesis investigates the differentiation and function of human adipocytes and how they are altered in metabolic disease. The aim was to modulate the expressions of miR-107, miR-221-3p, or THRSP and study how adipocyte differentiation and metabolic functions were affected. In study I, we revealed miR-107 to inhibit Simpson-Golabi Behmel syndrome (SGBS) adipogenesis and lipid storage. We showed that the differentiation defect is mediated via downregulation of cyclin-dependent kinase 6 (CDK6) and Notch3. In mature adipocytes, miR-107 impaired glucose uptake and triglyceride (TG) synthesis. Our results suggest that by distinct mechanisms, it promotes insulin resistance may contribute to ectopic lipid storage by impairing function of pre- and differentiated adipocytes. Study II focused on the role of miR-221-3p in the terminal differentiation of adipocytes and in cancer-associated adipocytes (CAAs). We showed that it inhibited terminal differentiation and identified 14-3-3γ as a potential mediator of the differentiation defect. We measured the lipidome in miR-221- 3p overexpressing adipocytes and analyzed the mechanisms behind the alterations. MiR-221-3p inhibited de novo lipogenesis (DNL) of TG, but increased ceramide and sphingomyelin concentrations while reducing diglycerides (DG). MiR-221-3p is connected to many cancers; here we showed that miR-221-3p expression was elevated in tumor proximal adipose tissue from patients with invasive breast cancer. Moreover, conditioned medium from miR-221-3p transfected adipocytes increased invasion and proliferation of cancer cells in vitro. Taken together, we characterized the role of miR-221- 3p in adipocyte dysfunction and showed that the alterations in CAAs are relevant for cancer progression. In study III, we found THRSP to be induced by insulin in humans by using the euglycemic insulin clamp technique in 36 subjects. We measured a 5- and 8-fold increase in THRSP mRNA after 180 and 360 min of in vivo euglycemic hyperinsulinemia. We identified functions of THRSP by conducting a transcriptomic analysis of THRSP-silenced adipocytes. We measured mitochondrial respiration, oxidation of radiolabeled oleate, and uptake of glucose, observing that THRSP silencing impaired mitochondrial function. Lipidomic analysis of THRSP-silenced adipocytes exhibited decreased hexosylceramide concentrations, accompanied by altered expression of sphingolipid metabolism genes. In summary, THRSP induction is impaired in insulin resistance. Its downregulation in human adipocytes could promote mitochondrial dysfunction and impaired sphingolipid metabolism may contribute to metabolic dysfunction. In conclusion, these findings reveal and demonstrate different mechanisms of adipocyte dysfunction. Adipocyte differentiation and function are impaired by overexpression of miR-107 and miR-221-3p, and downregulation of THRSP expression.