| Objective: To study liver fibrosis in rats caused by chronic drinking sodium arsenite,searching differences in protein in order to lay the foundation for further clarify the mechanism of liver fibrosis caused by arsenic. Methods: The 40 healthy 8-weeks-old Sprague wrote-Dawley(SD)male rats of Specefic pathogen Free(SPF) grade were randomly divided into 4 groups, which are control group(deionized water) and 0.68, 1.36,2.73 mg/kg sodium arsenite(iAs3+) infected group, respectively. Infected rats with drinking water freely in 24 consecutive weeks. Collected 24 h urine, blood and gathered the liver, using the high performance liquid chromatography(HPLC)-hydride generation atomic fluorescence spectrum(HPLC-HGAFS) method to detect the total arsenic content in the liver of rats and urine. Using enzyme-linked immunoassay(ELISA) to detect cereal third transaminase(ALT) and aspertate aminotransferase(AST) of liver function index,and liver fibrosis indexes. Liver tissue sections were stained with HE. To understand the pathological alteration under a light microscope and observe cell microstructure changes under electron microscope. Based on isobaric tags for relative and absolute quantitation(i TRAQ) reagent 8-plex experiment, combined with 2DLC-MS/MS to compare the differences of protein expression of the medium dose group and high dose group with control group in the rat liver tissue. Results: 1. Rats weight of high dose group is less than the control group at sodium arsenite exposured 20 weeks(P<0.05), the weight of high dose group is less than the control group, low dose group, medium dose group at24 week(P<0.05), and there was no statistically significant difference between the rest groups(P>0.05). 2. Medium and high dose group of organ coefficient of rat liver is higher than normal control group, the difference was statistically significant(P<0.05), high dosegroup is higher than low dose group, the difference was statistically significant(P< 0.05),there was no statistically significant difference between low dose and the middle dose group(P>0.05). 3. There was no obvious morphological change in low dose group under light and electron microscope, but fiber hyperplasia along with dose increased turn to obviously in medium and high dose group. 4. The level of UAs in each group increased with the infected of iAs, compared low and medium and high dose group with control group, the difference was statistically significant(P<0.05), UAs of low dose group is higher than the control group, middle dose group is higher than the low dose group, high dose group was higher than middle dose groups, differences were statistically significant(P<0.05). LAs differences among groups was statistically significant(P< 0.05), LAs levels is increased with the increase of the infected dose, LAs of low dose group is higher than the control group, middle dose group is higher than the low dose group, high dose group is higher than dose groups, differences were statistically significant(P<0.05). 5. The difference of AST and ALT between groups was statistically significant(P<0.05), the difference of AST of low dose group between control group has no statistically significant;Medium and high dose group were higher than low dose group and control group(P<0.05),there was no statistically significant difference in medium and high dose group(P>0.05).The ALT of low, medium and high dose group is higher than the controlgroup, there was no statistically significant difference between low dose group and the middle dose group(P>0.05), the difference between the rest groups were statistically significant(P<0.05),high dose group was higher than low, middle dose group(P<0.05). 6. “Four indexes of liver fibrosis” difference has statistically significant(P<0.05), In multiple comparison of HA, PCⅢ, LN, there was no statistically significant difference between low dose group and control group. Medium and high dose group were higher than low dose group and control group, the difference was statistically significant, and Significant statistics difference were found between high dose group and middle dose group(P<0.05). The content of Ⅳ-C of middle and high dose group is higher than the control group and low dose group(P<0.05), and between rest two groups there was no statistically significant difference(P>0.05). 7. Combined ITRAQ with 2 DLC-MS/MS, selecting the confidence threshold of protein(Unused ProtScore)>1.3, at least 1 matched peptides within the 95%confidence interval for protein identification results, identified 2948 proteins. Compared by Venn diagram found three groups were detected for 2162 kinds of proteins, remove significant difference proteins in control group, the number of up-regulation proteins were687 kinds, down regulation were 548 kinds in middle dosage group; the number ofup-regulation proteins were 633 kinds, down regulation were 519 kinds in high dosage group; The differences of protein expression between medium and high dose group was not statistically significant(P>0.05). 8. Differences in expression related to the metabolism including AS3 MT, SHMT, CHDH, CTH, CSAD BHMT, up-regulated in middle and high dose group; There’re 2 kinds of protein of MTR, METK1 up-regulated,F1LRB8 down-regulated. There’re 18 were detected associated to the GSH protein,including Gsta1, Gsta4, Gsta5, Gstt1, Gstt2, Gstk1, Gstp1, Gstm1, Gstm2, Gstm3, Gss,Gpx1, Gpx4, Esd, Hagh, Glo1, Mgst1, B6DYQ5 are all up-regulated. Proteins associated with liver fibrosis were detected Hic-5, Gss and six kinds of Tpm, including two kinds of Tpm1, three kinds of Tpm2 and one kind of Tpm3, which expressing are all raised.Conclusion: There is accumulation of arsenic in the liver after chronic arsenic exposure and resulting in liver fibrosis and decline of liver function, and AS3 MT, MTR, MAT,SHMT, BHMT, CHDH, CTH, CSAD expression up-regulated, arsenic metabolism methionine cycle, folic acid cycle, sulfur transfer pathways are closely related. GSH play an important role in arsenic metabolism and liver fibrosis, Hic-5, GSS, TPM may be associated with the occurrence of liver fibrosis. |
PDF Full Text Request