Volume 117, Issue 4, Pages 953-961 (October 1999) Secretory leukocyte protease inhibitor in mice regulates local and remote organ inflammatory injury induced by hepatic ischemia/reperfusion  Alex B. Lentsch, Hiroyuki Yoshidome, Roscoe L. Warner, Peter A. Ward, Michael J. Edwards  Gastroenterology  Volume 117, Issue 4, Pages 953-961 (October 1999) DOI: 10.1016/S0016-5085(99)70355-0 Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 1 Effects of SLPI on liver and lung injury induced by hepatic ischemia/reperfusion. (A) Hepatocellular injury was assessed by measuring ALT levels in serum. (B) Liver and (C) lung wet to dry weight ratios were determined. Mice underwent sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Saline or SLPI (100 μg) was infused intravenously before induction of ischemia and again at the onset of reperfusion. Values represent mean ± SEM; n = 10 (A and B) or n = 5 (C) per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 2 Effects of SLPI on (A) liver and (B) lung neutrophil accumulation. Tissue MPO content was determined in mice undergoing sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Saline or SLPI (100 μg) was infused intravenously before induction of ischemia and again at the onset of reperfusion. Values represent mean ± SEM; n = 10 (A) or n = 5 (B) per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 3 Effects of SLPI on serum levels of (A) TNF-α and (B) MIP-2. Serum contents of TNF-α and MIP-2 were determined by ELISA in mice undergoing sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Saline or SLPI (100 μg) was infused intravenously before induction of ischemia and again at the onset of reperfusion. Values represent mean ± SEM; n = 10 per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 4 Effects of SLPI on liver NF-κB activation induced by hepatic ischemia/reperfusion. NF-κB activation in liver tissue harvested from mice undergoing sham surgery or 90 minutes of hepatic ischemia and 4 hours of reperfusion was analyzed by EMSA. Saline or SLPI (100 μg) was infused intravenously before induction of ischemia and again at the onset of reperfusion. Results are representative of 2 independent experiments. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 5 Expression of SLPI mRNA in liver during hepatic ischemia/reperfusion injury. (A) Liver RNA extracts were analyzed by RT-PCR. (B) Ethidium bromide–stained products were digitized using image analysis, and SLPI mRNA expression is represented as the densitometric ratio of SLPI mRNA to β-actin mRNA. Results are representative of 2 independent experiments. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 6 Expression of SLPI protein in liver during hepatic ischemia/reperfusion injury. (A) Immunostaining for SLPI in frozen liver sections from sham controls showed no staining. (B) Irregular hepatic lobular staining is observed in livers from mice undergoing 90 minutes of ischemia. (C) After hepatic ischemia and 1 hour of reperfusion, there was nonuniform staining for SLPI in hepatic lobules. Inset shows a higher magnification of SLPI-positive cells, which were identified as hepatocytes. (D) After hepatic ischemia and 4 hours of reperfusion, very little staining for SLPI remains. Where present, only small areas of hepatic lobules still retain staining for SLPI. Hepatic parenchyma shows evidence of focal necrosis. (Original magnification: A-D, 20×; C inset, 50 ×; counterstained with hematoxylin.) Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 7 Effects of anti-SLPI on serum levels of (A) TNF-α and (B) MIP-2. Serum contents of TNF-α and MIP-2 were determined by ELISA in mice undergoing sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Preimmune IgG (200 μg) or anti-SLPI (200 μg) was administered intravenously immediately before ischemia. Values represent mean ± SEM; n = 6 per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 8 Effects of anti-SLPI on (A) liver and (B) lung neutrophil accumulation. Tissue MPO content was determined in mice undergoing sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Preimmune IgG (200 μg) or anti-SLPI (200 μg) was administered intravenously immediately before ischemia. Values represent mean ± SEM; n = 6 per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 9 Effects of anti-SLPI on liver and lung injury induced by hepatic ischemia and reperfusion. (A) Hepatocellular injury was assessed by measuring serum levels of ALT. (B) Liver and (C) lung wet to dry weight ratios were determined. Mice underwent sham surgery (▧) or 90 minutes of hepatic ischemia and 4 hours of reperfusion (■). Preimmune IgG (200 μg) or anti-SLPI (200 μg) was administered intravenously immediately before ischemia. Values represent mean ± SEM; n = 6 per group. Gastroenterology 1999 117, 953-961DOI: (10.1016/S0016-5085(99)70355-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions