The Studies Of The Protective Effect Of Hydrogen-rich Saline On Ethanol-induced Liver Injury In Vitro And In Vivo

Aim: To investigate the protective effects of hydrogen-rich saline on alcoholic liver injury disease in vitro and in vivo.Method: First of all,in vitro we build ethanol-induced liver cell injury model by using hepatocyte cell lines(LO2)to study whether hydrogen-rich culture medium(HCM)can protect against alcoholic liver cell damage.In vivo experiments,we use ICR mouse to construct two kinds of experimental mouse models in our present study,first we build acute alcoholic liver injury mouse model by intragastric administration with high-dose alcohol in a short time and then we evaluate the protective effects of hydrogen-rich saline(HS)on mice with acute alcoholic liver injury.In addition,in order to further demonstrate the protective effects of HS on ALD,we here use the same kind mouse to build chronic alcoholic liver injury mouse model,yet,we adopt a new method that we use chronic intragastric administration with alcohol of various concentrations which from low dose to high dose to construct chronic alcoholic liver injury mouse model so as to prevent happen of high mouse mortality.Specifically,we first use CCK-8 assay kit to measure cell proliferation activity for determining the optimal ethanol concentration which is utilized to build ethanol-induced liver cell damage model later.Second,we then test levels of cell viability and some biomarkers for comparing degree of liver cell injury in HCM group with other groups.In vivo experiments,we here use 50% ethanol to build acute and chronic alcoholic liver injury models,at the same time,mice in different groups are subjected to various treatments including normal saline,low-dose HS(5 ml/kg),high-dose HS(10 ml/kg)and Vitamin C solution(200 mg/kg),respectively.Then,assay kits such AST,ALT,SOD,GSH assay kits are used to measure levels of biomarkers in serum and in liver tissues that reflect liver functions,besides we make HE staining liver tissue slices to observe changes of hepatic structure in different groups.Results: In vitro experiments,CCK-8 results indicated that cell proliferation rates are significantly(p<0.05)increased in HCM treatment group when compared with other groups.In addition,HCM treatment can markedly reverse the level changes of some biomarkers in culture medium and in cells.In vivo experiments,we found that HS supplement can restrain the increase of biochemical indices levels or the decrease of their levels induced by alcohol whether in acute alcoholic liver injury models or in chronic alcoholic liver injury models.More importantly,the results of HE staining liver tissue slices also showed that HS treatment can alleviate liver injury resulted from alcohol.Conclusion: In our present study,we proved that hydrogen supplement can relief liver injury induced by ethanol in vitro and in vivo according to a series of detailed experiments designed by us.Accordingly,drinking hydrogen-rich water could be regarded as a kind of therapeutic methods in clinic to treat patients with alcoholic liver injury disease in the future.