Protein kinase C mediates tumor necrosis factor-alpha-induced inhibition of obese gene expression and leptin secretion in brown adipocytes

Previously we showed that tumor necrosis factor-alpha (TNF-alpha) inhibits lipoprotein lipase (LPL) activity and its gene expression, an early marker of adipocyte differentiation, in cultured brown adipocytes. To know whether TNF-alpha also affects late events in brown adipocyte maturation, we examined the effect of TNF-alpha on obese gene expression and leptin secretion in mouse brown adipocytes differentiated in culture. TNF-alpha caused a concentration-dependent decrease in leptin accumulation in culture medium and leptin mRNA amount in brown adipocytes which constitutively express the ob gene. Time-course study showed that TNF-alpha significantly suppressed leptin secretion during incubation for 16, 24 and 48 h. Since some effect of TNF-alpha is mediated by activation of protein kinase C (PKC), the role of PKC in TNF-alpha-induced downregulation of ob gene expression and leptin secretion was studied. The suppressive effect of TNF-alpha on both ob gene expression and leptin secretion was blocked by PKC inhibitors such as bisindolylmaleimide I (BIM) and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7). Incubation of brown adipocytes with TNF-alpha (20 ng/ml, 15 min) caused a rapid shift of PKC activity from the cytosolic to the membrane fraction, suggesting an activation of PKC by TNF-alpha in brown adipocytes. This effect of TNF-alpha was blocked by a selective PKC inhibitor, BIM. These results suggest that TNF-alpha promotes dedifferentiation of the brown adipocytes as evidenced by a downregulation in ob gene expression and leptin secretion via PKC-dependent mechanisms.