Objective To investigate the effect of CD226 gene knockout(CD226-/-)on carbon tetrachloride(CCl4)-induced liver fibrosis in mice.Methods Eight 6-8 week-old male wild-type and CD226-/-C57BL/6 mice were used,with weights of(22.16±2.24)g and(21.84±2.50)g,respectively.Four mice each from wild-type and CD226-/-mice were injected intraperitoneally with CCl4 to establish a liver fibrosis model,namely,the liver fibrosis group and the CD226-/-liver fibrosis group,and the remai-ning four mice each corresponded to the control group and the CD226-/-control group.The effect of CD226-/-on liver mass,liver index and Ishak fibrosis score were observed.HE,Masson,and Sirius red staining were performed to observe the liver injury and liver fibrosis.Immunohistochemistry and Western blot were used to detect the protein expression levels of collagen type Ⅰ(Collagen Ⅰ)and α-smooth muscle actin(α-SMA)in liver tissues.Flow cytometry was used to analyze the proportion of T lymphocytes and their intracellular IFN-γ secretion levels in the peripheral blood and liver tissues.The mRNA expression level of transforming growth factor-β1(TGF-β1)in the liver tissues was detected by real-time fluorescence quantita-tive PCR(qPCR).Results Compared to the liver fibrosis group,the CD226-/-liver fibrosis group showed an increase in liver mass[(2.00±0.14)g vs.(1.41±0.16)g]and liver index(7.25±0.59 vs.5.33±0.30),and a decrease in Ishak's fibrosis score[(6.75±0.96)score vs.(10.00±1.41)score],and the differences were statistically significant(all P＜0.05).HE staining showed disorganization of the liver tissue structure,and degeneration and necrosis of the hepatocytes both occurred in the liver fibro-sis group and CD226-/-liver fibrosis group,but the degree of inflammatory cell recruitment was milder in the CD226-/-liver fibrosis group than the liver fibrosis group.Sirius red and Masson staining revealed reduced collagen fibre deposition in the CD226-/-liver fibrosis group compared to the liver fibrosis group.Immunohistochemical staining showed that the expression of Collagen Ⅰ[(4.38±0.51)％vs.(6.55±1.40)％]and α-SMA[(0.77±0.20)％vs.(1.20±0.24)％]were reduced in the CD226-/-liver fibrosis group compared to the liver fibrosis group,and the differences were statistically significant(both P＜0.05).Western blot indicated that the relative expression level of Collagen Ⅰ was reduced in the CD226-/-liver fibrosis group compared to the liver fibrosis group(0.35±0.14 vs.1.66±0.73),and the difference was statistically significant(P＜0.05).Flow cytometry revealed increased intracellular IFN-γ secretion by T lymphocytes in the peripheral blood in the CD226-/-liver fibrosis group compared to the liver fibrosis group.qPCR result revealed that the relative mRNA expressions of TGF-β1 was reduced in the CD226-/-liver fibrosis group compared to the liver fibrosis group(0.38±0.18 vs.0.61±0.28),and the difference was statistically significant(P＜0.05).Conclusion CD226-/-attenuates CCl4-induced hepatic inflam-matory infiltration and hepatic fibrosis,which may be related to its affected intrahepatic T lymphocytes and intracellular IFN-γ secretion.
维普
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