Therapeutic And Prevention Effects Of Caffeine On Alcohol-induced Liver Fibrosis In Rats And Its Partial Mechanisms

Alcoholic liver disease(ALD) is the result of long-term heavy dringking,the mainly symptoms are varying degrees of liver cells damages.Basing on the the different progresses of disease,there are alcoholic fatty liver,alcoholic hepatitis,alcoholic hepatic fibrosis and alcoholic cirrhosis,eventually,the liver cells are widespread putrescenced and the function of the liver lost. Alcohol-induced liver fibrosis is characterized as extracellular matrix(ECM) hyperplasia stimulated by alcohol,which is the central link of the process of ALD,effective treatment of alcohol-induced liver fibrosis will block or delay the process of chronic ALD.However,at present there are no effective medicine that can prevent alcohol-induced liver fibrosis.Caffeine is a the main ingredient of coffee and tea,which are the usual drink in life,serving drinks containing caffeine is widely spread in the world.Rencent years,more and more studies reported that having drink that containing caffeine will reduce the probability of suffering from the liver disease,but the mechanism of it has not yet been elaborated completely.So whether the intaking of caffeine can also affect alcohol-induced liver fibrosis which is the central link of the process of ALD and what's the possible mechanism? Basing on a review of relevant literatures,in this research,at first we established the alcohol-induced hepatic fibrosis model in rats by increasing concentration of alcohol twice a day for 12 weeks,on this basis, we stimulated the HSC by acetaldehyde to establish the alcohol-induced hepatic fibrosis model in vitro,then explored the therapeutic and prevention effects of low concentration of caffeine on alcohol-induced hepatic fibrosis and the effects on proliferation of HSC-T6 stimulated by acetaldehyde and its partial mechanisms.The main contents summarized as follows:1. The therapeutic and prevention effects of caffeine on alcohol-induced liver fibrosis The model of alcohol-induced liver fibrosis was duplicated by filling the stomach with increasing concentration of alcohol two times a day for 12 weeks,at the same time,the treatment groups were fed with corresponding concentration of caffeine(5,10,20mg/kg). At the end of 8 weeks and 12 weeks,compared with that of model group, caffeine groups can significantly reduce the serum alanine aminotransferase(ALT),aspartate aminotransferase(AST ),hyaluronic acid(HA ),laminin(LN),Procollagen III ( PIIINP ),Procollagen IV ( CIV ) contents ( P<0.01 ) ..Liver histopathological examination and sirius-red staining showed that in the end of 8 weeks,the liver tissue of model group have inflammatory cells infiltrationing and different sizes of fat drops,part of the hepatic lobule have necrosis,the red collagen fibers surrounded the edge of the portal areas,in the end of 12 weeks,large amount of inflammatory cells infiltrationing the liver tissue of model group , in the portal areas,the red collagen fibers extended to the hepatic lobule,the high dose of caffeine group had a lesser degree of inflammatory cells infiltrationing,the connective tissue proliferation reduced,the fibrous septa tended narrow and the structure tended to be the normal tissue. Immunohistochemical detections showed thatα-SMA in the caffeine groups tend to be reduced and the expressions of DR4(death receptor 4),DR5(death receptor 5)increased.These results suggest that caffeine has the therapeutic and prevention effects on alcohol-induced hepatic fibrosis and the mechanim may be the upregulated expressions of DR4,DR5 and the formation ofα-SMA reduced by caffeine ,thus alleviating the alcohol-induced hepatic fibrosis. 2 The effects of caffeine on the proliferation and apoptosis of HSC-T6 stimulated by acetaldehyde and its partial mechanisms.The HSC-T6 pretreatmented with different doses of caffeine were stimulated by 200μmol·L-1 acetaldehyde.In the time of 24,48,72 hours,we use MTT to explore the proliferation of HSC-T6.The result showed that caffeine can inhibite the proliferation of HSC-T6 stimulated by acetaldehyde and the 50% inhibiting concentration (IC50 )were 21.404,3.332,1.784 mmol·L^-1 respectively.The apoptotic rates and the changes of cell cycle were detected by flow cytometer(FCM) when HSC cells stimulated by acetaldehyde were treated by caffeine after 48h ,compared with the model group,the apoptotic rates increase remarkably, the cells increased in G₀/G₁ and G₂/M periods and decreased in the S period.Isolating the toal RNAs of HSC-T6 stimulated by acetaldehyde,the result of RT-PCR showed that the mRNA expression levels ofα-Smooth mucle actin(α-SMA),Procollagen I(PI),Procollagen III(PIIINP) decreased,and the mRNA expression levels of DR4 and DR5 tend to be increased.These results suggest that the proliferation of HSC-T6 stimulated by acetaldehyde can be inhibited by caffeine and the mechanism may be related to the upregulating the mRNAs expressions of DR4 and DR5 ,then induce the apoptosis of HSC-T6.The research provided a promising approach to using caffeine as an anti-alcoholic liver fibrosis drug and provided a new idea for evaluation,screening effective treatment of alcoholic liver fibrosis.