Antifibrotic effects of ZK14, a novel nitric oxide-donating biphenyldicarboxylate derivative, on rat HSC-T6 cells and CCl4-induced hepatic fibrosis.
OBJECTIVE
To study the pharmacologic effect of ZK(14), a novel nitric oxide-donating biphenyldicarboxylate (DDB) derivative, on HSC-T6 cells and on CCl(4)-induced hepatic fibrosis.


METHODS
Inhibition of HSC-T6 cell growth by ZK(14) was evaluated by MTT assay. The effect of ZK(14) on the percentage of HSC-T6 cells undergoing apoptosis was measured using Annexin-V/PI double-staining and TUNEL assay. Mitochondrial membrane potential (MMP) and caspase activities were tested. Hepatic fibrosis was induced in Sprague-Dawley rats by intraperitoneal injection with 14% CCl(4). Rats with hepatic fibrosis were randomly divided into four groups: model control, ZK(14) (20 mg/kg), ZK(14) (10 mg/kg) and DDB (5 mg/kg). Levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), hyaluronic acid (HA), type III collagen (PCIII), and nitric oxide (NO) were assessed, and liver samples were stained with hematoxylin-eosin. The NO level in cells treated with ZK(14) in vitro was also measured.


RESULTS
The effect of ZK(14) on HSC-T6 cell apoptosis was concentration- and time-dependent, with up to 50% of cells becoming apoptotic when exposed to 100 mumol/L ZK(14) for 18 h. ZK(14) treatment resulted in mitochondrial membrane depolarization and activation of caspases 3 and 9. At a dose of 20 mg/kg, ZK(14) significantly decreased serum transaminase (AST, ALT) activities and fibrotic index (HA, PCIII) levels and significantly inhibited fibrogenesis.


CONCLUSIONS
These data indicate that ZK(14), a novel NO-donating DDB derivative, promotes HSC-T6 apoptosis in vitro through a signaling mechanism involving mitochondria and caspase activation and it inhibits CCl(4)-induced hepatic fibrosis in vivo. The results suggest that ZK(14) has potential therapeutic value in the treatment of hepatic fibrosis.
Dai
Li
L
Department of Pharmacology, China Pharmaceutical University, Nanjing, China.
Ji
Hui
H
Kong
Xiang-wen
XW
Zhang
Yi-hua
YH
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001713:Biphenyl Compounds / Q000008:administration & dosage / Q000494:pharmacology*; D002251:Carbon Tetrachloride; D053148:Caspase 3 / Q000187:drug effects / Q000378:metabolism; D053453:Caspase 9 / Q000187:drug effects / Q000378:metabolism; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D055166:Hepatic Stellate Cells / Q000187:drug effects / Q000378:metabolism; D020287:In Situ Nick-End Labeling; D008106:Liver Cirrhosis, Experimental / Q000188:drug therapy* / Q000503:physiopathology; D008111:Liver Function Tests; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D010069:Oxadiazoles / Q000008:administration & dosage / Q000494:pharmacology*; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
C547632:4,4'-dimethoxy-5,6,5',6'-dimethylenedioxy-2-methoxycarbonyl-2'-(N-(alanine 2-(3-phenylsulfonyl-1,2,5-oxadiazole-2-oxide-4-) oxyethoxyethyl ester)) carbamoyl biphenyl; D001713:Biphenyl Compounds; D020030:Nitric Oxide Donors; D010069:Oxadiazoles; D002251:Carbon Tetrachloride; D053148:Caspase 3; D053453:Caspase 9