Article

1H-NMR-based metabolic signatures of mild and severe ischemia/reperfusion injury in rat kidney transplants

March 2005
Kidney International 67(3):1142-51

March 2005
67(3):1142-51

DOI:10.1111/j.1523-1755.2005.00181.x

Source
PubMed

Authors:

Natalie J Serkova

University of Colorado

Jost Klawitter

University of Colorado

Show all 5 authorsHide

Severe ischemia/reperfusion (IR) injury is a risk factor for delayed graft function. Delayed graft function remains difficult to predict, and it currently relies primarily on serum creatinine (SCr), urine output, and occasionally on graft biopsy. (1)H-NMR (nuclear magnetic resonance spectroscopy) based metabolomics was used to establish IR-specific metabolic markers in both blood and kidney tissue. These markers were compared to SCr and graft histology. Male Lewis rats were used for kidney transplantation. Two cold ischemia (CI) groups (24- and 42-hour) and two transplantation groups [after 24 (TX24) and after 42 hours (TX42) of CI] were compared to a control group. Whole blood and kidney tissue were collected for further analysis. SCr levels taken 24 hours after transplantation were 1.6 +/- 0.12 mg/dL (TX24) and 2.1 +/- 0.5 mg/dL (TX42), (P= n.s.). Histology samples revealed mild injury in the TX24 group and severe injury in the TX42 group. A significantly decreased level of polyunsaturated fatty acids (PUFA) and elevated levels of allantoin, a marker of oxidative stress, was found in the renal tissue. In the blood, both trimethylamine-N-oxide (TMAO), a marker of renal medullary injury, and allantoin were significantly increased. Allantoin levels were low in both the control and CI groups. Levels were significantly increased after reperfusion (control 0.02 +/- 0.03 micromol/mL, TX24 1.13 +/- 0.22, and TX42 1.89 +/- 0.38, P < 0.001), and correlated with cold ischemia time (r= 0.96) and TMAO (r= 0.94). The (1)H-NMR metabolic profiles of both the mild and severe IR groups revealed significant changes consistent with graft histology, while the SCr did not.

Magnetic resonance spectroscopy of rat kidney in vivo at 9.4 T

Article

Full-text available

Jul 2019

Glutamate (Glu), glutamine (Gln), myo-insotol (Ins), taurine (Tau), choline (Cho), and betaine (Bet) are kidney metabolites whose levels are relevant to the study of Renal Cell Carcinoma (RCC). The presented work investigates the quantification of Glx (Glu + Gln), Ins, and Tau, relative to Cho + Bet with magnetic resonance spectroscopy (MRS) in rat kidney at 9.4 T. The Point-Resolved Spectroscopy (PRESS) pulse sequence was employed for non-invasive MRS acquisition in vivo from the right kidney of four rats. LCModel was utilized for peak fitting and spectral analysis of the in-vivo spectra. Relative metabolite concentrations and LCModel Cramér-Rao Lower Bounds (CRLBs) are reported. Relative metabolite concentrations are reported because the absolute concentration of any metabolite is unknown. The concentrations of Glx, Ins, and Tau, relative to Cho + Bet, the largest peak, were found to be, on average, 2.16, 1.40, and 2.17, with average CRLB values of 12.5%, 12.8%, and 16.8%, respectively. The average CRLB of Cho + Bet was 4.8%. To our knowledge, this is the first non-invasive MRS study of rat kidney.

Metabonomic profiling of chronic intermittent hypoxia in a mouse model

Article

Mar 2018
RESP PHYSIOL NEUROBI

Chronic intermittent hypoxia (ChIH) is a dominant feature of obstructive sleep apnoea (OSA) and is associated to metabolic alterations and oxidative stress (OS). Although management of OSA is well established, the research of new biomarkers that are independent of confounding factors remains necessary to improve the early detection of comorbidity and therapeutic follow-up. In this study, the urinary metabonomic profile associated to intermittent hypoxia was evaluated in a mouse model. When exposed to intermittent hypoxia, animals showed a significant alteration in energy metabolism towards anaerobic pathways and signs of OS imbalance. A compensatory response was observed over time. Our data also indicates an excess production of vitamin B3, liver function modulations and a stimulation of creatine synthesis which could be used to evaluate the ChIH repercussions. As well, TMAO and allantoin could constitute interesting biomarker candidates, respectively in the context of cardiovascular risk and OS associated to OSA.

The Role of Metabolomics in Current Concepts of Organ Preservation

Article

Full-text available

Sep 2020
INT J MOL SCI

In solid organ transplantation (Tx), both survival rates and quality of life have improved dramatically over the last few decades. Each year, the number of people on the wait list continues to increase, widening the gap between organ supply and demand. Therefore, the use of extended criteria donor grafts is growing, despite higher susceptibility to ischemia-reperfusion injury (IRI) and consecutive inferior Tx outcomes. Thus, tools to characterize organ quality prior to Tx are crucial components for Tx success. Innovative techniques of metabolic profiling revealed key pathways and mechanisms involved in IRI occurring during organ preservation. Although large-scale trials are needed, metabolomics appears to be a promising tool to characterize potential biomarkers, for the assessment of graft quality before Tx and evaluate graft-related outcomes. In this comprehensive review, we summarize the currently available literature on the use of metabolomics in solid organ Tx, with a special focus on metabolic profiling during graft preservation to assess organ quality prior to Tx.

The interplay between microbiota‐dependent metabolite trimethylamine N‐oxide, Transforming growth factor β/SMAD signaling and inflammasome activation in chronic kidney disease patients: A new mechanistic perspective

Article

Full-text available

Apr 2019

Background Chronic kidney disease (CKD) signifies a frequently life‐threatening condition influencing kidney structure and function. Despite its irrefutable importance, its exact pathogenesis is not completely clarified. However, CKD is known to be associated with accumulated uremic toxins/metabolites, interstitial fibrosis, and systemic inflammation. So we aimed to investigate the role of microbiota‐dependent metabolite trimethylamine N‐oxide (TMAO), transforming growth factor β (TGFβ)/SMAD signaling, and inflammasome activation in CKD pathogenesis through its different stages. Subjects and Methods Eighty patients with CKD of stages 2 to 4 in addition 15 healthy control subjects were enrolled. SMAD3 and nucleotide‐binding oligomerization domain‐, leucine‐rich repeat‐ and pyrin domain‐containing 3 (NLRP3) messenger RNA (mRNA) expressions from whole blood were assessed by quantitative real‐time polymerase chain reaction (RT‐PCR). Serum TGF‐β1 and interleukin‐1β (IL‐1β) levels were estimated by the enzyme‐linked immunosorbent assay. Plasma and urinary TMAO levels were measured. Oxidative stress markers were also assessed. Results SMAD3 and NLRP3 mRNA expressions were significantly upregulated in patients with CKD. Likewise, serum TGF‐β1 and IL‐1β levels were significantly elevated in patients with CKD, with increase in plasma and urinary TMAO levels and altered redox status throughout different CKD stages. Conclusion The study documented that TMAO could be used as a reliable biomarker to evaluate CKD progression; being linked to TGF‐β/SMAD signaling, NLRP3 inflammasome activation as well as being a noninvasive applicable technique.

