Upregulation of heme oxygenase-1 by degranulation in rat basophilic leukemia cells.
Heme oxygenase (HO)-1, which is a rate-limiting enzyme involved in the catabolism of heme, is upregulated by a variety of stresses including oxidative stresses and inflammatory cytokines, in many cell types. Recent studies have suggested that upregulation of HO-1 might provide cytoprotection and immunomodulatory functions in addition to its obvious role in heme metabolism. In this study, we examined whether HO-1 was upregulated following degranulation in mast cells that initiate vigorous immunity reactions. To trigger degranulation, rat basophilic leukemia (RBL)-2H3 cells were passively sensitized using an antiserum collected from ovalbumin (OA) immunized-Brown Norway rats, and the cells were stimulated by treatment with OA. Degranulation was confirmed by measuring the release of beta-hexosaminidase. HO-1 mRNA and presence of HO-1 protein were detected using Northern blot and Western blot analyses, respectively. The effect of the antioxidant N-acetyl-L-cysteine (NAC) on HO-1 expression was also tested. HO-1 mRNA transiently increased at 1--2 h after RBL-2H3 cells were stimulated to degranulate. Its mRNA increases were dependent on the extent of degranulation. Following the upregulation of HO-1 mRNA, HO-1 protein was also increased. We also detected intracellular production of reactive oxygen species following degranulation in RBL-2H3 cells. NAC attenuated the HO-1 expression in a dose-dependent manner. This is the first report to reveal induction of both HO-1 mRNA and protein by degranulation in RBL-2H3 cells. We showed that NAC inhibited HO-1 upregulation. These results suggest that oxidative stress in activated RBL-2H3 cells results in the upregulation of HO-1.
Yasui
Yumiko
Y
Department of Pharmacology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University. yumikoy@ph.bunri-u.ac.jp
Sasao
Emiko
E
Sakata
Masaru
M
Matsui
Nobuaki
N
Fukuishi
Nobuyuki
N
Akagi
Reiko
R
Akagi
Masaaki
M
D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D001391:Azo Compounds / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D015550:Cell Degranulation; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D005453:Fluorescence; D016166:Free Radical Scavengers / Q000494:pharmacology; D051547:Heme Oxygenase-1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015471:Leukemia, Basophilic, Acute / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D008665:Metalloporphyrins / Q000494:pharmacology; D010047:Ovalbumin / Q000276:immunology / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011914:Rats, Inbred BN; D017382:Reactive Oxygen Species / Q000378:metabolism; D015854:Up-Regulation
C110400:2',7'-dichlorodihydrofluorescein diacetate; D001391:Azo Compounds; D004791:Enzyme Inhibitors; D005452:Fluoresceins; D016166:Free Radical Scavengers; D008665:Metalloporphyrins; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; C055421:tin mesoporphyrin; D010047:Ovalbumin; D051547:Heme Oxygenase-1; C032756:ponceau S; D000111:Acetylcysteine