Adipose cells are extrathyroidal targets of thyroid-stimulating hormone (TSH). TSH stimulates interleukin-6 (IL-6) release from adipocytes. We examined TSH responsiveness as a function of stage of differentiation or adipose tissue depot in cultured adipose cells and determined the effect of TSH on extrathyroidal IL-6 production in vivo. Stromal preadipocytes, isolated from human abdominal subcutaneous or omental adipose tissue, and their differentiated counterparts were studied. IL-6 protein concentration in the medium was measured after TSH stimulation. Basal IL-6 release was greater for preadipocytes than differentiated adipocytes, whether derived from subcutaneous or omental fat depots. A depot-dependent effect (omental > subcutaneous) on basal IL-6 release was observed for preadipocytes (1.6-fold, P < 0.05); a similar trend for differentiated adipocytes was not significant (6.2-fold, P > 0.05). IL-6 responsiveness to TSH was observed upon differentiation, but only for subcutaneous adipocytes (1.9-fold over basal, P < 0.001). To determine if TSH could stimulate IL-6 release from extrathyroidal tissues in vivo, we measured serum IL-6 levels from five thyroidectomized patients who received recombinant human (rh) TSH and found that levels increased by threefold on days 3 and 4 ( P < 0.05) after its administration. Our data demonstrate that stage of differentiation and fat depot origin affect basal and TSH-stimulated IL-6 release from adipose cells in culture. Furthermore, rhTSH elevates serum IL-6 response in thyroidectomized patients, indicating an extrathyroidal site of TSH action.