Urinary metabolites predict prolonged duration of delayed graft function in DCD kidney transplant recipients

Article

Full-text available

May 2018

Extending kidney donor criteria, including donation after circulatory death (DCD), has resulted in increased rates of delayed graft function (DGF) and primary nonfunction. Here, we used Nuclear Magnetic Resonance (NMR) spectroscopy to analyze the urinary metabolome of DCD transplant recipients at multiple time points (days 10, 42, 180 and 360 after transplantation). The aim was to identify markers that predict prolonged duration of functional DGF (fDGF). Forty‐seven metabolites were quantified and their levels were evaluated in relation to fDGF. Samples obtained at day 10 had a different profile than samples obtained at the other time points. Furthermore, at day 10 there was a statistically significant increase in eight metabolites and a decrease in six metabolites in the group with fDGF (N=53) vis‐à‐vis the group without fDGF (N=22). In those with prolonged fDGF (≥21 days) (N=17) urine lactate was significantly higher and pyroglutamate lower than in those with limited fDGF (<21 days) (N=36). In order to further distinguish prolonged fDGF from limited fDGF, the ratios of all metabolites were analyzed. In a logistic regression analysis, the sum of branched‐chain amino acids (BCAAs) over pyroglutamate and lactate over fumarate, predicted prolonged fDGF with an AUC of 0.85. In conclusion, kidney transplant recipients with fDGF can be identified based on their altered urinary metabolome. Furthermore, two ratios of urinary metabolites, lactate/fumarate and BCAAs/pyroglutamate, adequately predict prolonged duration of fDGF. This article is protected by copyright. All rights reserved.

Correction: Elucidating time-dependent changes in the urinary metabolome of renal transplant patients by a combined 1 H NMR and GC-MS approach

Article

Full-text available

Jan 2017

Correction for 'Elucidating time-dependent changes in the urinary metabolome of renal transplant patients by a combined (1)H NMR and GC-MS approach' by Kienana Muhrez et al., Mol. BioSyst., 2015, 11, 2493-2510.

Potential applications of lipid peroxidation products - F4-neuroprostanes, F3-neuroprostanesn-6 DPA, and F2-dihomo-isoprostanes and F2-isoprostanes - in the evaluation of the allograft function in renal transplantation.

Article

Jan 2017
FREE RADICAL BIO MED

F4-neuroprostanes, F3-neuroprostanesn-6 DPA, and F2-dihomo-isoprostanes, metabolites of non-enzymatic lipid peroxidation of polyunsaturated fatty acids [docosahexaenoic acid, n-6 docosapentanoic acid, and adrenic acid respectively], have become important biomarkers for oxidative stress in several diseases like epilepsy and alzheimer. These biomarkers and the 15-F2t-isoprostane (also known as 8-iso-PGF2α), a F2-isoprostane isomer measured as reference oxidative marker at systemic level, were analyzed by UHPLC-QqQ-MS/MS in the urine of 60 renal recipients from cadaveric donors of the Nephrology Unit of the University Hospital Virgen de la Arrixaca, at six different times during the first six months after renal transplantation, and were compared with a control group of 60 healthy subjects from the same hospital. A total of 11 metabolites were analyzed and different patterns were observed. A tendency to decrease was observed in three metabolites (4-epi-4-F3t- NeuroPn-6 DPA, ent-7(RS)-7-F2t-dihomo-IsoP, and ent-7(S)-7-F2t-dihomo-IsoP) and in our reference oxidative marker (15-F2t-IsoP) when kidney function improved and the excretion of urine proteins decreased. These results suggest that these three biomarkers of oxidative stress could be useful to assess renal function in the postransplant phase. Unfortunately, little is known about this kind of biomarker in this cohort of patients, so further investigation would be required in the clinical field to clarify the relationship between oxidative stress and the graft function, as well as the usefulness of these biomarkers as rejection markers.

Nuclear Magnetic Resonance Metabolomic Profiling of Mouse Kidney, Urine and Serum Following Renal Ischemia/Reperfusion Injury

Article

Full-text available

Sep 2016
PLOS ONE

Background Ischemia/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. Methods Renal 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). Results Urine 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. Conclusions Our 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.

Metabolic Responses to the Yukon Arctic Ultra: Longest and Coldest in the World

Article

Full-text available

Sep 2016

Purpose The Yukon Arctic Ultra is considered the longest and coldest ultraendurance event in the world. Cold exposure and exercise has been reported to influence circulating levels of myokines, adipokines, and hepatokines that may influence considerable alterations in the regulation of metabolism. The purpose of the study was to evaluate the influence of the Yukon Arctic Ultra (430-mile event) on potential activators of brown fat, metabolites, and body composition in healthy individuals. Methods Eight male and female participants (mean ± SEM: age, 44 ± 3 yr; body mass index, 23.4 ± 0.9) were recruited for participation. Blood samples were collected at pre-event, mid-event, and post-event checkpoints. Results The temperature during the event ranged from −45°C to −8°C. Because of these extremely challenging conditions, 50% of the participants withdrew from competition by the 300-mile mark, and those that surpassed 300 miles lost a significant (P = 0.002; P = 0.01) amount of body weight (76 ± 5 kg to 73 ± 4 kg) and fat mass (13 ± 1 kg to 12 ± 3 kg), respectively. With respect to serum irisin, there was a trend (P = 0.06) toward significance from pre-event (1033 ± 88 ng·mL⁻¹), mid-event (1265 ± 23 ng·mL⁻¹) to post-event (1289 ± 24 ng·mL⁻¹). Serum meteorin-like and fibroblast growth factor-21 remained stable throughout the event. There were no changes in creatinine, acetoacetate, acetate, and valine. Serum lactate decreased (P = 0.04) during the event. Conclusions The influence of cold exposure and extreme physical exertion may promote substantial increases in serum irisin, and specific alterations in substrate metabolism that largely preserve skeletal muscle and physiological resilience.

