Urine/Plasma Neutrophil Gelatinase Associated Lipocalin Ratio Is a Sensitive and Specific Marker of Subclinical Acute Kidney Injury in Mice

Background Detection of acute kidney injury (AKI) is still a challenge if conventional markers of kidney function are within reference range. We studied the sensitivity and specificity of NGAL as an AKI marker at different degrees of renal ischemia. Methods Male C57BL/6J mice were subjected to 10-, 20- or 30-min unilateral renal ischemia, to control operation or no operation, and AKI was evaluated 1 day later by histology, immunohistochemistry, BUN, creatinine, NGAL (plasma and urine) and renal NGAL mRNA expression. Results A short (10-min) ischemia did not alter BUN or kidney histology, but elevated plasma and urinary NGAL level and renal NGAL mRNA expression although to a much smaller extent than longer ischemia. Surprisingly, control operation elevated plasma NGAL and renal NGAL mRNA expression to a similar extent as 10-min ischemia. Further, the ratio of urine to plasma NGAL was the best parameter to differentiate a 10-min ischemic injury from control operation, while it was similar in the non and control-operated groups. Conclusions These results suggest that urinary NGAL excretion and especially ratio of urine to plasma NGAL are sensitive and specific markers of subclinical acute kidney injury in mice

LPS-induced delayed preconditioning is mediated by hsp90 and involves the heat shock response in mouse kidney.

INTRODUCTION: We and others demonstrated previously that preconditioning with endotoxin (LPS) protected from a subsequent lethal LPS challenge or from renal ischemia-reperfusion injury (IRI). LPS is effective in evoking the heat shock response, an ancient and essential cellular defense mechanism, which plays a role in resistance to, and recovery from diseases. Here, by using the pharmacological Hsp90 inhibitor novobiocin (NB), we investigated the role of Hsp90 and the heat shock response in LPS-induced delayed renal preconditioning. METHODS: Male C57BL/6 mice were treated with preconditioning (P: 2 mg/kg, ip.) and subsequent lethal (L: 10 mg/kg, ip.) doses of LPS alone or in combination with NB (100 mg/kg, ip.). Controls received saline (C) or NB. RESULTS: Preconditioning LPS conferred protection from a subsequent lethal LPS treatment. Importantly, the protective effect of LPS preconditioning was completely abolished by a concomitant treatment with NB. LPS induced a marked heat shock protein increase as demonstrated by Western blots of Hsp70 and Hsp90. NB alone also stimulated Hsp70 and Hsp90 mRNA but not protein expression. However, Hsp70 and Hsp90 protein induction in LPS-treated mice was abolished by a concomitant NB treatment, demonstrating a NB-induced impairment of the heat shock response to LPS preconditioning. CONCLUSION: LPS-induced heat shock protein induction and tolerance to a subsequent lethal LPS treatment was prevented by the Hsp90 inhibitor, novobiocin. Our findings demonstrate a critical role of Hsp90 in LPS signaling, and a potential involvement of the heat shock response in LPS-induced preconditioning

Oxidative/Nitrative Stress and Inflammation Drive Progression of Doxorubicin-Induced Renal Fibrosis in Rats as Revealed by Comparing a Normal and a Fibrosis-Resistant Rat Strain

Chronic renal fibrosis is the final common pathway of end stage renal disease caused by glomerular or tubular pathologies. Genetic background has a strong influence on the progression of chronic renal fibrosis. We recently found that Rowett black hooded rats were resistant to renal fibrosis. We aimed to investigate the role of sustained inflammation and oxidative/nitrative stress in renal fibrosis progression using this new model. Our previous data suggested the involvement of podocytes, thus we investigated renal fibrosis initiated by doxorubicin-induced (5 mg/kg) podocyte damage. Doxorubicin induced progressive glomerular sclerosis followed by increasing proteinuria and reduced bodyweight gain in fibrosis-sensitive, Charles Dawley rats during an 8-week long observation period. In comparison, the fibrosis-resistant, Rowett black hooded rats had longer survival, milder proteinuria and reduced tubular damage as assessed by neutrophil gelatinase-associated lipocalin (NGAL) excretion, reduced loss of the slit diaphragm protein, nephrin, less glomerulosclerosis, tubulointerstitial fibrosis and matrix deposition assessed by periodic acid-Schiff, Picro-Sirius-red staining and fibronectin immunostaining. Less fibrosis was associated with reduced profibrotic transforming growth factor-beta, (TGF-beta1) connective tissue growth factor (CTGF), and collagen type I alpha 1 (COL-1a1) mRNA levels. Milder inflammation demonstrated by histology was confirmed by less monocyte chemotactic protein 1 (MCP-1) mRNA. As a consequence of less inflammation, less oxidative and nitrative stress was obvious by less neutrophil cytosolic factor 1 (p47phox) and NADPH oxidase-2 (p91phox) mRNA. Reduced oxidative enzyme expression was accompanied by less lipid peroxidation as demonstrated by 4-hydroxynonenal (HNE) and less protein nitrosylation demonstrated by nitrotyrosine (NT) immunohistochemistry and quantified by Western blot. Our results demonstrate that mediators of fibrosis, inflammation and oxidative/nitrative stress were suppressed in doxorubicin nephropathy in fibrosis-resistant Rowett black hooded rats underlying the importance of these pathomechanisms in the progression of renal fibrosis initiated by glomerular podocyte damage

