| Objective: The incidence of non-alcoholic fatty liver disease(NAFLD) is increasing.The condition covers the spetrum from steatosis to steatohepatitis,fibrosis and cirrhosis.No pharmacological treatment is currently approved for the treatment of NAFLD.Glucagon-like peptide-1(GLP-1) secreted by intestinal L endocrine cells which belongs to one of an important incretin. GLP-1 analogues-liraglutide as antidiabetic drugs have been widely used clinically.Pancreatic outside effectiveness its role out of pancreatic has attracted wide attention in the field of diabetes. Resistance of organ fibrosis mediated by GLP-1 receptor has been preliminary confirmed. Its mechanism is very complicated, and in different organs there may be different functions, and some key cells,such as hepatic stellate cells,are involved rather possibly.As the key cells of the liver fibrosis, does GLP-1 receptor have expression in the HSC or not?This study was designed to find out the expression of GLP-1 receptor in rat hepatic stellate cells, the effects of liraglutide for rat hepatic stellate cells under high glucose conditions and the GLP-1 influence towards NAFLD and fibrosis.Methods:1 The cultivation of hepatic stellate cells : The rat hepatic stellate cells were vaccinated in germ-free plastic bottles at 37℃ and the volume fraction of 5% CO2 saturated humidity incubator with 1640 medium which including 10% fetal bovine serum. Changed medium once a day,extracted the sample to identify morphology under a microscope, and analysed for subsequent experients.2 Used Western blot method to detect the GLP-1 receptor protein expression in rat hepatic stellate cells.3 Used real-time PCR method to detect the GLP-1 receptor m RNA expression in rat hepatic stellate cells.4 Intervened and divided into groups: divided the rat hepatic stellate cells cultured for 48 hours into 7 groups,respectively, low glucose control group(5.5 mmol/L), hypertonic control group(5.5mmol/L glucose+19.5mmol/L mannitol), high glucose control group(25.0mmol/L), high glucose+low concentration of liraglutide group(50nmol/L), high glucose+medium concentration liraglutide group(100nmol/L), high glucose+high concentration of liraglutide group(200nmol/L), low glucose+high concentration of liraglutide group(200 nmol/L).5 Used MTS method to detect the proliferation of the cells from 7 groups as described above.6 Data analysis: The data were presented as mean±standard deviation( sx ?) and the statistical analysis ussd SPSS 13.0 software. Used analysis of variance for Optical density(OD) values of different groups at different times, the Bonferroni method were used to discuss concrete difference of different groups of the same point in time. For the high sugar + high concentrations of liraglutide group used paired t test to study the effects of time for OD values. P <0.05 was considered statistically significant.Results:1 The morphology identification of rat hepatic stellate cells.The activating hepatic stellate cells observed under inverted microscope were flat,with large cell body,and there are projections around the cell body.2 The rat hepatic stellate cells had the expression of GLP-1 receptor proved by the method of western blot and real-time PCR. GLP-1 protein and m RNA expression was assessed in five samples.3 The proliferation of hepatic stellate cells:The proliferation of high glucose control group compaired with low glucose control group increased(p<0.05), At the time of 48 hours and 72 hours high glucose with high concentrations of liraglutide(200 nmol/L) intervention group was lower than high glucose control group(p<0.05), but higher than low glucose control group(p<0.05). Moreover, the proliferation of high concentrations of liraglutide(200 nmol/L) was lower than the low(50 nmol/L), medium(100 nmol/L) concentrations of liraglutide group(p<0.05), but low concentrations of liraglutide group had no difference with medium concentrations of liraglutide group, hypertonic control group and low glucose with high liraglutide group had no difference with the low glucose control group(p>0.05).Conclusion:1 The rat hepatic stellate cells had the expression of GLP-1 receptor.2 High glucose could increase the proliferation of rat hepatic stellate cells,However liraglutide could fight the proliferation, and was correlated with its concentration and action time. |
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