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Objective To investigate the role of CD4+ CD25+ regulatory T lymphocyte(Tregs)in the immunological pathogenesis of liver fibrosis.Methods Twenty-six rats were divided into two groups:control group(6 rats)and model group(twenty rats).The rat model of liver fibrosis was induced bv subcutaneous injection of carbon tetrachloride(CCl4).The serums were collected for detection of hepatic function and fibrosis parameters.Hepatic tissue samples were used to observe the histopathological changes.The flow cytometry was used to detect the proportions of Tregs in both peripheral blood and spleen.The data were evaluated by t-test.The relationship between two variables was analyzed using Pearson linear correlation.Results The levels of serum alanine aminotransferase (ALT)and aspartate aminotransferase (AST) were significantly increased,but the level of serum albumin (Alb) was obviously decreased.The concentrations of serum hyaluronic acid (HA),procollagen type Ⅲ(PCⅢ),collagen type Ⅳ(CⅣ)of the model group increased to(177.42±61.25)μg/L,(34.86±7.47)μg/L and(7.32±3.71)μg/L,respectively,which were higher than those in the control group(t=-3.670,-5.661,-3.950,respectively;all P0.05).Conclusion These findings suggest that the reduction of CD4+ CD25+ Tregs probably play an important role in the immunological pathogenesis of liver fibrosis.
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Objective To investigate the dynamic expressions of exchange protein directly activated by cyclic adenosine monophosphate (cAMP) (Epac) in rat model of hepatic fibrosis(HF).Methods Forty-two male SD rats were divided into control group (n = 6) and model group (n = 36)which was divided into six subgroups of day 4, week 1, week 2, week 4,week 6 and week 8 with six rats in each subgroup. The rat model of HF was established by intraperitoneal injection of dimethylnitrosamine (DMN). The pathological changes of liver were observed by Hematoxylin-Eosin and Masson staining. Reverse transcription-polymerase chain reaction (RT-PCR),immunohistochemistry and Western blot were employed to detect the mRNA and protein expressions of Epac1, Epac2 and transforming gronth factor (TGF)β1 during the process of modeling and localization in the liver. The statistical analysis was done using one-factor ANOVA, LSD-t test,Dunnett T3 test and Pearson linear correlation analysis. Results Rat model of liver fibrosis was established successfully. In control group, Epac1 (0. 031 28±0. 008 96) and Epac2 protein (0.034 43±0. 002 45) mainly expressed in the cytoplasm of hepatocytes. In model group, the level of Epac1 decreased at day 4 (0. 023 97±0. 003 81) and week 1 (0. 015 81±0. 002 48) ,then began to increase at week 2 of modeling and peaked at week 6 (0. 039 54±0. 001 43), which had statistical significance compared to the control group (t= 5.47,11.58 and - 6.18, respectively; all P＜0.05). Epac2 protein expression declined after modeling, reached the lowest level at week 4 (0. 011 21 ±0. 001 32), which had statistical significance compared to the control group (t= 24. 50, P＜0. 05). TGFβ1 protein expression increased after modeling and peaked at week 4 (0. 011 30±0.001 03) which had statistical significance (t= -23. 36, P＜0. 05) compared to the control group (0. 002 08 ±0. 000 18). The expressions of Epac1, Epac2 and TGFβ1 mRNA were consistent with the trend of protein levels.Correlation analysis showed that Epac1 protein was positively correlated with the course of HF (r =0. 703, P＜0.01 ), while Epac2 protein was negatively correlated (r = - 0. 409, P＜0.05). Conclusions During the progression of HF, Epac1 expression tends to decrease firstly and increase afterwards,while Epac2 expression declines continually. Epac may be involved in the pathogenesis of HF.
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Objective To explore the expression and significance of uncoupling protein (UCP)2in rats models of acute liver failure (ALF). Methods Thirty-six healthy male SD rats were randomly divided into normal control group and model group, and the model group was divided into 5 subgroups:6, 12, 24, 36 and 48 hours sub groups with 6 rats in each sub group. The rat model of ALF was established by intraperitoneal injections of D-galactosamine (D-Gal) and lipopolysaccharide (LPS).Sections of liver tissue were stained with hematoxylin and eosin and observed under optical microscope.UCP2 and UCP2 mRNA in rat liver were determined at different time points with immunohistochemical method and reverse transcription-polymerase chain reaction ( RT-PCR ),respectively. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels and malondialdehyde (MDA) concentration in the liver tissues were analyzed at the same time points.Comparisons among all the experimental groups were done by SNK test. Results Infiltration of inflammatory cells and necrosis of hepatic cells were marked in model group,and ALT, AST and MDA in model group were significantly higher than those in control group [(24. 0 ± 2. 0) U/L, (82. 3±16. 9) U/L, (2. 55±0. 22)μmol/g] at all time points. And they reached a peak at 24 h [(8346. 7±1363. 1) U/L, (9766. 7±1274. 1) U/L, (8. 34±1. 13) μmol/g; all P<0. 05]. UCP2 and UCP2 mRNA expressed scarcely in the liver tissues of control group, while increased markedly from 6 to 48 hours after D-Gal/LPS challenge in model group (P<0. 05). They both reached a peak at 24 h. And the discrepancy between consecutive experimental group had statistical significance ( P < 0. 05).Conclusions The rat model of ALF was established successfully by intraperitoneal injections of D-gal and LPS. The expression levels of UCP2 mRNA and UCP2 are consistent with the extent of liver injury and the level of oxidative stress in the rat model of ALF.
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Objective To study the protective role of ulinastatin in acute liver failure (ALF) and the effect on the expression of hemeoxygenase-1 (HO-1).Methods Sixty-six S-D rats were divided into three groups:control group,ALF group (model group) and ulinastatin group (intervention group).The rat model of ALF was induced by intraperitoneal injection of D-galactosamine (D-Gal) and lipopolysaccharide (LPS). The serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and malondialdehyde (MDA) were detected dynamically after 6,12,24,36 and 48 h injection.HO-1 mRNA expression in liver tissue was determined by reverse transcriptionpolymerase chain reaction (RT-PCR), and the expression of HO-1 protein was detected by immunohistochemistry.The differences among multiple groups were compared by univariate ANOVA and pairwise comparison was done by least significant difference (LSD). Results D-Gal/LPS injections successfully induced ALF rat model which presented with elevated levels of serum ALT,AST and liver MDA after 6 h injections (F=23.864,38.446,18.051,respectively,all P＜0.01),and peaked at 12-24 h after injections.Twenty-four hours after D-Gal/LPS treatment,the levels of serum ALT,AST and MDA in model group and intervention group were (8 346.7±1 363.1) U/L vs(4 151.3±970.0) U/L,(9 766.7±1.274.1) U/L vs (4 696.7±1 476.9) U/L,(8.34±1.13)μmol/g vs (4.66±0.91 ) μmol/g,respectively,which were significantly higher than those e (24.0±2.0) U/L,(82.3±16.9) U/L,(2.55±0.22) μmol/g,respectively] in control group (F=55.684,55.501,47.843,respectively,all P＜0.01);while those in intervention group were much lower than those in model group (P＜0.01).The expressions of HO-1 mRNA and protein in model group were significantly increased than those in control group (P＜0.01),while those in intervention group were even higher (P＜0.01).Conclusion Ulinastain could up-regulate the expressions of HO-1 mRNA and protein,which indicates that ulinastain may play anti-oxidant and anti-inflammatory roles in ALF through HO-1 pathway.