The huge world of small RNAs: Regulating networks of microRNAs (Review)

MicroRNAs (miRNAs) are a recently discovered class of small, non-coding RNAs which do not code proteins. MiRNAs regulate gene expression by inhibiting protein translation from the messenger RNA. MiRNAs may function in networks, forming a complex relationship with diseases. Furthermore, specific miRNAs have significant correlation with diseases of divergent origin. After identification of disease-associated miRNAs, their tissue expression could be altered in a beneficial way by inhibiting or mimicking their effects. Thus, modifying the expression of miRNAs is a potential future gene-therapeutic tool to influence post-transcriptional regulation of multiple genes in a single therapy. In this review we introduce the biogenesis, mechanism of action and future aspects of miRNAs. Research on the post-transcriptional regulation of gene expression by miRNA may reshape our understanding of diseases and consequently may bring new diagnostic markers and therapeutic agents. Therapeutic use of miRNAs is already under clinical investigation in RNA interference trials

Activation of the miR-17 Family and miR-21 During Murine Kidney Ischemia-Reperfusion Injury.

Background: Ischemia-reperfusion (I/R) is the main cause of acute kidney injury (AKI) in patients. We investigated renal microRNA (miRNA) expression profiles and the time course of changes in selected miRNA expressions after renal I/R to characterize the miRNA network activated during development and recovery from AKI. Methods and Results: One day after lethal (30 minutes) and sublethal (20 minutes) renal ischemia, AKI was verified by renal histology (tubular necrosis, regeneration), blood urea nitrogen (BUN) level, renal mRNA expression, and plasma concentration of neutrophil gelatinase-associated lipocalin (NGAL) in C57BL/6J mice. On the first day after 30-minute, lethal I/R miR-21, miR-17-5p, and miR-106a were elevated out of the 21 miRNAs successfully profiled on the Luminex multiplex assay. After 20-minute, sublethal I/R, renal miR-17-5p and miR-106a expressions were elevated on the first and second days of reperfusion, while miR-21 expression increased later and lasted longer. Renal miR-17-5p and miR-21 expressions correlated with each other. Renal function returned to normal on the fourth day after sublethal I/R. Conclusions: Our results demonstrate that besides miR-21, miR-17-5p, and miR-106a are additionally activated during the maintenance and recovery phases of renal I/R injury. Furthermore, a correlation between renal miR-17-5p and miR-21 expressions warrants further investigation of how they may influence each other and the outcome of renal ischemia-reperfusion injury

Body temperature regulation determines immune reactions and species longevity

Temperature variations in various species have marked changes in body metabolism with higher temperatures associated with increased ageing. The observation that diets with calorie restriction are associated with species longevity are now related to the heat shock genes and body temperature regulation. Temperature increases that induce temperature dysregulation are connected to non alcoholic fatty liver disease (NAFLD) and the induction of diabetes and neurodegenerative diseases. Specific microRNAs are associated with heat shock gene regulation and override body temperature regulation relevant to adipose tissue-liver defects and insulin resistance. In geriatrics and diabetics complete heat shock gene inactivation is associated with mitochondrial apoptosis relevant to defective hepatic dietary fat and drug metabolism. The connections between core body temperature defects and autoimmune disease have now become important to determine programmed cell death in many cells and tissues with relevance to the global chronic disease epidemic and species survival