Protective effects of n-Butanol extract and iridoid glycosides of Veronica ciliata Fisch. against ANIT-induced cholestatic liver injury in mice

Ethnopharmacological relevance: Veronica ciliata Fisch. is a traditional medical herb that present in more than 100 types of Tibetan medicine prescriptions, most of which are used for liver disease therapy. Iridoid glycosideshave been identified as the major active components of V.ciliata with a variety of biological activities.

Aims of the study: The aim of this study is to explore the protective effect and potential mechanism of n-Butanol extract (BE) and iridoid glycosides (IG) from V.ciliata against ɑ-naphthyl isothiocyanate (ANIT)-induced hepatotoxicity and cholestasis in mice.

Materials and methods: Mice were intragastrically (i.g.) given BE and IG at different dose or positive control ursodeoxycholic acid (UCDA) once a day for 14 consecutive days, andwere treated with ANIT to cause liver injury on day 12^th.Serum levels of hepatic injury markers and cholestasis indicators, liver index and liver histopathology were measured to evaluate the effect of BE and IG on liver injury caused by ANIT. Theprotein levels of tumor necrosis factor-α (TNF-α), nuclear factor kappa B(NF-κB), interleukin-6 (IL-6), Na+/taurocholate cotransporting polypeptide (NTCP), bile salt export pump (BSEP), multidrug resistance-associated protein 2 (MRP2), and the levels of oxidative stress indicators in liver tissuewere investigatedto reveal the underlying protective mechanisms of BE and IG against ANIT-induced hepatotoxicity and cholestasis.

Results: The n-Butanol extract (BE) and iridoid glycosides (IG) isolated fromV.ciliata significantly decreased serum level of cholestatic liver injury markers aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), γ-Glutamyl transferase (GGT), total bile acid (TBA), total bilirubin (TBIL), and direct bilirubin (DBIL) in ANIT-treated mice. Histopathology of the liver tissue showed that pathological damages were relieved upon BE and IG treatment. Meanwhile, the results indicated BE and IG notably restored relative liver weights, inhibited oxidative stress induced by ANIT through increasing hepatic level of superoxide dismutase (SOD), reduced glutathione (GSH), catalase (CAT) and decreasing hepatic content of malondialdehyde (MDA). Western blot revealed that BE and IG inhibited the expression of pro-inflammatory factors TGF-α, IL-6 and NF-κB. Furthermore, the decreased protein expression of bile acid transporters NTCP, BSEP, MRP2 were upregulated by BE and IG in a dose-dependent manner.

Conclusion: The results have demonstrated that BE and IG exhibited a dose-dependently protective effect against ANIT-induced liver injury with acute intrahepatic cholestasis in mice, which might be related to the regulation of oxidative stress, inflammatory response and bile acid transporters. In addition, these findings pointed out that iridoid glycosides as main active components of V.ciliata play a critical role in hepatoprotective effect of V.ciliata.

Keywords: ANIT; Bile acid transporter; Cholestasis; Iridoid glycoside; Veronica ciliata Fisch..