Mechanism of infliximab in preventing autoimmune hepatitis in mice model / 中华消化杂志

ABSTRACT

Objective:To observe the preventive effects of infliximab in autoimmune hepatitis (AIH) and to explore its mechanism.Methods:The mice AIH model was established by injecting concanavalin A (Con-A) into the caudal vein. Forty mice were divided into prevention group and control group, with 20 mice in each group. The mice of prevention group were injected intravenously with infliximab (20 mg/kg) one hour before Con-A injection and the mice of control group were administrated with 200 μL phosphate buffered saline (PBS). Serum was collected 3, 8, 12 and 24 h after Con-A/PBS injection. The serum level of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) was detected by colorimetry. The level of cytokine interleukin (IL)-6, IL-1β, interferon gamma (IFN-γ), IL-4, IL-17A, IL-10 and chemokine C-X-C motif ligand 10 (CXCL10) was measured by enzyme-linked immunosorbent assay (ELISA). Liver samples were taken 12 h after Con-A/PBS injection for hematoxylin-eosin staining. Liver infiltrated lymphocytes were assessed by flow cytometry. The expression of T-box transcription factor 21 ( TBX21), GATA binding protein 3 ( GATA3), RAR related orphan receptor C ( RORC) and CXCL10 at mRNA level was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The expression of CXCL10 in liver was detected by Western blotting. Paired t test and one-way analysis of variance were used for statistic analysis. Results:At 8, 12, and 24 h after Con-A injection, the serum ALT level, AST level, IL-1β and IFN-γ of prevention group were all lower than those of control group ((545.8±190.3) U/L vs. (865.8±237.7) U/L, (947.6±267.9) U/L vs. (1 448.0±403.5) U/L, (508.6±131.1) U/L vs. (976.6±207.6) U/L; (620.7±132.0) U/L vs. (952.9±106.8) U/L, (801.6±212.0) U/L vs. (1 424.8±236.0) U/L, (632.1±117.8) U/L vs. (1 008.3±187.5) U/L; (31.38±10.12) ng/L vs. (48.12±11.53) ng/L, (39.34±11.40) ng/L vs. (60.00±14.17) ng/L, (29.49±8.22) ng/L vs. (46.89±5.50) ng/L; and (432.93±66.82) ng/L vs. (674.66±97.88) ng/L, (655.09±169.17) ng/L vs. (937.90±166.36) ng/L, (263.40±54.97) ng/L vs. (410.74±86.64) ng/L), and the differences were statistically significant ( t = 2.350, 2.308, 4.263, 4.374, 4.860, 3.806, 2.440, 2.541, 3.939, 4.560, 2.660 and 3.210; all P<0.05). The serum IL-6 levels 3, 8, 12 and 24 h after Con-A injection of prevention group were all lower than those of control group ((1 075.79±303.77) ng/L vs. (1 914.48±317.80) ng/L, (1 945.97±271.85) ng/L vs. (2 100.80±378.42) ng/L, (1 578.60±504.54) ng/L vs. (2 525.40±406.55) ng/L, (1 020.64±280.03) ng/L vs. (1 582.00±311.96) ng/L), and the differences were statistically significant ( t=4.266, 2.903, 3.267 and 2.994; all P < 0.05). At 3 h after Con-A injection, serum CXCL10 level and CXCL10 mRNA expression in liver tissues of prevention group were both lower than those of control group ((1 755.8±148.1) ng/L vs. (2 102.0±334.0) ng/L and 7.20±3.00 vs. 27.60±1.90), and the differences were statistically significant ( t=2.356 and 2.623, both P<0.05). At 3 and 8 h after Con-A injection, T- bet expression at mRNA level in liver tissues of prevention group was lower than that of control group (6.94±2.29 vs. 15.20±3.48 and 9.38±3.48 vs. 18.17±4.48), and the differences were both statistically significant ( t = 4.427 and 3.673, both P<0.05). However, 3, 8, 12 and 24 h after Con-A injection, there were no statistically significant differences in serum IL-4, IL-17A, IL-10, or GATA3 or RORC expression at mRNA level between prevention group and control group (all P > 0.05). Conclusions:Infliximab has certain preventive effects in mice AIH model, which may be achieved by antagonizing TNF-α and decreasing the expression of CXCL10 in liver, reducing the infiltration of T-helper 1 cells and CD8 + T cells into liver, and by reducing T lymphocyte activation induced by inflammatory cytokines thus alleviating the damage of T lymphocytes to hepatocytes.