Metabolites and lipoproteins may predict the severity of early Acute Pancreatitis in a South African cohort

Preprint

Full-text available

Nov 2023

Background: Acute pancreatitis (AP) is a common clinical disease with varying severity. The Revised Atlanta Classification (RAC) categorises AP into mild, moderately severe, and severe (MAP, MSAP and SAP) respectively. Despite the availability of different scoring systems to triage patients, these are not always suitable for predicting the course and outcome of certain patients during admission. In this study, untargeted metabolomics was used to identify metabolic parameters that can potentially be used as prognostic markers for stratifying the risk profile of patients for improved management and treatment. Methods: Serum isolated from blood samples collected from 30 AP patients (8 MAP, 14 MSAP, and 8 SAP) and 9 healthy control (HC) individuals was analysed using 1H nuclear magnetic resonance (NMR) spectroscopy. Wilcoxon and Kruskal Wallis rank-sum tests were used to compare differences in numerical covariates. A liposcale test was used for lipoprotein characterisation and the Spearman rank test was conducted for correlation of the data. P-values < 0.05 were considered significant. Results: Elevated levels of lactate, (rho = 0.67; p-value < 0.001, FDR = 0.001), 3-hydroxybutyrate (rho = 0.46; p-value < 0.003, FDR = 0.013), acetoacetate (rho = 0.63; p-value < 0.001, FDR < 0.001) and lipid alpha-CH2 (rho = 0.45; p-value = 0.004, FDR = 0.013) were associated with AP severity as was decreased levels of ascorbate (rho = 0.46; p-value < 0.003, FDR = 0.013), methanol (rho = 0.46; p-value < 0.003, FDR = 0.013), glutamine (rho = -0.55; p-value < 0.001, FDR = 0.002), ethanol (rho = 0.64; p-value < 0.001, FDR< 0.001), protein-NH (rho= -0.75; p-value < 0.001, FDR<0.001) among others. HDL-C decreased while IDL-C and VLDL-C increased across all the AP metabolic phenotypes compared to the HC. Conclusion: Dysregulated metabolites and lipids can potentially add to the understanding of the pathophysiological conditions of AP and can aid in the early prognosis and stratification of patients for specialist care.

Uncovering the relationship between gut microbial dysbiosis, metabolomics, and dietary intake in type 2 diabetes mellitus and in healthy volunteers: a multi-omics analysis

Article

Full-text available

Oct 2023

Type 2 Diabetes Mellitus has reached epidemic levels globally, and several studies have confirmed a link between gut microbial dysbiosis and aberrant glucose homeostasis among people with diabetes. While the assumption is that abnormal metabolomic signatures would often accompany microbial dysbiosis, the connection remains largely unknown. In this study, we investigated how diet changed the gut bacteriome, mycobiome and metabolome in people with and without type 2 Diabetes.1 Differential abundance testing determined that the metabolites Propionate, U8, and 2-Hydroxybutyrate were significantly lower, and 3-Hydroxyphenyl acetate was higher in the high fiber diet compared to low fiber diet in the healthy control group. Next, using multi-omics factor analysis (MOFA2), we attempted to uncover sources of variability that drive each of the different groups (bacterial, fungal, and metabolite) on all samples combined (control and DM II). Performing variance decomposition, ten latent factors were identified, and then each latent factor was tested for significant correlations with age, BMI, diet, and gender. Latent Factor1 was the most significantly correlated. Remarkably, the model revealed that the mycobiome explained most of the variance in the DM II group (12.5%) whereas bacteria explained most of the variance in the control group (64.2% vs. 10.4% in the DM II group). The latent Factor1 was significantly correlated with dietary intake (q < 0.01). Further analyses of the impact of bacterial and fungal genera on Factor1 determined that the nine bacterial genera (Phocaeicola, Ligilactobacillus, Mesosutterella, Acidaminococcus, Dorea A, CAG-317, Caecibacter, Prevotella and Gemmiger) and one fungal genus (Malassezia furfur) were found to have high factor weights (absolute weight > 0.6). Alternatively, a linear regression model was fitted per disease group for each genus to visualize the relationship between the factor values and feature abundances, showing Xylose with positive weights and Propionate, U8, and 2-Hydroxybutyrate with negative weights. This data provides new information on the microbially derived changes that influence metabolic phenotypes in response to different diets and disease conditions in humans.

Urinary Metabolomics Study on the Protective Role of Cocoa in Zucker Diabetic Rats via 1H-NMR-Based Approach

Article

Full-text available

Oct 2022

Cocoa constitutes one of the richest sources of dietary flavonoids with demonstrated anti-diabetic potential. However, the metabolic impact of cocoa intake in a diabetic context remains unexplored. In this study, metabolomics tools have been used to investigate the potential metabolic changes induced by cocoa in type 2 diabetes (T2D). To this end, male Zucker diabetic fatty rats were fed on standard (ZDF) or 10% cocoa-rich diet (ZDF-C) from week 10 to 20 of life. Cocoa supplementation clearly decreased serum glucose levels, improved glucose metabolism and produced significant changes in the urine metabolome of ZDF animals. Fourteen differential urinary metabolites were identified, with eight of them significantly modified by cocoa. An analysis of pathways revealed that butanoate metabolism and the synthesis and degradation of branched-chain amino acids and ketone bodies are involved in the beneficial impact of cocoa on diabetes. Moreover, correlation analysis indicated major associations between some of these urine metabolites (mainly valine, leucine, and isoleucine) and body weight, glycemia, insulin sensitivity, and glycated hemoglobin levels. Overall, this untargeted metabolomics approach provides a clear metabolic fingerprint associated to chronic cocoa intake that can be used as a marker for the improvement of glucose homeostasis in a diabetic context.

Serum Metabolomic and Lipoprotein Profiling of Pancreatic Ductal Adenocarcinoma Patients of African Ancestry

Article

Full-text available

Sep 2021

Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer with a characteristic dysregulated metabolism. Abnormal clinicopathological features linked to defective metabolic and inflammatory response pathways can induce PDAC development and progression. In this study, we investigated the metabolites and lipoproteins profiles of PDAC patients of African ancestry. Nuclear Magnetic Resonance (NMR) spectroscopy was conducted on serum obtained from consenting individuals (34 PDAC, 6 Chronic Pancreatitis, and 6 healthy participants). Seventy-five signals were quantified from each NMR spectrum. The Liposcale test was used for lipoprotein characterization. Spearman’s correlation and Kapan Meier tests were conducted for correlation and survival analyses, respectively. In our patient cohort, the results demonstrated that levels of metabolites involved in the glycolytic pathway increased with the tumour stage. Raised ethanol and 3-hydroxybutyrate were independently correlated with a shorter patient survival time, irrespective of tumour stage. Furthermore, increased levels of bilirubin resulted in an abnormal lipoprotein profile in PDAC patients. Additionally, we observed that the levels of a panel of metabolites (such as glucose and lactate) and lipoproteins correlated with those of inflammatory markers. Taken together, the metabolic phenotype can help distinguish PDAC severity and be used to predict patient survival and inform treatment intervention.

