RT Journal Article
SR Electronic
T1 Effectiveness of Novel Imidazole-Dioxolane Heme Oxygenase Inhibitors in Renal Proximal Tubule Epithelial Cells
JF Journal of Pharmacology and Experimental Therapeutics
JO J Pharmacol Exp Ther
FD American Society for Pharmacology and Experimental Therapeutics
SP 763
OP 770
DO 10.1124/jpet.107.119800
VO 323
IS 3
A1 Robert T. Kinobe
A1 Yanbin Ji
A1 Jason Z. Vlahakis
A1 Roberto Motterlini
A1 James F. Brien
A1 Walter A. Szarek
A1 Kanji Nakatsu
YR 2007
UL http://jpet.aspetjournals.org/content/323/3/763.abstract
AB To enhance our understanding of the physiological roles of heme oxygenase (HO) isozymes, HO-1 (inducible) and HO-2 (constitutive), we developed novel imidazole-based HO inhibitors. Unlike the metalloporphyrins, these imidazole-dioxolane compounds are selective for the in vitro inhibition of HO with minimal effects on other heme-dependent enzymes such as nitric oxide synthase and soluble guanylyl cyclase. In the current study, we tested the hypothesis that these novel HO inhibitors are effective in intact cells by extending their application to cultured, renal proximal tubule epithelial cells (LLC-PK1). HO-1 and HO-2 protein expression was enhanced by pretreatment of cells with hemin, transduction with adenovirus encoding human HO-1, and transfection with cDNA for HO-2, respectively. Total HO activity was measured by determining the formation of carbon monoxide (CO), whereas cell viability and apoptosis were measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and the expression of activated caspase-3. Gliotoxin/tumor necrosis factor-α (TNF-α) produced cytotoxicity in wild-type LLC-PK1 cells (P < 0.05) but not in HO-1 and HO-2 overexpressing or wild type cells pretreated with hemin (10 μM). The presence of imidazole-dioxolane HO inhibitors (2–25 μM) decreased cell viability (P < 0.05). A CO-releasing molecule reversed, in a dose-dependent manner, the cytotoxic effects and caspase-3 activation induced by the combination of gliotoxin/TNF-α and the HO inhibitors, suggesting an important role for CO in protection against renal toxicity. These data demonstrate a protective role of both HO-1 and HO-2 against gliotoxin/TNF-α-induced cytotoxicity in LLC-PK1 cells. The novel imidazole-dioxolane compounds can be used as effective inhibitors of HO activity in cell culture. The American Society for Pharmacology and Experimental Therapeutics