Introduction: Leptin is a pro-inflammatory cytokine and excess leptin in obesity may induce autoimmune disease. The suppressors of cytokine signalling (SOCS) are negative regulators of cytokine signalling and therefore a target for therapy. SOCS3 modulates leptin actions but there is no data regarding other SOCS proteins and leptin signalling.
Aim: To develop a bioassay and examine the actions of SOCS proteins on leptin signalling.
Materials and Methods: MCF-7 cells were transfected with mammalian expression vectors for the leptin receptor (ObRb), SIE-luciferase reporter (STAT3 reporter), STAT3, and Renilla luciferase. Co-transfection with SOCS1-3 and CIS expression vectors was performed. Transfected cells were treated with leptin (0-100 ng/ml) for 6 hours. Luciferase activity was determined and results reported as fold induction over cells not exposed to leptin.
Results: In the absence of both ObRb and STAT3 no SIE activation was seen. The maximal fold induction in cells transfected with ObRb in the absence of a STAT3 expression vector was 4.97 plus/minus 0.64 at 100 ng/ml leptin, however co-transfection with STAT3 augmented fold induction to 21.44 plus/minus 1.9. SOCS1 and SOCS3 inhibited leptin signalling by 95.7% (p<0.0001) and 93.8% (p<0.0001), respectively. SOCS2 showed a 41% (p<0.001) reduction in leptin signalling and co-transfection of CIS showed no effect.
Conclusions: We describe a robust leptin bioassay based on the detection of STAT3 activation. SOCS1 and SOCS3 blocked leptin signalling, SOCS2 showed some inhibition but CIS showed no effect on leptin signalling.
04 - 06 Apr 2005
British Endocrine Societies