Human resistin stimulates the pro-inflammatory cytokines TNF-a and IL-12 in macrophages by NF-jB-dependent pathway Nirupama Silswal a,1 , Anil K. Singh b,1 , Battu Aruna a , Sangita Mukhopadhyay b , Sudip Ghosh a , Nasreen Z. Ehtesham a, * a Molecular Biology Unit, National Institute of Nutrition (ICMR), Hyderabad 500007, India b Laboratory of Molecular and Cellular Biology, CDFD, Hyderabad 500076, India Received 29 June 2005 Available online 14 July 2005 Abstract Resistin, a recently discovered 92 amino acid protein involved in the development of insulin resistance, has been associated with obesity and type 2 diabetes. The elevated serum resistin in human diabetes is often associated with a pro-inflammatory milieu. How- ever, the role of resistin in the development of inflammation is not well understood. Addition of recombinant human resistin protein (hResistin) to macrophages (both murine and human) resulted in enhanced secretion of pro-inflammatory cytokines, TNF-a and IL- 12, similar to that obtained using 5 lg/ml lipopolysaccharide. Both oligomeric and dimeric forms of hResistin were able to activate these cytokines suggesting that the inflammatory action of resistin is independent of its conformation. Heat denatured hResistin abrogated cytokine induction while treatment of recombinant resistin with polymyxin B agarose beads had no effect thereby ruling out the role of endotoxin in the recombinant hResistin mediated cytokine induction. The pro-inflammatory nature of hResistin was further evident from the ability of this protein to induce the nuclear translocation of NF-jB transcription factor as seen from elec- trophoretic mobility shift assays. Induction of TNF-a in U937 cells by hResistin was markedly reduced in the presence of either dominant negative IjBa plasmid or PDTC, a pharmacological inhibitor of NF-jB. A protein involved in conferring insulin resis- tance is also a pro-inflammatory molecule that has important implications. Ó 2005 Elsevier Inc. All rights reserved. Keywords: Resistin; IL-12; TNF-a; NF-jB; Inflammation; Monocyte/macrophage; Insulin resistance; Obesity; Type 2 diabetes; Resistin confor- mation Resistin, an adipocytokine, is found to be elevated in genetic and diet-induced mouse models of obesity. This protein is expressed exclusively in adipocytes in rodents. However, in humans it is secreted mainly by macro- phages. Reduced insulin-stimulated glucose uptake in mice that were administered recombinant resistin and reversal of the same by anti-resistin IgG indicates a role for this molecule in the development of insulin resistance [1]. Plasma resistin levels are elevated in individuals with type 2 diabetes mellitus [2]. Diabetics with insulin resis- tance and reduced glucose uptake suffer from cytokine- induced acute-phase inflammation [3]. Inflammation in relation to obesity and insulin resis- tance has often been correlated with the over-production of the pro-inflammatory cytokine TNF-a [4]. TNF-a levels have also been found to be markedly increased in different murine models of obesity namely ob/ob mouse, db/db mouse, and fa/fa Zucker rats. Gene expression profiling by microarray in obese mice re- vealed that the largest class of genes altered significantly were those involved in inflammation, reminiscent of macrophages [5]. Although a positive correlation be- tween resistin and inflammatory disorders has been de- scribed [6], the mechanism(s) by which they are 0006-291X/$ - see front matter Ó 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2005.06.202 * Corresponding author. Fax: +91 40 27019074. E-mail address: nas_ehtesham@yahoo.com (N.Z. Ehtesham). 1 These two authors contributed equally. www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 334 (2005) 1092–1101 BBRC