title: Free heme is a danger signal inducing expression of proinflammatory proteins in cultured cells derived from normal rat hearts.
abstract: Endogenous molecules from damaged tissue act as danger signals to trigger or amplify the immune/inflammatory response. In this study, we examined whether free heme induced pro-inflammatory proteins in cultured cells derived from normal hearts and investigated the cells targeted by heme, together with its mechanism of action in these cells. We cultured collagenase-isolated heart-derived cells from normal rats and examined whether free heme induced pro-inflammatory proteins, reactive oxygen species (ROS) production and NF-κB activation, by quantitative RT-PCR, ELISA and flow cytometry. Free heme increased mRNA of various pro-inflammatory proteins in cultured cardiac resident cells (CCRC) (at least 100-fold) and induced intracellular ROS formation. Approximately 85-90% of CCRC are fibroblast/smooth muscle cells and 10-15% are CD11bc-positive macrophages; therefore to examine individual target cells, macrophage-deleted (CD11bc-negative) CCRC, primary cultured cells (cardiac fibroblasts, arterial smooth muscle cells and cardiac microvascular endothelial cells) and macrophage cells lines (NR8383) were similarly treated. Free heme activated NF-κB and induced expression of some pro-inflammatory proteins, including IL-1 and TNF-α in NR8383. On the other hand, macrophage-deleted CCRC strongly increased expression of these proteins on treatment with IL-1 or TNF-α, but not free heme. Induction of expression of pro-inflammatory proteins by free heme was not inhibited by intracellular ROS reduction, but by protease and proteasome inhibitors capable of regulating NF-κB. These data suggest that free heme strongly induces various pro-inflammatory proteins in injured hearts through NF-κB activation in cardiac resident macrophages and through cross-talk between macrophages and fibroblast/smooth muscle cells mediated inter alia by IL-1, TNF-α.
authors.affiliation: Division of Cardiology, Niigata University Graduate School of Medical and Dental Sciences, 1-757 Asahimachi-dori, Niigata 951-8120, Japan.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Hao
authors.forename: Haruo
authors.initials: H
authors.lastname: Hanawa
authors.forename: Limin
authors.initials: L
authors.lastname: Ding
authors.forename: Yoshimi
authors.initials: Y
authors.lastname: Ota
authors.forename: Kaori
authors.initials: K
authors.lastname: Yoshida
authors.forename: Ken
authors.initials: K
authors.lastname: Toba
authors.forename: Minako
authors.initials: M
authors.lastname: Ogura
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ito
authors.forename: Makoto
authors.initials: M
authors.lastname: Kodama
authors.forename: Yoshifusa
authors.initials: Y
authors.lastname: Aizawa
mesh_terms: D000818:Animals; D018845:CD11 Antigens / Q000378:metabolism; D048430:Cell Shape / Q000187:drug effects; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects; D006418:Heme / Q000494:pharmacology*; D006427:Hemin / Q000494:pharmacology; D018836:Inflammation Mediators / Q000378:metabolism*; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism*; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008297:Male; D009206:Myocardium / Q000166:cytology* / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011917:Rats, Inbred Lew; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*
chemical_list: D018845:CD11 Antigens; D018836:Inflammation Mediators; D053583:Interleukin-1beta; D016328:NF-kappa B; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D006418:Heme; D006427:Hemin