The Meaning Of CMKLR1 Over Expression In Liver Tissue Of SD Rat With Nonalcoholic Fatty Hepatitis

Objective:To take SD male rats as the research object,feed them with high-fat diet cause nonalcoholic fatty liver disease,and make CMKLR1 over expression in liver tissue via slow virus,we look forward to that this basic research may provide direction for clinical treatment of NAFLD.Methods:Prepare lentivirus vectors which carry CMKLR1 gene of rat and identificate it in advance.42 male SD rats adaptive feeding 1 week in the appropriate illumination,temperature and humidity environment,then divide them into 4 groups at random: The control group,model group and transfection group(SV-CMKLR1)12 each,empty transfection group(CSV)6,control group feed with normal diet,others with high-fat diet.On the first day of experiment,inject PBS 100 ul to control group and model group via tail vein;SV-CMKLR1 group in tail vein injection with SV-EGFP 2×109pfu100ul;CSV group in tail vein injection with CSV 2×109pfu 100 ul.In the experimental7,14 and 21 day,sacrifice two rats in CSV group to evaluate the transfection efficiency.Tracking rats normally,record weight of rats each week in the process of experiment.In the experimental 8 and 12 week,Take 6 rats in control group,model group and transfection group each at random,After 12 h forbidden diet,anesthesia abdominal cavity With chloral hydrate,Then quickly cut open it,observe the liver morphology and take blood from abdominal aorta,Let liver tissue frozen in liquid nitrogen and then move it to 80 ℃ freezer until total RNA extraction.At the same time take part of the liver tissue with 10% formalin fixed,then do HE staining after paraffin section,observe liverhistology change and evaluate NAFLD activity score of each rat through microscope,Applicatting Real-time PCR detective the expression level of Adiponectin and CMKLR1 mRNA in each rat liver tissue.Applicating Protein immunoblot method to detective the expression,level of Adiponectin and CMKLR1 protein in each rat liver tissue.Results:(1)In comparison the Weight and liver index of rats at the same period: model group> control group(P ￡ 0.05),model group > transfection group(P ￡ 0.05).(2)Liver HE staining results showed that: The control group rats Liver cells arranged rules,have no inflammatory cell infiltration or cavity;In Model group rats 8 weeks,Liver tissue can appear liver cell swelling,liver cords become Disordered arrangement,we can see balloon sample change,characterized by simple fatty change;12 weeks,almost all of the liver cell swelling,cytoplasm is loose,a large number of spherical lipid drops appeared and a large number of inflammatory cells infiltration,we can see point necrosis and bridging necrosis,progress to NASH;Transfection group is also characterized by simple fatty liver on 8week and progress to the NASH on 12 week,But the degree is mild than model group in same period.(3)In comparison the Kay Murray level in the serum of rats In the same period:model group > control group(P>0.05),transfection group >model group(P ￡ 0.05).(4)In comparison the Adiponectin level in the serum of rats at same period:model group<control group(P ￡ 0.05),transfection group >model group(P ￡ 0.05).(5)In comparison the rats at same period,PCR detection CMKLR1 mRNA and adiponectin mRNA expression in liver tissue,the results showed that,The expression of CMKLR1 mRNA lever: model group > control group(P>0.05),transfection group >model group(P ￡ 0.05);The expression of adiponectin mRNA lever: control group >model group(P ￡ 0.05),transfection group > model group(P ￡ 0.05).(6)Protein immunoblot results at the same period showed that,The CMKLR1 protein content:model group > control group(P>0.05),transfection group > model group(P ￡ 0.05);Theexpression of adiponectin protein content : control group >model group(P ￡ 0.05),transfection group >model group(P￡ 0.05).Conclusion:Slow virus mediated CMKLR1 over expression in liver tissue of SD rats with nonalcoholic fatty hepatitis can improve the liver tissue pathology significantly.