Suppression of Tumor Necrosis Factor  Production by cAMP in Human Monocytes: Dissociation with mRNA Level and Independent of Interleukin-10 Brian D. Shames, M.D., Robert C. McIntyre, Jr., M.D., Denis D. Bensard, M.D.,* Edward J. Pulido, M.D., Craig H. Selzman, M.D., Leonid L. Reznikov, Ph.D., Alden H. Harken, M.D., and Xianzhong Meng, M.D., Ph.D. 1 Department of Surgery, University of Colorado Health Sciences Center, Denver, Colorado 80262; and *Department of Pediatric Surgery, The Children’s Hospital, Denver, Colorado 80262 Submitted for publication January 29, 2001; published online May 24, 2001 Background. Elevation of cellular cAMP inhibits lipopolysaccharide (LPS)-stimulated tumor necrosis factor  (TNF-) production and increases the expres- sion of interleukin (IL)-10 in mononuclear cells. TNF- gene expression obligates activation of the transcrip- tion factor nuclear factor B (NF-B). Exogenous IL-10 inhibits NF-B in monocytes and thus attenuates TNF- production. We examined the role of endoge- nous IL-10 in the regulation of NF-B activation and TNF- production in human monocytes by cAMP. Methods. Human monocytes were stimulated with Escherichia coli LPS (100 ng/ml) with and without for- skolin (FSK, 50 M) or dibutyryl cyclic AMP (dbcAMP, 100 M). Cytokine (TNF- and IL-10) release was mea- sured by immunoassay. TNF- mRNA was measured by reverse transcription polymerase chain reaction, and NF-B DNA binding activity was assessed by gel mobility shift assay. Results. cAMP-elevating agents inhibited LPS- stimulated TNF- release (0.77  0.13 ng/10 6 cells in LPS  dbcAMP and 0.68  0.19 ng/10 6 cells in LPS  FSK, both P < 0.05 vs 1.61  0.34 ng/10 6 cells in LPS alone). Conversely, cAMP enhanced LPS-stimulated IL-10 release (100  21.5 pg/10 6 cells in LPS  dbcAMP and 110  25.2 pg/10 6 cells in LPS  FSK, both P < 0.05 vs 53.3  12.8 pg/10 6 cells in LPS alone). Neither TNF- mRNA expression nor NF-B activation stimulated by LPS was inhibited by the cAMP-elevating agents. Neu- tralization of IL-10 with a specific antibody did not attenuate the effect of cAMP-elevating agents on TNF- production. Conclusion. The results indicate that cAMP inhibits LPS-stimulated TNF- production through a posttran- scriptional mechanism that is independent of endoge- nous IL-10. © 2001 Academic Press Key Words: nuclear factor B; gene expression; lipo- polysaccharide; forskolin; dibutyryl cAMP. INTRODUCTION Tumor necrosis factor  (TNF-) orchestrates in- flammation in a manner that can provoke both acute and chronic disease. Evidence for this central role of TNF- is derived from successful treatment of rheuma- toid arthritis [1] and inflammatory bowel disease [2] with anti-TNF- therapies. Furthermore, the hemody- namic changes associated with the sepsis syndrome are mediated by the production of TNF- and interleukin-1 (IL-1) [3]. Regulatory strategies inhibit- ing TNF- production are of immediate clinical appli- cation. Sepsis also induces anti-inflammatory factors, in- cluding catecholamines and interleukin-10 (IL-10) [4, 5]. Catecholamines inhibit TNF- production through  2 -adrenoreceptor-mediated elevation of intracellular cyclic adenosine monophosphate (cAMP) and activa- tion of protein kinase A (PKA) [4, 6, 7]. Agents that increase intracellular cAMP, including exogenous dibutyryl cAMP (dbcAMP, a cell-permeable cAMP an- alog), adenylyl cyclase activator, and phosphodiester- ase inhibitors, decrease TNF- production by mononu- clear cells [8, 9]. IL-10 was initially identified as a “cytokine synthesis 1 To whom correspondence should be addressed at Department of Surgery, University of Colorado Health Sciences Center, Box C-320, 4200 East Ninth Avenue, Denver, CO 80262. Fax: (303) 315-0007. E-mail: xianzhong.meng@uchsc.edu. Journal of Surgical Research 99, 187–193 (2001) doi:10.1006/jsre.2001.6178, available online at http://www.idealibrary.com on 187 0022-4804/01 $35.00 Copyright © 2001 by Academic Press All rights of reproduction in any form reserved.