Echinomycin Decreases Induction of Vascular Endothelial Growth Factor and Hepatocyte Regeneration in Acetaminophen Toxicity in Mice

Abstract

Up-regulation of vascular endothelial growth factor (VEGF) is important to hepatocyte regeneration in the late stages of acetaminophen (APAP) toxicity in the mouse. The following study was conducted to examine the relationship of hypoxia inducible factor 1α (HIF-1α) to VEGF and hepatocyte regeneration in APAP toxicity by using an inhibitor of HIF-1α DNA-binding activity, echinomycin (EC). B6C3F1 male mice were treated with APAP (200 mg/kg IP), followed by EC (0.15 mg IP) and killed at 4 hr. Serum ALT, necrosis, hepatic glutathione (GSH) and APAP protein adducts were comparable in the APAP/EC and the APAP/veh mice at 4 hr. Additional studies showed that high dose EC (0.3 mg) reduced hepatic VEGF but also lowered hepatic GSH. Subsequent studies were performed using the 0.15 mg dose of EC. Although EC 0.15 mg had no effect on hepatic VEGF levels at 8 hr, by 24 hr VEGF levels were decreased by 40%. Toxicity (ALT and histopathology) were comparable in the APAP and APAP/EC groups at 24 and 48 hr. Proliferating cell nuclear antigen expression was reduced by both western blot analysis and immunohistochemical staining in the APAP/EC mice at 48 hr. The data support the hypothesis that induction of HIF-1α, its binding to DNA and subsequent expression of vascular endothelial growth factor are important factors in hepatocyte regeneration in APAP toxicity in the mouse.

Ringer's lactate improves liver recovery in a murine model of acetaminophen toxicity

“…In this study, the number of PCNA positive cells was large, and the vast majority of PCNA positive cells were inflammatory cells, this is different from the other studies which have shown hepatocytes at bordering area of necrosis express PCNA during injury progression [ 37 , 38 ]. There are two possibilities for this phenomenon: (1) The specificity and the sensitivity of the first antibody to PCNA might markedly influence the results, because the other two studies used monoclonal antibody to PCNA from DakoCytomation (Carpinteria, CA, USA) and Abcam, (Cambridge, MA), our PCNA staining kit was purchased from Invitrogen (Camarillo, CA, USA).…”

“…There are two possibilities for this phenomenon: (1) The specificity and the sensitivity of the first antibody to PCNA might markedly influence the results, because the other two studies used monoclonal antibody to PCNA from DakoCytomation (Carpinteria, CA, USA) and Abcam, (Cambridge, MA), our PCNA staining kit was purchased from Invitrogen (Camarillo, CA, USA). (2) Animal species might also influence the results, because 24 hrs after APAP administration, PCNA expression was apparent in B6C3F1 mice [ 37 ] and B6J129 SVF2 mice [ 38 ], however, PCNA staining was negative or occasional in C57/BL6 mice in our study at 24 h time point, and our result was consistent with another study [ 32 ] in which BrdU (5-bromo-2-deoxyuridine) immunohistochemistry was performed to detect hepatocyte regeneration in C57/BL6 mice.…”

Effect of trifluoperazine on toxicity, HIF-1α induction and hepatocyte regeneration in acetaminophen toxicity in mice

“…Echinomycin, a small molecule inhibitor of HIF-1 DNA binding, lowered VEGF protein expression in APAP toxicity in mice (Micheli-Halle, 2011). Despite lower HIF-1α induction, APAP/TFP mice had relatively higher levels of hepatic VEGF at 8 and 24 h, compared to the APAP mice (Fig.…”

New Insights into the Acetaminophen Hepatotoxicity Research

“…animal species also markedly influence the results: 24 hrs after APAP administration, PCNA expression is apparent in B6C3F1 mice [42] and B6J129 SVF2 mice [43], however, BrdU and PCNA (Invitrogen) methods only show negative or occasional positive staining in C57/BL6 mice [10,11,44]; more importantly, 48 hrs after APAP subjected to C57/BL6 mice, both PCNA (Invitrogen) and BrdU methods show that the extent of hepatic PCNA / BrdU expression depends mainly on the extent of liver damage, because the expression of hepatic PCNA /BrdU is significantly correlated with the area of hepatocyte necrosis for each mouse: larger size of necrosis has larger number of PCNA/BrdU positive cells, while improved liver repair with smaller necrotic size has smaller number of PCNA/BrdU positive cells, this is not conventionally expected [10,11,44]. In addition, the number of PCNA-positive non-parenchymal cells is 10 times larger than the number of BrdU-positive hepatocyte in C57/BL6 mice during APAP hepatotoxicity [11], suggesting that currently the nonparenchymal cells are likely underestimated, and the number of the non-parenchymal cell loss could be larger than the hepatocyte loss during APAP toxicity, the latter could explain the impaired immune function and the higher incidence of gram-negative infection during hepatotoxicity [11,45], further investigation is needed to focus on non parenchymal cell loss in APAP overdose.…”