Metabolic and Lipoprotein Profiling of Pancreatic Ductal Adenocarcinoma Patients of African Ancestry

Preprint

Full-text available

Aug 2021

Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer with a characteristic dysregulated metabolism. Abnormal clinicopathological features linked to defective metabolic and inflammatory response pathways can induce PDAC development and progression. In this study, we investigated the metabolites and lipoproteins profiles of PDAC patients of African ancestry. Nuclear Magnetic Resonance (NMR) spectroscopy was conducted on serum obtained from consenting individuals (34 PDAC, 6 Chronic Pancreatitis, and 6 healthy participants). Seventy-five signals were quantified from each NMR spectrum. The Liposcale test was used for lipoprotein characterization. Spearman's correlation and Kapan Meier tests were conducted for correlation and survival analyses respectively. In our patient cohort, the results demonstrated that levels of metabolites involved in the glycolytic pathway increased with the tumour stage. Raised ethanol and 3-hydroxybutyrate were independently correlated with a shorter patient survival time, irrespective of tumour stage. Furthermore, increased levels of bilirubin resulted in an abnormal lipoprotein profile in PDAC patients. Additionally, we observed that the levels of a panel of metabolites (such as glucose, lactate) and lipoproteins correlated with those of inflammatory markers. Taken together, the metabolic phenotype can help distinguish PDAC severity and be used in predicting patient survival and in informing treatment intervention.

A UPLC-Q-TOF/MS-Based Metabolomics Study on the Effect of Corallodiscus flabellatus (Craib) B. L. Burtt Extract on Alzheimer’s Disease

Article

Full-text available

May 2021
EVID-BASED COMPL ALT

A UPLC-Q-TOF/MS-based metabolomics study was carried out to explore the intervening mechanism of Corallodiscus flabellatus (Craib) B. L. Burtt (CF) extract on Alzheimer’s disease (AD). The AD model group consisted of senescence-accelerated mouse prone 8 (SAMP8) mice, and the control group consisted of senescence-accelerated mouse resistant 1 (SAMR1) mice. UPLC-Q-TOF/MS detection, multivariate statistical analysis, and pathway enrichment were jointly performed to research the change in metabolite profiling in the urine of AD mice. The result suggested that the metabolite profiling of SAMP8 mice significantly changed at the sixth month compared with SAMR1 mice of the same age, and the principal component analysis (PCA) score scatter plots of the CF group closely resembled those of the control and positive drug (huperzine A, HA) group. A total of 28 metabolites were considered potential biomarkers associated with the metabolism of beta-alanine, glycine, serine, threonine, cysteine, methionine, arginine, proline, and purines in AD mice. Furthermore, the CF group was clustered with the control and positive group and was clearly separated from the model group in the heat map. In conclusion, significant anti-AD effects were firstly observed in mice after treatment with the CF extract, and the urinary metabolomics approach assisted with dissecting the underlying mechanism.

Studien zur Rolle von Trimethylamin-N-oxid in Lipid- und Glukosestoffwechsel

Thesis

Jan 2019

Astrid Ines Knell

Nuclear magnetic resonance (NMR)-based metabolome profile evaluation in dairy cows with and without displaced abomasum

Article

Full-text available

Jan 2020

Background: Displaced abomasum (DA) is a condition of dairy cows that severely impacts animal welfare and causes huge economic losses. Objective: To assess the metabolic status of the disease using metabolomics in serum, urine and liver samples aimed at both water soluble and lipid soluble fractions. Methods: Fifty Holstein multiparous cows with DA (42 left, 8 right) and 20 clinically healthy Holstein multiparous cows were used. Left DA was associated with concomitant ketosis in 19 animals and right in two. NMR-based metabolomics approach and hematological and biochemical analyses were performed. Statistical analysis was carried out on ¹HNMR data after they have been normalized using PQN method. Results: Contrary to generated PCA score plots the OPLS-supervised method revealed differences between healthy animals and diseased ones based on serum water-soluble samples. While water and lipid soluble metabolites decreased in serum samples, fatty acid fractions and cholesterol were increased in liver samples in DA affected cows. The metabolomic and chemical profiles clearly revealed that cows with DA (especially with LDA) were at risk of ketosis and fatty liver. Serum hippuric acid concentration was significantly higher in healthy cows in comparison with LDA, whereas serum glycine concentration was reported higher for healthy when compared to RDA affected animals. Conclusion: A biochemical network and pathway mapping revealed ‘valine, leucine and isoleucine biosynthesis’ and ‘phenylalanine, tyrosine and tryptophan biosynthesis’ as the most probable altered metabolic pathway in DA condition. Serum was advocated as the optimal biological matrix for the 1H-NMR analysis.

Tissue, urine and blood metabolite signatures of chronic kidney disease in the 5/6 nephrectomy rat model

Article

Full-text available

Aug 2019
METABOLOMICS

Introduction Progressive chronic kidney disease (CKD) is an important cause of morbidity and mortality. It has a long asymptomatic phase, where routine blood tests cannot identify early functional losses, and therefore identifying common mechanisms across the many etiologies is an important goal. Objectives Our aim was to characterize serum, urine and tissue (kidney, lung, heart, spleen and liver) metabolomics changes in a rat model of CKD. Methods A total of 17 male Wistar rats underwent 5/6 nephrectomy, whilst 13 rats underwent sham operation. Urine samples were collected weekly, for 6 weeks; blood was collected at weeks 0, 3 and 6; and tissue samples were collected at week 6. Samples were analyzed on a nuclear magnetic resonance spectroscopy platform with multivariate and univariate data analysis. Results Changes in several metabolites were statistically significant. Allantoin was affected in all compartments. Renal asparagine, creatine, hippurate and trimethylamine were significantly different; in other tissues creatine, dimethylamine, dimethylglycine, trigonelline and trimethylamine were significant. Benzoate, citrate, dimethylglycine, fumarate, guanidinoacetate, malate, myo-inositol and oxoglutarate were altered in urine or serum. Conclusion Although the metabolic picture is complex, we suggest oxidative stress, the gut-kidney axis, acid–base balance, and energy metabolism as promising areas for future investigation.

