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Human miRNome profiling identifies microRNAs differentially present in the urine after kidney injury.

ABSTRACT: BACKGROUND:Extracellular microRNAs (miRNAs) have been proposed as potentially robust and stable biomarkers of various disease conditions. The primary objective of this study was to identify miRNAs differentially occurring in the urine that could serve as potential biomarkers of acute kidney injury (AKI), because traditional AKI markers have limitations with respect to sensitivity, specificity, and timeliness of diagnosis. METHODS:We profiled 1809 miRNAs in pooled urine samples from 6 patients with AKI and from 6 healthy controls. We measured the 378 stably detectable miRNAs in the 12 samples individually and selected the top 7 miRNAs that were most different in the urine of patients with AKI compared with the non-AKI control individuals. These miRNAs were assessed in a larger cohort of patients with AKI (n = 98: 71 AKI patients in the intensive care unit (ICU) and 27 kidney transplantation patients with biopsy-proven tubular injury) and patients without AKI (n = 97: 74 healthy volunteers and 23 ICU patients without AKI). RESULTS:We identified 4 miRNAs capable of significantly differentiating patients with AKI from individuals without AKI: miR-21 (P = 0.0005), miR-200c (P < 0.0001), miR-423 (P = 0.001), and miR-4640 (P = 0.0355). The combined cross-validated area under the ROC curve for these 4 miRNAs was 0.91. The imprecision with respect to miRNA isolation and reverse transcription efficiency was <9% across 224 samples. CONCLUSIONS:In this study we determined the entire miRNome of human urine and identified a panel of miRNAs that are both detectable noninvasively and diagnostically sensitive indicators of kidney damage.

SUBMITTER: Ramachandran K

PROVIDER: S-EPMC3870155 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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Human miRNome profiling identifies microRNAs differentially present in the urine after kidney injury.

Ramachandran Krithika K Saikumar Janani J Bijol Vanesa V Koyner Jay L JL Qian Jing J Betensky Rebecca A RA Waikar Sushrut S SS Vaidya Vishal S VS

Clinical chemistry 20131023 12

<h4>Background</h4>Extracellular microRNAs (miRNAs) have been proposed as potentially robust and stable biomarkers of various disease conditions. The primary objective of this study was to identify miRNAs differentially occurring in the urine that could serve as potential biomarkers of acute kidney injury (AKI), because traditional AKI markers have limitations with respect to sensitivity, specificity, and timeliness of diagnosis.<h4>Methods</h4>We profiled 1809 miRNAs in pooled urine samples fro ...[more]

PMID: 24153252

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Project description:Bone injury healing is an orchestrated process that starts with an inflammatory phase followed by repair and remodelling of the bone defect. The initial inflammation is characterized by local changes in immune cell populations and molecular mediators, including microRNAs (miRNAs). However, the systemic response to bone injury remains largely uncharacterized. Thus, this study aimed to profile the changes in the plasma miRnome after bone injury and determine its biological implications. Methods: A rat model of femoral bone defect was used, and animals were evaluated at days 3 and 14 after injury. Non-operated (NO) and sham operated animals were used as controls. Blood and spleen were collected and peripheral blood mononuclear cells (PBMC) and plasma were separated. Plasma miRnome was determined by RT-qPCR array and bioinformatics Ingenuity pathway analysis (IPA) was performed. Proliferation of bone marrow mesenchymal stem/stromal cells (MSC) was evaluated by Ki67 staining and high-throughput cell imaging. Candidate miRNAs were evaluated in splenocytes by RT-qPCR, and proteins found in the IPA analysis were analysed in splenocytes and PBMC by Western blot. Results: Bone injury resulted in timely controlled changes to the miRNA expression profile in plasma. At day 3 there was a major down-regulation of miRNA levels, which was partially recovered by day 14 post-injury. Interestingly, bone injury led to a significant up-regulation of let-7a, let-7d and miR-21 in plasma and splenocytes at day 14 relative to day 3 after bone injury, but not in sham operated animals. IPA predicted that most miRNAs temporally affected were involved in cellular development, proliferation and movement. MSC proliferation was analysed and found significantly increased in response to plasma of animals days 3 and 14 post-injury, but not from NO animals. Moreover, IPA predicted that miRNA processing proteins Ago2 and Dicer were specifically inhibited at day 3 post-injury, with Ago2 becoming activated at day 14. Protein levels of Ago2 and Dicer in splenocytes were increased at day 14 relative to day 3 post-bone injury and NO animals, while in PBMC, levels were reduced at day 3 (albeit Dicer was not significant) and remained low at day 14. Ephrin receptor B6 followed the same tendency as Ago2 and Dicer, while Smad2/3 was significantly decreased in splenocytes from day 14 relative to NO and day 3 post-bone injury animals. Conclusion: Results show a systemic miRNA response to bone injury that is regulated in time and is related to inflammation resolution and the start of bone repair/regeneration, unravelling candidate miRNAs to be used as biomarkers in the monitoring of healthy bone healing and as therapeutic targets for the development of improved bone regeneration therapies.

