Cobalt protoporphyrin inhibition of lipopolysaccharide or lipoteichoic acid-induced nitric oxide production via blocking c-Jun N-terminal kinase activation and nitric oxide enzyme activity |
| |
Authors: | Hui-Yi Lin Shing-Chuan Shen Cheng-Wei Lin Yen-Chou Chen |
| |
Institution: | a Graduate Institute of Medical Sciences, Taipei Medical University, Taipei 110, Taiwan b Graduate Institute of Pharmacy, School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan c Department of Gastroenterology, Taipei Medical University Wan Fan Hospital, Taipei 110, Taiwan d Cancer Research Center and Orthopedics Research Center, Taipei Medical University Hospital, Taipei 110, Taiwan |
| |
Abstract: | In the present study, low doses (0.5, 1, and 2 μM) of cobalt protoporphyrin (CoPP), but not ferric protoporphyrin (FePP) or tin protoporphyrin (SnPP), significantly inhibited lipopolysaccharide (LPS) or lipoteichoic acid (LTA)-induced inducible nitric oxide (iNOS) and nitric oxide (NO) production with an increase in heme oxygenase 1 (HO-1) protein in RAW264.7 macrophages under serum-free conditions. IC50 values of CoPP inhibition of NO and iNOS protein individually induced by LPS and LTA were around 0.25 and 1.7 μM, respectively. This suggests that CoPP is more sensitive at inhibiting NO production than iNOS protein in response to separate LPS and LTA stimulation. NO inhibition and HO-1 induction by CoPP were blocked by the separate addition of fetal bovine serum (FBS) and bovine serum albumin (BSA). Decreasing iNOS/NO production and increasing HO-1 protein by CoPP were observed with CoPP pretreatment, CoPP co-treatment, and CoPP post-treatment with LPS and LTA stimulation. LPS- and LTA-induced NOS/NO productions were significantly suppressed by the JNK inhibitor, SP600125, but not by the ERK inhibitor, PD98059, through a reduction in JNK protein phosphorylation. Transfection of a dominant negative JNK plasmid inhibited LPS- and LTA-induced iNOS/NO production and JNK protein phosphorylation, suggesting that JNK activation is involved in LPS- and LTA-induced iNOS/NO production. Additionally, CoPP inhibition of LPS- and LTA-induced JNK, but not ERK, protein phosphorylation was identified in RAW264.7 cells. Furthermore, CoPP significantly reduced NO production in a cell-mediated, but not cell-free, iNOS enzyme activity assay accompanied by HO-1 induction. However, attenuation of HO-1 protein stimulated by CoPP via transfection of HO-1 siRNA did not affect NO's inhibition of CoPP against LPS stimulation. CoPP effectively suppressing LPS- and LTA-induced iNOS/NO production through blocking JNK activation and iNOS enzyme activity via a HO-1 independent manner is first demonstrated herein. |
| |
Keywords: | HO-1 heme oxygenase-1 NO nitric oxide LPS lipopolysaccharide iNOS inducible nitric oxide synthase CO carbon monoxide MTT 3-(4 5-dimethylthiazol-2-yl)-2 5-diphenyl tetrazolium bromide DTT dithiothreitol L-NAME N-nitro-L-arginine methyl ester NBT nitroblue tetrazolium BCIP 5-bromo-4-chloro-3-indolyl phosphate COX cyclooxygenase CoPP cobalt protoporphyrin SnPP tin protoporphyrin |
本文献已被 ScienceDirect 等数据库收录! |
|