| Objective : To establish a rat model of chronic toxic liver injury induced by dimethylformamide(DMF)and observe its pathological features and the expression of mitochondrial apoptotic pathway-related proteins in the hepatic tissue of rats chronically toxicated by DMF.To investigate the role of apoptosis in the pathogenesis of DMF.Methods: 1.Establish a rat model of chronic toxic liver injury induced by DMF: SD rats were randomly divided into two groups with eight rats in each group;the DMF liver injury group: 40% DMF solution was injected intraperitoneally at the dosage of 1 ml/100 g once a week;the normal control group underwent intraperitoneal injection of an equal volume of 0.9% sodium chloride once a week;blood was drawn 2 days at week 8 of administration and liver biochemistry(ALT,AST,ALB)was tested.The tissues from the sacrificed rats were examined pathologically and tested for apoptosis-related proteins.2.Liver biochemistry was performed using an automatic biochemical analyzer.3.Histopathological examination of the liver: Liver tissues were fixed in 10% neutral formaldehyde solution,routinely embedded with paraffin,stained with HE and reticular fibers,and examined under a microscope.4.Apoptosis detection: Mitochondrial apoptosis pathway-related proteins including Bax,Bcl-2,Cyt-c,Caspase-3,Caspase-9 were immunohistochemically stained and the results were analyzed using the method of semi-quantitative integration and Image J software.Results: 1.Rats' general condition: Compared with the normal control group,the rats in the DMF group lost weight Significant differences were found in body weight between the DMF groupand the control group at week 8(450.2±12.6g vs 312.8±25.9g,t=12.51,p? 0.001).2.Liver biochemistry: At week 8,both groups had similar ALT levels(87.83±28.63 vs 82.00±19.9,t=1.38,p=2.61)and AST levels(126.12±44.25 vs 122.2±26.88,t=0.35,p=0.77.Compared with the normal control group,ALB significantly decreaseed in the DMF group(28.03±1.56 vs 27.49±0.71,t=0.89,p=0.03).3.Liver histopathological lesions: The lobular structure of liver tissue in the DMF group was still intact.Mild hepatocyte edema and vacuolar degeneration were observed.Cellular eosinophilic lesions and apoptotic bodies were found around the central vein.The bile ducts were slightly dilated with cholestasis.Fibrosis was observed peri-sinus using fibrosis staining.The liver tissue was normal in the control group.4.Cell apoptosis: Bax was highly expressed in the DMF group,prominentlyin hepatocytes around the central vein.It was negative in the control group.The semi-quantitative integration method was used to compare the scores between the two groups,it was statistically significant(8.71 + 0.49 vs 3.50 + 0.53,t=15.59,p <0.001).The percentage of positive staining areas calculated by grayscale analysis in the two groups(11.40 + 0.36 vs 3.59 + 0.17,t= 55.11,p <0.001)was also statistically significant.Cyt-c,Caspase-3 and Caspase-9 were also highly expressed in the DMF group,while the Bcl-2 expression was low.The expression of apoptotic proteins were most prominent in the liver cells around the central vein,but were not expressed in the normal control group.5.Comparison of apoptotic proteins using the semi-quantitative analysis: Bax,Cyt-c,Caspase-3 and Caspase-9 were all highly expressed in the DMF group,and Bcl-2 was low expressed.The difference between high expression protein and low expression protein Bcl-2 was statistically significant(p <0.001).Conclusions: 1.The mitochondrial apoptotic pathway-related proteins Bax,Bcl-2,Cyt-c,Caspase-3,and Caspase-9 are highly expressed in rat livers with DMF-induced chronic toxic liver injury,but Bcl-2 is low expressed in rat livers with DMF-induced chronic toxic liver injury.2.Liver apoptosis is a common pathological lesion in chronic toxic liver injury induced by DMF in rats,which is prominent in hepatocytes surrounding the central vein. |
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