Specific role of interleukin-1 in hepatic neutrophil recruitment after ischemia/reperfusion

Abstract

Hepatic ischemia/reperfusion injury is caused primarily by the products of neutrophils recruited into the liver after reperfusion. The mediators responsible for the development of this inflammatory response are thought to be tumor necrosis factor-alpha and interleukin (IL)-1. Although there is abundant evidence to support a role for tumor necrosis factor-alpha, much less is known about the function of IL-1 in this injury. In the present studies, we investigated whether IL-1 was a critical mediator for the induction of liver inflammation after ischemia/reperfusion. Wild-type and IL-1 receptor I-knockout (IL-1RI(-/-)) mice were exposed to 90 minutes of partial hepatic ischemia and up to 24 hours of reperfusion. In wild-type mice, IL-1beta expression was maximal after ischemia and 8 hours of reperfusion. At the same time, both wild-type and IL-1RI(-/-) mice had severe liver injury as assessed by serum alanine aminotransferase levels and hepatic histopathology. However, IL-1RI(-/-) mice had significantly less neutrophil accumulation in liver tissues as measured by liver myeloperoxidase content and histology. The reduction in hepatic neutrophil recruitment in IL-1RI(-/-) mice was associated with decreased activation of the transcription factor, nuclear factor-kappaB, and reduced expression of the CXC chemokine, macrophage inflammatory protein-2. These data suggest that IL-1 functions to augment neutrophil accumulation, but does not play an essential role in this response.

Figure 1.

Hepatic expression of IL-1β mRNA during ischemia/reperfusion injury. RNA extracts from wild-type mice were analyzed for IL-1β and GAPDH expression by RNase protection assay. Autoradiographs were digitized and IL-1β mRNA expression was expressed as percent GAPDH mRNA expression.

Figure 2.

Serum levels of IL-1β after hepatic ischemia/reperfusion. Serum samples from wild-type mice were analyzed by enzyme-linked immunosorbent assay. Data are expressed as mean ± SEM with n = 5 to 10 per group. *, P < 0.05 compared to the sham group.

Figure 3.

Serum levels of TNF-α in wild-type and IL-1RI^−/− mice after hepatic ischemia and 8 or 24 hours of reperfusion. Serum samples were analyzed by enzyme-linked immunosorbent assay. Data are expressed as mean ± SEM with n = 8 to 10 per group.

Figure 4.

Liver injury in wild-type and IL-1RI^−/− mice after hepatic ischemia/reperfusion. Hepatocellular injury was assessed by measuring serum ALT levels. Data are expressed as mean ± SEM with n = 8 to 10 per group.

Figure 5.

Histological analysis of livers from wild-type and IL-1RI^−/− mice after hepatic ischemia/reperfusion. Normal liver architecture was observed in sham-operated wild-type (A) and IL-1RI^−/− (B) mice. After ischemia and 8 hours of reperfusion, similar degrees of hepatocyte necrosis were observed in wild-type (C) and IL-1RI^−/− (D) mice. After ischemia and 24 hours of reperfusion, there were similar degrees of necrosis and areas of normal liver architecture, indicating resolution of liver injury. In wild-type mice, ischemia/reperfusion induced abundant neutrophil accumulation around central veins and more diffusely in sinusoids (C and E). In IL-1RI^−/− mice, neutrophils were observed primarily around central veins (D and F). Original magnifications, ×100.

Figure 6.

Liver neutrophil recruitment in wild-type and IL-1RI^−/− mice after hepatic ischemia/reperfusion. Liver neutrophil accumulation was assessed by liver content of MPO. Data are expressed as mean ± SEM with n = 8 to 10 per group. *, P < 0.05 compared to wild-type mice.

Figure 7.

Expression of MIP-2 in wild-type and IL-1RI^−/− mice during hepatic ischemia/reperfusion injury. A: MIP-2 and β-actin mRNA expression were analyzed by RT-PCR in sham-operated mice and mice undergoing hepatic ischemia and 1 or 8 hours of reperfusion. B: Serum levels of MIP-2 were measured in wild-type and IL-1RI^−/− mice after hepatic ischemia/reperfusion. Data are expressed as mean ± SEM with n = 8 to 10 per group. *, P < 0.05 compared to wild-type mice.

Figure 8.

Activation of the transcription factors, NF-κB and AP-1, in wild-type and IL-1RI^−/− mice during hepatic ischemia/reperfusion injury. Liver nuclear extracts from sham-operated mice and mice undergoing hepatic ischemia and 1 or 8 hours of reperfusion were subjected to electrophoretic mobility shift assay. Results are representative of duplicate experiments.