OBJECTIVE: Obesity and insulin resistance in men are linked to decreased testosterone and increased estradiol (E2) levels. Aromatase (ARO) converts testosterone into E2, and this occurs mainly in adipose tissue in men. E2 acts through estrogen receptors ESR1 and ESR2, and they potentially affect development of type 2 diabetes (T2D). This study explored alterations in ARO, ESR1 and ESR2 in men with obesity or T2D. METHODS: Subcutaneous adipose tissue (SAT) from men with or without obesity or T2D was analyzed for ARO, ESR1, and ESR2 gene and protein expression. Data were compared across groups and correlated with markers of obesity, glycaemia, insulin resistance, and sex hormones. Moreover, SAT was incubated with E2 or testosterone for ex vivo glucose uptake measurements. RESULTS: Aromatase ARO levels were higher in SAT from men with obesity compared to non-obese men, and gene expression correlated positively with adiposity, hyperglycaemia and insulin resistance. No association was found between ARO and circulating E2. Men with obesity had lower levels of ESR1 and ESR1:ESR2 ratio, but not ESR2. ESR1 gene expression in SAT correlated negatively with adiposity and insulin resistance markers as well as with ARO expression, and tended to be lower in men with T2D. E2 reduced insulin-stimulated glucose uptake, while testosterone increased basal glucose uptake in adipocytes. CONCLUSION: Elevated ARO in SAT was found in obese men, and this was linked to insulin resistance and glycaemia, supporting that local estrogen production contributes to metabolic dysregulation. ESR1 was reduced in men with T2D and was linked to adiposity and insulin resistance. Taken together, high ARO and low ESR1 expression in SAT in obese men may contribute to insulin resistance and T2D development.