Flavin-containing monooxygenase 3 (FMO3): genetic variants and their consequences for drug metabolism and disease

Article

Jul 2019

1. The review focuses on genetic variants of human flavin-containing monooxygenase 3 (FMO3) and their impact on enzyme activity, drug metabolism and disease. 2. The majority of FMO-mediated metabolism in adult human liver is catalyzed by FMO3. Some drugs are metabolized in human liver predominantly by FMO3, but most drug substrates of FMO3 are metabolized also by other enzymes, particularly cytochromes P-450, and the FMO3-catalyzed reaction is not the major route of metabolism. 3. Rare variants that severely affect production or activity of FMO3 cause the disorder trimethylaminuria and impair metabolism of drug substrates of FMO3. More common variants, particularly p.[(Glu158Lys);(Glu308Gly)], can moderately affect activity of FMO3 in vitro and reduce metabolism of drug substrates in vivo, in some cases increasing drug efficacy or toxicity. 4. Common variants of FMO3 have been associated with a number of disorders, but additional studies are needed to confirm or refute such associations. 5. Elevated plasma concentrations of trimethylamine N-oxide, a product of an FMO3-catalyzed reaction, have been implicated in certain diseases, particularly cardiovascular disease. However, the evidence is often contradictory and additional work is required to establish whether trimethylamine N-oxide is a cause, effect or biomarker of the disease. 6. Genetic variants of other FMOs are also briefly discussed.

Metabolomics Approaches for the Diagnosis and Understanding of Kidney Diseases

Article

Full-text available

Feb 2019

Diseases of the kidney are difficult to diagnose and treat. This review summarises the definition, cause, epidemiology and treatment of some of these diseases including chronic kidney disease, diabetic nephropathy, acute kidney injury, kidney cancer, kidney transplantation and polycystic kidney diseases. Numerous studies have adopted a metabolomics approach to uncover new small molecule biomarkers of kidney diseases to improve specificity and sensitivity of diagnosis and to uncover biochemical mechanisms that may elucidate the cause and progression of these diseases. This work includes a description of mass spectrometry-based metabolomics approaches, including some of the currently available tools, and emphasises findings from metabolomics studies of kidney diseases. We have included a varied selection of studies (disease, model, sample number, analytical platform) and focused on metabolites which were commonly reported as discriminating features between kidney disease and a control. These metabolites are likely to be robust indicators of kidney disease processes, and therefore potential biomarkers, warranting further investigation.

Negative Energy Balance Does Not Alter Fat-Free Mass During the Yukon Arctic Ultra-The Longest and the Coldest Ultramarathon

Article

Full-text available

Dec 2018

Purpose: The objective of this study was to determine alterations in caloric balance, body composition, metabolites, and cytokines in athletes participating in the Yukon Arctic Ultra. Methods: Ten participants traveling on foot in the 2017 692-km event were recruited for the study. Measurements and samples were obtained at pre-event, 278 km (C1), 384 km (C2), and post-event. Body composition measurements were obtained using bioelectrical impedance analysis. Accelerometer devices were utilized to provide an estimation of caloric expenditure and dietary recalls provided assessments of caloric intake. Blood serum samples were collected, processed, and analyzed using enzyme-linked immunosorbent assays or nuclear magnetic resonance. Results were analyzed using linear mixed model, presented as means ± SD, and considered significant at p < 0.05. Results: Participants (8 males, 2 females; age: 37 ± 10 years; body mass index: 24.4 ± 2.5 kg/m²) were recruited. Four males and one female completed the entire event in 260 ± 19 h. Caloric intake/expenditure was 4,126 ± 1,115 kcal/day and 6,387 ± 781 kcal/day, respectively, indicating a caloric deficit of 2,261 ± 1,543 kcal/day. Total mass, body mass index, and fat mass were reduced at each time point of the event. Fat-free mass (FFM) was unchanged throughout the event. Follistatin was increased at C1 (1,715 ± 876 pg/ml) in comparison to baseline. Acetoacetate increased significantly at post-event (6.1 ± 1.5 mg/ml). Conclusions: Despite a pronounced caloric deficit and sustained activity under extreme cold conditions, FFM was preserved with an increase in serum follistatin and acetoacetate. Future studies should be directed at the role of nutrient strategies and/or training methods on the retention of FFM under these conditions.

Article

Full-text available

Feb 2018
METABOLOMICS

1 H NMR spectra from urine can yield information-rich data sets that offer important insights into many biological and biochemical phenomena. However, the quality and utility of these insights can be profoundly affected by how the NMR spectra are processed and interpreted. For instance, if the NMR spectra are incorrectly referenced or inconsistently aligned, the identification of many compounds will be incorrect. If the NMR spectra are mis-phased or if the baseline correction is flawed, the estimated concentrations of many compounds will be systematically biased. Furthermore, because NMR permits the measurement of concentrations spanning up to five orders of magnitude, several problems can arise with data analysis. For instance, signals originating from the most abundant metabolites may prove to be the least biologically relevant while signals arising from the least abundant metabolites may prove to be the most important but hardest to accurately and precisely measure. As a result, a number of data processing techniques such as scaling, transformation and normalization are often required to address these issues. Therefore, proper processing of NMR data is a critical step to correctly extract useful information in any NMR-based metabolomic study. In this review we highlight the significance, advantages and disadvantages of different NMR spectral processing steps that are common to most NMR-based metabolomic studies of urine. These include: chemical shift referencing, phase and baseline correction, spectral alignment, spectral binning, scaling and normalization. We also provide a set of recommendations for best practices regarding spectral and data processing for NMR-based metabolomic studies of biofluids, with a particular focus on urine.

Ischemia/reperfusion-associated tubular cells injury in renal transplantation: Can metabolomics inform about mechanisms and help identify new therapeutic targets?

Article

Jan 2018

Tubular cells are central targets of ischemia-reperfusion (I/R) injury in kidney transplantation. Inflammation and metabolic disturbances occurring within these cells are deleterious by themselves but also favor secondary events, such as activation of immune response. It is critical to have an in depth understanding of the mechanisms governing tubular cells response to I/R if one wants to define pertinent biomarkers or to elaborate targeted therapeutic interventions. As oxidative damage was shown to be central in the patho-physiological mechanisms, the impact of I/R on proximal tubular cells metabolism has been widely studied, contrary to its effects on expression and activity of membrane transporters of the proximal tubular cells. Yet, temporal modulation of transporters over ischemia and reperfusion periods appears to play a central role, not only in the induction of cells injury but also in graft function recovery. Metabolomics in cell models or diverse biofluids has the potential to provide large pictures of biochemical consequences of I/R. Metabolomic studies conducted in experimental models of I/R or in transplanted patients indeed retrieved metabolites belonging to the pathways known to be particularly affected. Interestingly, they also revealed that metabolic disturbances and transporters activities are in very close mutual interplay. As well as helping to select diagnostic biomarkers, such analyses could also contribute to identify new pharmacological targets and to set up innovative nephroprotective strategies for the future. Even if various therapeutic approaches have been evaluated for a long time to prevent or treat I/R injuries, metabolomics has helped identifying new ones, those related to membrane transporters seeming to be of particular interest. However, considering the very complex and multifactorial effects of I/R in the context of kidney transplantation, all tracks must be followed if one wants to prevent or limit its deleterious consequences.