Project description:Heat stroke can induce a systemic inflammatory response, which may lead to multi‑organ dysfunction including acute kidney injury (AKI) and electrolyte disturbances. To investigate the pathogenesis of heat stroke (HS)‑related AKI, a mouse model of HS was induced by increasing the animal's core temperature to 41˚C. Blood samples obtained from the tail vein were used to measure plasma glucose and creatinine levels. Micro‑positron emission tomography‑computed tomography (micro‑PET/CT), H&E staining and transmission electron microscopy were conducted to examine metabolic and morphological changes in the mouse kidneys. Immunohistochemistry (IHC) and western blot analyses were performed to investigate the expression of apoptosis‑inducing factor mitochondria‑associated 2 (Aifm2), high‑mobility group box 1 (HMGB1) and receptor for advanced glycosylation end products (RAGE). Liquid chromatography‑mass spectrometry analysis was conducted to find differential metabolites and signaling pathways. The HS mouse model was built successfully, with significantly increased creatinine levels detected in the serum of HS mice compared with controls, whereas micro‑PET/CT revealed active metabolism in the whole body of HS mice. H&E and TUNEL staining revealed that the kidneys of HS mice exhibited signs of hemorrhage and apoptosis. IHC and western blotting demonstrated significant upregulation of Aifm2, HMGB1 and RAGE in response to HS. Finally, 136 differential metabolites were screened out, and enrichment of the 'biosynthesis of unsaturated fatty acids' pathway was detected. HS‑associated AKI is the renal manifestation of systemic inflammatory response syndrome, and may be triggered by the HMGB1/RAGE pathway. Metabolomics indicated increased adrenic acid, docosahexaenoic acid and eicosapentaenoic acid may serve as metabolic biomarkers for AKI in HS. The findings suggested that a correlation between the HMGB1/RAGE pathway and biosynthesis of unsaturated fatty acids may contribute to the progression of HS‑related AKI.

Project description:BackgroundIschemia/reperfusion (I/R) is the most common cause of acute kidney injury (AKI). Its pathophysiology remains unclear. Metabolomics is dedicated to identify metabolites involved in (patho)physiological changes of integrated living systems. Here, we performed 1H-Nuclear Magnetic Resonance metabolomics using urine, serum and kidney samples from a mouse model of renal I/R.MethodsRenal 30-min ischemia was induced in 12-week-old C57BL/6J male mice by bilaterally clamping vascular pedicles, and was followed by 6, 24 or 48-hour reperfusion (n = 12/group). Sham-operated mice were used as controls. Statistical discriminant analyses, i.e. principal component analysis and orthogonal projections to latent structures (OPLS-DA), were performed on urine, serum and kidney lysates at each time-point. Multivariate receiver operating characteristic (ROC) curves were drawn, and sensitivity and specificity were calculated from ROC confusion matrix (with averaged class probabilities across 100 cross-validations).ResultsUrine OPLS-DA analysis showed a net separation between I/R and sham groups, with significant variations in levels of taurine, di- and tri-methylamine, creatine and lactate. Such changes were observed as early as 6 hours post reperfusion. Major metabolome modifications occurred at 24h post reperfusion. At this time-point, correlation coefficients between urine spectra and conventional AKI biomarkers, i.e. serum creatinine and urea levels, reached 0.94 and 0.95, respectively. The area under ROC curve at 6h, 24h and 48h post surgery were 0.73, 0.98 and 0.97, respectively. Similar discriminations were found in kidney samples, with changes in levels of lactate, fatty acids, choline and taurine. By contrast, serum OPLS-DA analysis could not discriminate sham-operated from I/R-exposed animals.ConclusionsOur study demonstrates that renal I/R in mouse causes early and sustained metabolomic changes in urine and kidney composition. The most implicated pathways at 6h and 24h post reperfusion include gluconeogenesis, taurine and hypotaurine metabolism, whereas protein biosynthesis, glycolysis, and galactose and arginine metabolism are key at 48h post reperfusion.

Project description:Acute kidney injury (AKI) is a common cause of morbidity after congenital heart disease surgery. Progress on diagnosis and therapy remains limited, however, in part due to poor mechanistic understanding and a lack of relevant translational models. Metabolomic approaches could help identify novel mechanisms of injury and potential therapeutic targets. In the present study, we used a piglet model of cardiopulmonary bypass with deep hypothermic circulatory arrest (CPB/DHCA) and targeted metabolic profiling of kidney tissue, urine, and serum to evaluate metabolic changes specific to animals with histological acute kidney injury. CPB/DHCA animals with acute kidney injury were compared with those without acute kidney injury and mechanically ventilated controls. Acute kidney injury occurred in 10 of 20 CPB/DHCA animals 4 h after CPB/DHCA and 0 of 7 control animals. Injured kidneys showed a distinct tissue metabolic profile compared with uninjured kidneys (R2 = 0.93, Q2 = 0.53), with evidence of dysregulated tryptophan and purine metabolism. Nine urine metabolites differed significantly in animals with acute kidney injury with a pattern suggestive of increased aerobic glycolysis. Dysregulated metabolites in kidney tissue and urine did not overlap. CPB/DHCA strongly affected the serum metabolic profile, with only one metabolite that differed significantly with acute kidney injury (pyroglutamic acid, a marker of oxidative stress). In conclusion, based on these findings, kidney tryptophan and purine metabolism are candidates for further mechanistic and therapeutic investigation. Urine biomarkers of aerobic glycolysis could help diagnose early acute kidney injury after CPB/DHCA and warrant further evaluation. The serum metabolites measured at this early time point did not strongly differentiate based on acute kidney injury.NEW & NOTEWORTHY This project explored the metabolic underpinnings of postoperative acute kidney injury (AKI) following pediatric cardiac surgery in a translationally relevant large animal model of cardiopulmonary bypass with deep hypothermic circulatory arrest. Here, we present novel evidence for dysregulated tryptophan catabolism and purine catabolism in kidney tissue and increased urinary glycolysis intermediates in animals who developed histological AKI. These pathways represent potential diagnostic and therapeutic targets for postoperative AKI in this high-risk population.

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Human miRNome profiling identifies microRNAs differentially present in the urine after kidney injury.

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Human miRNome profiling identifies microRNAs differentially present in the urine after kidney injury.

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