title: Anti-inflammatory effects of miR-21 in the macrophage response to peritonitis.
abstract: We investigated the role of microRNA-21 in the macrophage response to peritonitis; microRNA-21 expression increases in peritoneal macrophages after lipopolysaccharide stimulation but is delayed until 48 hours after cecal ligation and puncture. MicroRNA-21-null mice and bone marrow-derived cell lines were exposed to cecal ligation and puncture or lipopolysaccharide, and survival, microRNA-21 levels, target messenger RNAs and proteins, and cytokines were assayed. Macrophages were also transfected with microRNA-21 mimics and antagomirs, and similar endpoints were measured. Survival in microRNA-21-null mice was significantly decreased after lipopolysaccharide-induced peritonitis but unchanged after cecal ligation and puncture compared with similarly treated wild-type mice. MicroRNA-21 expression, tumor necrosis factor-α, interleukin 6, and programmed cell death protein 4 levels were increased after lipopolysaccharide addition in peritoneal cells. Pelino1 and sprouty (SPRY) messenger RNAs were similarly increased early, whereas programmed cell death protein 4 messenger RNA was decreased after lipopolysaccharide, and all microR-21 target messenger RNAs were subsequently decreased by 24 hours after lipopolysaccharide. Transfection with mimics and antagomirs led to appropriate responses in microRNA-21 and tumor necrosis factor-α. Knockdown of microRNA-21 in bone marrow-derived cells showed increased tumor necrosis factor-α and decreased interleukin 10 in response to lipopolysaccharide. Target proteins were unaffected by knockdown as was extracellular signal-regulated kinase; however, the nuclear factor κB p65 subunit was increased after lipopolysaccharide in the microRNA-21 knockout cells. In contrast, there was little change in these parameters after cecal ligation and puncture induction between null and wild-type mice. MicroRNA-21 is beneficial to survival in mice following lipopolysaccharide peritonitis. Overexpression of microRNA-21 decreased tumor necrosis factor-α secretion, whereas suppression of microRNA-21 expression increased tumor necrosis factor-α and interleukin 6, and decreased interleukin 10 levels after lipopolysaccharide. Protein targets of microRNA-21 were not different following suppression of microRNA-21. Nuclear factor κB was increased by suppression of microRNA-21. These findings demonstrate microRNA-21 is beneficial in modulating the macrophage response to lipopolysaccharide peritonitis and an improved understanding of the anti-inflammatory effects of microRNA-21 may result in novel, targeted therapy against peritonitis and sepsis.
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Rebecca Elise
authors.initials: RE
authors.lastname: Barnett
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Conklin
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Lindsey
authors.initials: L
authors.lastname: Ryan
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Keskey
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Vikram
authors.initials: V
authors.lastname: Ramjee
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Ernesto A
authors.initials: EA
authors.lastname: Sepulveda
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Sanjay
authors.initials: S
authors.lastname: Srivastava
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA.
authors.forename: Aruni
authors.initials: A
authors.lastname: Bhatnagar
authors.affiliation: *Hiram C. Polk, Jr., MD, Department of Surgery, School of Medicine, and Division of Cardiovascular Medicine, Department of Medicine, University of Louisville, Louisville, Kentucky, USA; and Robley Rex Veterans Affairs Medical Center, Louisville, Kentucky, USA wg.cheadle@louisville.edu.
authors.forename: William G
authors.initials: WG
authors.lastname: Cheadle
mesh_terms: D000818:Animals; D002432:Cecum / Q000293:injuries; D002461:Cell Line, Transformed; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D016753:Interleukin-10 / Q000096:biosynthesis / Q000235:genetics; D015850:Interleukin-6 / Q000096:biosynthesis / Q000235:genetics; D007416:Intestinal Perforation / Q000150:complications; D008070:Lipopolysaccharides / Q000633:toxicity; D008264:Macrophages / Q000502:physiology*; D017737:Macrophages, Peritoneal / Q000502:physiology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D058990:Molecular Targeted Therapy; D016328:NF-kappa B / Q000378:metabolism; D009841:Oligonucleotides / Q000494:pharmacology; D010538:Peritonitis / Q000139:chemically induced / Q000209:etiology / Q000276:immunology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics
chemical_list: C556062:IL10 protein, mouse; D015850:Interleukin-6; D008070:Lipopolysaccharides; C531950:MIRN21 microRNA, mouse; D035683:MicroRNAs; D016328:NF-kappa B; D009841:Oligonucleotides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; C577686:interleukin-6, mouse; C482199:lipopolysaccharide, E coli O55-B5; D016753:Interleukin-10
keywords: Toll-like receptor-4; endotoxin; inflammation; septic shock