Metabolic markers for differentiation between renal allograft rejection and immunosuppressant toxicity in rat urine

Article

Full-text available

Jul 2017
METABOLOMICS

IntroductionAlthough current immunosuppressive protocols have dramatically improved 1-year survival of kidney transplants, there has been less progress in terms of long-term graft survival over the last two decades. The key to avoiding late graft loss is early diagnosis and differentiation between anti-allograft immune processes and immunosuppressant toxicity (IS-Tox). Modern bioanalytical technologies have opened new opportunities for the development of sensitive and specific diagnostic tools. There is an immediate need for biomarkers that are able to differentiate between renal allograft rejection and immunosuppressant toxicity. Objective To test our hypothesis that changes of metabolite patterns in urine have the potential to serve as a non-invasive combinatorial biomarker that can differentiate between allograft immune reactions and IS-Tox. Methods We used 1H-NMR spectroscopy and Luminex multiplexing for metabolic profiling of rat urine and the analysis of protein biomarkers in urine and plasma, respectively, to compare the effects of chronic allograft rejection in a Fisher-to-Lewis rat transplant model with IS-Tox induced by cyclosporine, tacrolimus and/or sirolimus in Lewis rats. ResultsOur results showed that, while IS-Tox caused changes in metabolite patterns that are typically associated with proximal tubule damage, rejection caused more profuse changes not specifically focused on a particular kidney region. Moreover, metabolite pattern changes were more sensitive than changes in protein markers that were evident only during the later stages of rejection. Conclusion The present study provides first proof-of-concept that longitudinal monitoring of urine metabolite markers has the potential to differentiate between early renal allograft rejection and immunosuppressant nephrotoxicity.

Metabolomic profiling to characterize acute intestinal ischemia/reperfusion injury

Article

Full-text available

Jun 2017
PLOS ONE

Sepsis and septic shock are the leading causes of death in critically ill patients. Acute intestinal ischemia/reperfusion (AII/R) is an adaptive response to shock. The high mortality rate from AII/R is due to the severity of the disease and, more importantly, the failure of timely diagnosis. The objective of this investigation is to use nuclear magnetic resonance (NMR) analysis to characterize urine metabolomic profile of AII/R injury in a mouse model. Animals were exposed to sham, early (30 min) or late (60 min) acute intestinal ischemia by complete occlusion of the superior mesenteric artery, followed by 2 hrs of reperfusion. Urine was collected and analyzed by NMR spectroscopy. Urinary metabolite concentrations demonstrated that different profiles could be delineated based on the duration of the intestinal ischemia. Metabolites such as allantoin, creatinine, proline, and methylamine could be predictive of AII/R injury. Lactate, currently used for clinical diagnosis, was found not to significantly contribute to the classification model for either early or late ischemia. This study demonstrates that patterns of changes in urinary metabolites are effective at distinguishing AII/R progression in an animal model. This is a proof-of-concept study to further support examination of metabolites in the clinical diagnosis of intestinal ischemia reperfusion injury in patients. The discovery of a fingerprint metabolite profile of AII/R will be a major advancement in the diagnosis, treatment, and prevention of systemic injury in critically ill patients.

Nuclear magnetic resonance-based metabolomics of OCT-embedded frozen kidney samples in mouse and man following standardized pre-analytics

Article

Full-text available

Jun 2017
METABOLOMICS

Introduction Pre-analytical processing significantly affects tissue metabolomes. Since most frozen kidney samples are stored after embedding, standardization of cryoprotective medium removal before metabolomics is essential. Objectives We used rodent and human kidney samples to develop an easy and robust pre-analytical procedure compatible with ¹H-nuclear magnetic resonance (NMR)-based metabolomics. Methods In mice, renal ischemia was induced for 30 min, followed by 48-h reperfusion (I/R, n = 6). Right kidneys were transversally cut in two fragments, and snap-frozen in liquid nitrogen (LN2) or in Optimal Cutting Temperature ® (OCT) fixative. In man, double kidney biopsies were simultaneously obtained before transplantation (n = 15), and snap-frozen in LN2 or OCT. Results ¹H-NMR spectrum of pure OCT highlighted two major peaks, i.e. from 3.4 to 4.2 ppm (47.2%) and from 1.2 to 2.2 ppm (42.5%). ¹H-NMR spectra of mouse OCT kidneys were biased at 3.7. By contrast, ¹H-NMR analyses of mouse OCT kidneys iteratively rinsed in saline significantly discriminated sham versus I/R groups, with Q² at 0.695 (to be compared with Q² at 0.866 for LN2 sham vs. I/R kidneys). Discriminant metabolites were analogous in both OCT and LN2 kidneys, with a correlation coefficient of 0.83. In man, iteratively rinsing OCT kidneys in saline eliminated the spectral 3.7-peak, thereby making metabolomes of OCT kidneys interpretable and similar to LN2 samples, with a correlation coefficient of 0.73. Conclusion NMR metabolomics using OCT-frozen kidney samples is valuable in mouse and man, following standardized OCT removal. This may help use residual biobanked human tissues to better understand renal pathophysiology.

Liver Transplantation Biomarkers in the Metabolomics Era

Chapter

Jan 2017

The term biomarker usually refers to the biochemical molecules used in basic and clinical research, and also in the clinical practice, as surrogate markers that offer the advantage of being an objective, quantifiable, and reproducible measure. The most common applications of biomarkers include diagnosis, screening and monitoring of disease, assessment of response during therapy, risk assessment, and prognosis. Metabolomics or metabonomics enables the determination of hundreds of small molecules at the same time, which provides more comprehensive information than the determination of a single biomarker. Using metabolomics as an approach for searching biomarkers is supported by its capabilities to detect subtle metabolic changes triggered by external stimuli or perturbation. Metabolome changes are quite dynamic compared to genomics and transcriptomics, or even proteomics. Therefore, such metabolite alterations are found early in different samples, like tissues, cell lysates, blood, serum, plasma, feces, urine, etc. Application of metabolomics in liver transplantation is still in its early stages and has focused mainly on studying three aspects: post-reperfusion damage and rejection and dysfunction of the organ. In the current era when lack of organs suitable for transplantation is the most important limiting factor, the existence of an accepted functional assessment of grafts before transplantation would help to not only recover initially discarded organs but to also assess the therapies used to improve the quality of these organs. Different metabolic approaches have been used to search for objective markers of graft function and quality, but further analytical and clinical validation in multicentre studies is mandatory before they are incorporated into clinical routines.

