Distinct contributions of CD4+ T cell subsets in hepatic ischemia/reperfusion injury

Abstract

Helper T cells are known to mediate hepatic ischemia/reperfusion (I/R) injury. However, the precise mechanisms and subsets of CD4(+) T cells that contribute to this injury are still controversial. Therefore, we sought to determine the contributions of different CD4(+) T cell subsets during hepatic I/R injury. Wild-type, OT-II, or T cell receptor (TCR)-delta-deficient mice were subjected to 90 min of partial hepatic ischemia followed by 8 h of reperfusion. Additionally, wild-type mice were pretreated with anti-CD1d, -NK1.1, or -IL-2R-alpha antibodies before I/R injury. OT-II mice had diminished liver injury compared with wild-type mice, implicating that antigen-dependent activation of CD4(+) T cells through TCRs is involved in hepatic I/R injury. TCR-delta knockout mice had decreased hepatic neutrophil accumulation, suggesting that gammadelta T cells regulate neutrophil recruitment. We found that natural killer T (NKT) cells, but not NK cells, contribute to hepatic I/R injury via CD1d-dependent activation of their TCRs, as depletion of NKT cells by anti-CD1d antibody or depletion of both NKT cells and NK cells by anti-NK1.1 attenuated liver injury. Although regulatory T cells (Treg) are known to suppress T cell-dependent inflammation, depletion of Treg cells had little effect on hepatic I/R injury. The data suggest that antigen-dependent activation of CD4(+) T cells contributes to hepatic I/R injury. Among the subsets of CD4(+) T cells, it appears that gammadelta T cells contribute to neutrophil recruitment and that NKT cells directly injure the liver. In contrast, NK cells and Treg have little effects on hepatic I/R injury.

Fig. 1.

Response of OT-II mice to hepatic ischemia/reperfusion (I/R). A: liver injury, assessed by serum levels of alanine aminotransferase (ALT), was determined after 90 min of ischemia and 4 and 8 h of reperfusion. Data are means ± SE with n = 5 per group. *P < 0.05, compared with wild-type mice. B: neutrophil accumulation after 4 and 8 h of reperfusion was determined by liver content of myeloperoxidase (MPO). Data are means ± SE with n = 5 per group. C: liver histology. Normal hepatic architecture was observed in sham-operated wild-type (WT) mice. Sham-operated OT-II mice also showed normal hepatic architecture. After 8 h of reperfusion, livers from wild-type mice had large areas of necrosis, whereas livers from OT-II mice had less evidence of necrosis. Original magnification was ×50.

Fig. 2.

Effects of T cell receptor (TCR)-δ deficiency on hepatic I/R. A: liver injury, assessed by serum levels of ALT, was determined after 90 min of ischemia and 8 h of reperfusion. Data are means ± SE with n = 12–13 per group. KO, knockout. B: neutrophil accumulation after 8 h of reperfusion was determined by liver content of MPO. Data are means ± SE with n = 12–14 per group. *P < 0.05, compared with wild-type mice. C: liver histology. Normal hepatic architecture was observed in sham-operated wild-type mice and TCR-δ-deficient mice. After 8 h of reperfusion, livers from wild-type mice had large areas of necrosis with marked neutrophilic infiltrates, whereas TCR-δ-deficient mice had severe necrotic liver with reduced neutrophilic infiltrates. Original magnification was ×50.

Fig. 3.

Effects of depletion of natural killer T (NKT) cells on hepatic I/R. Wild-type mice were injected intraperitoneally with saline (control) or 100 μg of anti-CD1d antibody 4 h before operation. A: liver injury, assessed by serum levels of ALT, was determined after 90 min of ischemia and 8 h of reperfusion. Data are means ± SE with n = 5 per group. *P < 0.05, compared with control mice. B: neutrophil accumulation after 8 h of reperfusion was determined by liver content of MPO. Data are means ± SE with n = 5 per group. C: liver histology. Sham-operated wild-type mice had normal hepatic architecture. After 8 h of reperfusion, livers from wild-type mice had large areas of necrosis, whereas mice treated with anti-CD1d had less evidence of necrosis. Original magnification was ×50.

Fig. 4.

Effects of depletion of NK and NKT cells on hepatic I/R. Wild-type mice were injected intraperitoneally with saline (control) or 100 μg of PK136 48 h before operation. A: liver injury, assessed by serum levels of ALT, was determined after 90 min of ischemia and 8 h of reperfusion. Data are means ± SE with n = 10 per group. *P < 0.05, compared with control mice. B: neutrophil accumulation after 8 h of reperfusion was determined by liver content of MPO. Data are means ± SE with n = 10 per group.

Fig. 5.

Response of OT-II and wild-type mice depleted of NKT cells on hepatic I/R. Wild-type on OT-II mice were injected intraperitoneally with 100 μg of PK136 48 h before operation. A: liver injury, assessed by serum levels of ALT, was determined after 90 min of ischemia and 8 h of reperfusion. Data are means ± SE with n = 5 per group. B: neutrophil accumulation after 8 h of reperfusion was determined by liver content of MPO. Data are means ± SE with n = 5 per group. *P < 0.05; **P < 0.01; ***P < 0.001.