(A) The expressions of cleaved caspase-3 and CHOP were determined by immunoblotting assay

(A) The expressions of cleaved caspase-3 and CHOP were determined by immunoblotting assay. Quantitative analyses of the relative levels of protein expression. Cleaved caspase-3 and CHOP protein bands were quantified and normalized to -actin. The mean value obtained from sham-operated mice was arbitrarily defined as 1. There were 6 mice in each group and data were expressed as mean SD. *P 0.05 vs. sham-operated controls. CHOP knockout led to reduced apoptosis and attenuated renal IRI To evaluate possible differences between CHOP?/? and wild-type (WT) mice in susceptibility to renal ischemic injury, both strains were subjected to renal IR procedures. At 24 h after the initiation of reperfusion, the serum levels of creatinine (Cr) and blood urea nitrogen (BUN) were evaluated and the results were shown in Physique 2. Compared with WT controls, CHOP?/? mice exhibited a significant decrease in both Cr and BUN levels, suggesting attenuated renal dysfunction. Individual groups of mice were subjected to the same IR procedures and survival was observed and recorded for the following 7 days. No deaths occurred in both strains subjected to the sham operation (data not shown). However, no WT mice could survive the IR challenge and all of them died between 48 h and 72 Rabbit Polyclonal to OR2G3 h after IR. By contrast, most CHOP?/? mice survived (80%, P 0.05). The observations were reinforced by histological evidence in PAS staining, MPO activity and TUNEL assay. The evaluation of renal active caspase-3 expression also showed consistent results (Figure 2). Open in a separate window Figure 2 The effect of CHOP inactivation on renal IRI. CHOP?/? and wild-type mice L-Asparagine were subjected to right nephrectomy, followed by IR (left kidney) or sham-operation.Serum creatinine (A) and BUN (B) concentrations at 24 h after the initiation of reperfusion were shown (n?=?8 per group). (C) Survival of WT and CHOP?/? mice after renal IR operations (n?=?20 per group). CHOP knockout led to a significant survival advantage by Kaplan-Meier analysis (log-rank test, P 0.05). (D) Representative PAS-stained sections from post-ischemic kidneys harvested at 24 h (original magnification, 200). (E) Representative renal MPO staining (original magnification, 200). (F) TUNEL assay (original magnification, 400). (G) Representative immunoblotting results of the non-IR right kidneys and the operated left kidneys (6 hours after reperfusion). (HCI) Quantitative analyses of the relative levels of protein expression. Cleaved caspase-3 and CHOP protein bands were quantified and normalized to -actin. The mean value obtained from non-IR WT kidneys was arbitrarily defined as 1. There were 6 mice L-Asparagine in each group and data were expressed as mean SD. *P 0.05 vs. WT/IR L-Asparagine group. The alleviated renal IRI in CHOP?/? mice was not a result of CHOP deficiency in inflammatory cells, but in renal parenchymal cells Although the ultimate result of IRI was the death of renal parenchymal cells, the full development of injury was critically dependent on inflammatory responses. Since CHOP was reported to play a role in inflammatory injury in kidneys [8], we investigated whether CHOP induction in inflammatory cells was involved in IRI, by performing bone marrow transplantation (BMT) 30 days before renal IR procedures. At 24 h after IR serum Cr L-Asparagine and BUN levels were evaluated and shown in Figure 3. IRI was significantly attenuated in BMT (WTCHOP?/?), but not BMT (CHOP?/?WT) mice, indicating that CHOP expression in inflammatory cells didnt play a part in this setting. Survival observations.