Liver Transplantation Biomarkers in the Metabolomics Era

Chapter

Jan 2016

The term biomarker usually refers to the biochemical molecules used in basic and clinical research, and also in the clinical practice, as surrogate markers that offer the advantage of being an objective, quantifiable, and reproducible measure. The most common applications of biomarkers include diagnosis, screening and monitoring of disease, assessment of response during therapy, risk assessment, and prognosis. Metabolomics or metabonomics enables the determination of hundreds of small molecules at the same time, which provides more comprehensive information than the determination of a single biomarker. Using metabolomics as an approach for searching biomarkers is supported by its capabilities to detect subtle metabolic changes triggered by external stimuli or perturbation. Metabolome changes are quite dynamic compared to genomics and transcriptomics, or even proteomics. Therefore, such metabolite alterations are found early in different samples, like tissues, cell lysates, blood, serum, plasma, feces, urine, etc. Application of metabolomics in liver transplantation is still in its early stages and has focused mainly on studying three aspects: post-reperfusion damage, rejection and dysfunction of the organ. In the current era when lack of organs suitable for transplantation is the most important limiting factor, the existence of an accepted functional assessment of grafts before transplantation would help to not only recover initially discarded organs but to also assess the therapies used to improve the quality of these organs. Different metabolic approaches have been used to search for objective markers of graft function and quality, but further analytical and clinical validation in multicentre studies is mandatory before they are incorporated into clinical routines.

Metabolomic Profiling in Individuals with a Failing Kidney Allograft

Article

Full-text available

Jan 2017
PLOS ONE

Background Alteration of certain metabolites may play a role in the pathophysiology of renal allograft disease. Methods To explore metabolomic abnormalities in individuals with a failing kidney allograft, we analyzed by liquid chromatography-mass spectrometry (LC-MS/MS; for ex vivo profiling of serum and urine) and two dimensional correlated spectroscopy (2D COSY; for in vivo study of the kidney graft) 40 subjects with varying degrees of chronic allograft dysfunction stratified by tertiles of glomerular filtration rate (GFR; T1, T2, T3). Ten healthy non-allograft individuals were chosen as controls. Results LC-MS/MS analysis revealed a dose-response association between GFR and serum concentration of tryptophan, glutamine, dimethylarginine isomers (asymmetric [A]DMA and symmetric [S]DMA) and short-chain acylcarnitines (C4 and C12), (test for trend: T1-T3 = p<0.05; p = 0.01; p<0.001; p = 0.01; p = 0.01; p<0.05, respectively). The same association was found between GFR and urinary levels of histidine, DOPA, dopamine, carnosine, SDMA and ADMA (test for trend: T1-T3 = p<0.05; p<0.01; p = 0.001; p<0.05; p = 0.001; p<0.001; p<0.01, respectively). In vivo 2D COSY of the kidney allograft revealed significant reduction in the parenchymal content of choline, creatine, taurine and threonine (all: p<0.05) in individuals with lower GFR levels. Conclusions We report an association between renal function and altered metabolomic profile in renal transplant individuals with different degrees of kidney graft function.

Kidney Tissue Targeted Metabolic Profiling of Unilateral Ureteral Obstruction Rats by NMR

Article

Full-text available

Sep 2016

Renal interstitial fibrosis is a common pathological process in the progression of kidney disease. A nuclear magnetic resonance (NMR) based metabolomic approach was used to analyze the kidney tissues of rats with renal interstitial fibrosis (RIF), induced by unilateral ureteral obstruction (UUO). The combination of a variety of statistical methods were used to screen out 14 significantly changed potential metabolites, which are related with multiple biochemical processes including amino acid metabolism, adenine metabolism, energy metabolism, osmolyte change and induced oxidative stress. The exploration of the contralateral kidneys enhanced the understanding of the disease, which was also supported by serum biochemistry and kidney histopathology results. In addition, the pathological parameters (clinical chemistry, histological and immunohistochemistry results) were correlated with the significantly changed differential metabolites related with RIF. This study showed that targeted tissue metabolomic analysis can be used as a useful tool to understand the mechanism of the disease and provide a novel insight in the pathogenesis of RIF.

13C glucose labelling studies using 2D NMR are a useful tool for determining ex vivo whole organ metabolism during hypothermic machine perfusion of kidneys

Article

Full-text available

Dec 2016

Background The aim of this study is to determine the feasibility of using nuclear magnetic resonance (NMR) tracer studies (13C-enriched glucose) to detect ex vivo de novo metabolism in the perfusion fluid and cortical tissue of porcine kidneys during hypothermic machine perfusion (HMP). Methods Porcine kidneys (n = 6) were subjected to 24 h of HMP using the Organ Recovery Systems LifePort Kidney perfusion device. Glucose, uniformly enriched with the stable isotope 13C ([U-13C] glucose), was incorporated into KPS-1-like perfusion fluid at a concentration of 10 mM. Analysis of perfusate was performed using both 1D 1H and 2D 1H,13C heteronuclear single quantum coherence (HSQC) NMR spectroscopy. The metabolic activity was then studied by quantifying the proportion of key metabolites containing 13C in both perfusate and tissue samples. ResultsThere was significant enrichment of 13C in a number of central metabolites present in both the perfusate and tissue extracts and was most pronounced for lactate and alanine. The total amount of enriched lactate (per sample) in perfusion fluid increased during HMP (31.1 ± 12.2 nmol at 6 h vs 93.4 ± 25.6 nmol at 24 h p < 0.01). The total amount of enriched alanine increased in a similar fashion (1.73 ± 0.89 nmol at 6 h vs 6.80 ± 2.56 nmol at 24 h p < 0.05). In addition, small amounts of enriched acetate and glutamic acid were evident in some samples. Conclusions This study conclusively demonstrates that de novo metabolism occurs during HMP and highlights active metabolic pathways in this hypothermic, hypoxic environment. Whilst the majority of the 13C-enriched glucose is metabolised into glycolytic endpoint metabolites such as lactate, the presence of non-glycolytic pathway derivatives suggests that metabolism during HMP is more complex than previously thought. Isotopic labelled ex vivo organ perfusion studies using 2D NMR are feasible and informative.

