This study aims to investigate the role of Notch pathway in the renal ischemia/reperfusion injury (IRI)-associated inflammation and apoptosis. Materials and methods: Male Sprague-Dawley rats were divided into three groups: normal saline (NS)-treated sham rats, NS-treated ischemia/reperfusion (I/R) rats, and N- [N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT) (a γ-secretase inhibitor) treated I/R rats. I/R rat model underwent nephrectomy of the right kidney and was subjected to 60 min of left renal pedicle occlusion followed by 24 h, 48 h, and 72 h of reperfusion, respectively. The levels of creatinine, urea nitrogen (BUN), interleukin (IL)-6, tumor necrosis factor (TNF)-α in serum samples and urinary N-acety-β-D-glucosaminidase (NAG) were assayed. Histological examinations were performed. The protein expression of Notch2, hairy/enhancer of split 1 (hes-1), NF-κB2, monocyte chemoattractant protein (MCP)-1, B-cell lymphoma 2 (bcl-2), and bcl-2-associated X (bax) were detected and the degree of apoptosis of tubular cells was evaluated. Results: Renal IR induced severe tubular damage, caused significant increases in the Scr, BUN, IL-6, TNF-α, urinary NAG, Notch2, hes-1, NF-κB2, MCP-1, ratio of tubule cells apoptosis, and reduction in the ratio of bcl-2 to bax. However, DAPT treatment significantly reduced the level of Scr, BUN, IL-6, TNF-α, and NAG. Thus, I/R activates Notch2/hes-1 signaling and DAPT treatment can ameliorate the severity of tubular damage after renal IRI, lower the expression of NF-κB2, MCP-1, and bax protein, increase the expression of bcl-2 protein, and reduce the ratio of terminal 2-deoxyuridine 5-triphosphate nick end-labeling-positive cells. Conclusion: Notch signaling plays an important role in the renal IRI-associated inflammation and apoptosis. DAPT can protect against IRI through partly suppressing inflammation and apoptosis, which could constitute a new target for AKI.

BackgroundAccelerated muscle atrophy is associated with a three-fold increase in mortality in chronic kidney disease (CKD) patients. It is suggested that hyperphosphatemia might contribute to muscle wasting, but the underlying mechanisms remain unclear. Although evidence indicates that autophagy is involved in the maintenance of muscle homeostasis, it is not known if high phosphate levels can result in activation of autophagy, leading to muscle protein loss.MethodsImmortalized rat L6 myotubes were exposed to a high concentration of phosphate, with or without autophagy inhibition. Myotube atrophy was examined by phase contrast microscopy. Autophagic activity was assessed by measuring the expression of microtubule-associated protein 1 light chain 3 (LC3) and p62 using quantitative real-time polymerase chain reaction and western blot.ResultsPhosphate induced cell atrophy in L6 myotubes in a dose- and time-dependent manner, and these responses were not associated with calcification or osteogenesis. Phosphate also dose- and time-dependently increased the LC3-II/LC3-I ratio. Inhibition of autophagy with wortmannin or knockdown of Atg5 significantly suppressed myotube atrophy caused by high phosphate concentration.ConclusionsHigh phosphate concentration induces muscle cell atrophy through the activation of autophagy. Targeting autophagy could be a therapeutic strategy for preventing muscle wasting caused by hyperphosphatemia.

Vascular calcification (VC) is common in chronic kidney disease (CKD), where cardiovascular mortality remains the leading cause of death. Here, we examined the role of high-mobility group box1 (HMGB1), a nuclear DNA-binding protein involved in inflammation, in aortic calcification and renal dysfunction induced by high phosphate in a mouse model of CKD induced by 5/6 nephrectomy. HMGB1 and kidney function markers were measured by ELISA in the serum of CKD patients and in CKD mice. Sections of the aortas of mice were analyzed by immunofluorescence and Alizarin red staining, and protein lysates were generated to analyze the expression of related proteins in response to silencing of HMGB1 or β-catenin by western blotting. Our results showed that serum HMGB1 levels were significantly higher in CKD patients than in healthy controls and related to disease stage. High phosphate promoted the translocation of HMGB1 from the nucleus to the cytosol and aortic calcification in CKD mice in vivo, whereas HMGB1 knockdown ameliorated part of renal and vascular function. β-catenin silencing reversed high phosphate-induced calcification and restored renal marker levels. Taken together, our results suggest that HMGB1 is involved in VC associated with CKD via a mechanism involving the β-catenin.

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