Proanthocyanidin from blueberry leaves suppresses expression of subgenomic hepatitis C virus RNA.
Hepatitis C virus (HCV) infection is a major cause of chronic liver disease such as chronic hepatitis, cirrhosis, and hepatocellular carcinoma. While searching for new natural anti-HCV agents in agricultural products, we found a potent inhibitor of HCV RNA expression in extracts of blueberry leaves when examined in an HCV subgenomic replicon cell culture system. This activity was observed in a methanol extract fraction of blueberry leaves and was purified by repeated fractionations in reversed-phase high-performance liquid chromatography. The final purified fraction showed a 63-fold increase in specific activity compared with the initial methanol extracts and was composed only of carbon, hydrogen, and oxygen. Liquid chromatography/mass-ion trap-time of flight analysis and butanol-HCl hydrolysis analysis of the purified fraction revealed that the blueberry leaf-derived inhibitor was proanthocyanidin. Furthermore, structural analysis using acid thiolysis indicated that the mean degree of polymerization of the purified proanthocyanidin was 7.7, consisting predominantly of epicatechin. Proanthocyanidin with a polymerization degree of 8 to 9 showed the greatest potency at inhibiting the expression of subgenomic HCV RNA. Purified proanthocyanidin showed dose-dependent inhibition of expression of the neomycin-resistant gene and the NS-3 protein gene in the HCV subgenome in replicon cells. While characterizing the mechanism by which proanthocyanidin inhibited HCV subgenome expression, we found that heterogeneous nuclear ribonucleoprotein A2/B1 showed affinity to blueberry leaf-derived proanthocyanidin and was indispensable for HCV subgenome expression in replicon cells. These data suggest that proanthocyanidin isolated from blueberry leaves may have potential usefulness as an anti-HCV compound by inhibiting viral replication.
Takeshita
Masahiko
M
Research Division, Minami Nippon Dairy Co-op Co., Ltd., Miyazaki 885-0073, Japan.
Ishida
Yo-Ichi
Y
Akamatsu
Ena
E
Ohmori
Yusuke
Y
Sudoh
Masayuki
M
Uto
Hirofumi
H
Tsubouchi
Hirohito
H
Kataoka
Hiroaki
H
D029798:Blueberry Plants; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid / Q000379:methods; D024882:Drug Resistance, Viral; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D016174:Hepacivirus / Q000235:genetics*; D020128:Inhibitory Concentration 50; D013058:Mass Spectrometry / Q000379:methods; D009355:Neomycin / Q000494:pharmacology; D010936:Plant Extracts / Q000378:metabolism*; D018515:Plant Leaves / Q000378:metabolism*; D011108:Polymers / Q000737:chemistry; D044945:Proanthocyanidins / Q000378:metabolism*; D012367:RNA, Viral; D014779:Virus Replication / Q000187:drug effects*
D010936:Plant Extracts; D011108:Polymers; D044945:Proanthocyanidins; D012367:RNA, Viral; C013221:proanthocyanidin; D009355:Neomycin