Metabolic profiles in the response to supplementation with composite antimicrobial peptides in piglets challenged with deoxynivalenol

Article

Mar 2015

Deoxynivalenol (DON) causes various toxic effects in human and animals. However, our previous studies have shown that composite antimicrobial peptides (CAP) can have a protective effect in piglets challenged with DON. This study was conducted to evaluate the effect of the CAP GLAM 180# on the metabolism of piglets challenged with DON using a nuclear magnetic resonance (NMR)-based metabolomics approach. A total of 28 individually housed piglets (Duroc × Landrace × Large Yorkshire) weaned at 28 d of age were randomly assigned into 4 treatment groups (7 pigs/treatment) based on a 2 × 2 factorial arrangement that were fed, respectively, a basal diet (NC), basal diet + 0.4% CAP (basal + CAP), basal diet + 4 mg/kg DON (basal + DON), and basal diet + 4 mg/ kg DON + 0.4% CAP (DON + CAP). A 7-d adaptation period was followed by 30 d of treatment. Blood samples were then collected for metabolite analysis by proton NMR (1H-NMR) spectroscopy and liquid chromatography tandem mass spectrometry (LC-MS/MS). The combined results of1H-NMR spectroscopy and LC-MS/MS showed that DON increased (P < 0.05) the serum concentrations of low-density lipoprotein, glycoprotein, urea, trimethylamine-N-oxide (TMAO), and lactate as well as those of almost all essential AA and some nonessential AA but decreased the concentrations of high-density lipoprotein (HDL), unsaturated lipids, citrate, choline, and fumarate compared with those in NC treatment (P < 0.05). There was a significant interaction effect (P < 0.05) of supplementation with DON and CAP on some metabolites showed that the serum concentrations of HDL, unsaturated lipids, Pro, citrate, and fumarate were greater (P < 0.05) whereas those of glycoprotein, urea, TMAO, Gly, and lactate were lower in the DON + CAP treatment compared with those in the basal + DON treatment (P < 0.05). These findings indicated that DON causes disturbances in AA, lipid, and energy metabolism and that CAP could partially attenuate the above metabolic disturbances induced by DON. © 2015 American Society of Animal Science. All rights reserved.

Metabolomic characterization of renal ischemia and reperfusion in a swine model

Article

May 2016

Emerging approaches for the detection of trimethylamine N-oxide: A gut derived metabolite

Article

Nov 2023

Balancing the Equation: A Natural History of Trimethylamine and Trimethylamine- N -oxide

Article

Full-text available

Feb 2022

Trimethylamine (TMA) and its N-oxide (TMAO) are ubiquitous in prokaryote and eukaryote organisms as well as in the environment, reflecting their fundamental importance in evolutionary biology, and their diverse biochemical functions. Both metabolites have multiple biological roles including cell-signaling. Much attention has focused on the significance of serum and urinary TMAO in cardiovascular disease risk, yet this is only one of the many facets of a deeper TMA-TMAO partnership that reflects the significance of these metabolites in multiple biological processes spanning animals, plants, bacteria, and fungi. We report on analytical methods for measuring TMA and TMAO and attempt to critically synthesize and map the global functions of TMA and TMAO in a systems biology framework.

Preclinical models versus clinical renal ischemia reperfusion injury: A systematic review based on metabolic signatures

Article

Full-text available

Oct 2021

Despite decennia of research and numerous successful interventions in the preclinical setting, renal ischemia reperfusion (IR) injury remains a major problem in clinical practice, pointing towards a translational gap. Recently, two clinical studies on renal IR injury (manifested either as acute kidney injury or as delayed graft function), identified metabolic derailment as a key driver of renal IR injury. It was reasoned that these unambiguous metabolic findings enable direct alignment of clinical with preclinical data, thereby providing the opportunity to elaborate potential translational hurdles between preclinical research and the clinical context. A systematic review of studies that reported metabolic data in the context of renal IR was performed according to the PRISMA guidelines. The search (December 2020) identified thirty-five heterogeneous preclinical studies. The applied methodologies were compared, and metabolic outcomes were semi-quantified and aligned with the clinical data. This review identifies profound methodological challenges, such as the definition of IR injury, the follow-up time and sampling techniques, as well as shortcomings in the reported metabolic information. In light of these findings, recommendations are provided in order to improve the translatability of preclinical models of renal IR injury.

Intestinal microbiome and metabolome analyses reveal metabolic disorders in the early stage of renal transplantation

Article

Dec 2021

Renal transplantation is the most effective treatment for end-stage renal disease, but the long-term prognosis of organs after transplantation is not ideal. In recent years, the importance of gut microbes and metabolites in the study of disease mechanisms has gradually received attention. However, the coordination between gut microbes and the metabolism of renal transplant patients needs further study. We integrated 16s sequencing and metabolomics data to describe the changes in the serum and fecal metabolites of renal transplant patients. Our data revealed that the gut microbial diversity decreased and the relative abundance of many bacteria, such as Enterococcus and Streptococcus, significantly changed after transplantation. In addition, a large number of amino acids and peptides in serum and feces significantly changed, suggesting an abnormal amino acid metabolism after transplantation. Spearman's correlation analysis revealed the changes in the co-metabolism pattern between gut microbes and the host metabolism after transplantation. Furthermore, Enterococcus was found to be correlated with renal functions and metabolites reflecting renal damage. This study provides potential gut microbes and metabolites impacting renal health, which helps in understanding the renal damage in patients with kidney transplantation.

Monitoring of the influence of long‐term oxidative stress and ischemia on the condition of kidney using solid phase microextraction chemical biopsy coupled with liquid chromatography high resolution mass spectrometry

Article

Feb 2020

The limiting factor in conventional quality assessments of transplanted organs, namely the invasiveness of tissue sample collection, has prompted much research on the field of transplantology to focus on the development of alternative evaluation methods of organ quality. In the present project, we undertake the challenge to address the need for a new analytical solution for graft quality assessments by using a novel metabolomic diagnostic protocol based on low‐invasive solid‐phase microextraction. Solid‐phase microextraction probes of ca. 0.2 mm coated with 4 mm long mixed‐mode extraction phase were inserted into rabbit kidneys immediately following euthanasia and after 2, 4, 6, and 21 h of preservation. LC‐MS analysis of the extracts was performed with the use of a reversed phase column and a Q‐Exactive Focus mass spectrometer operated in positive ionization mode. Statistical analysis of significantly changing compounds revealed metabolic profile changes in kidneys induced by ischemia and oxidative stress as a function of the duration of cold storage. The most pronounced alterations were reflected in levels of essential amino acids and purine nucleosides. Our findings demonstrate that the proposed approach may be successfully used to monitor changes in the metabolic profile of organs over time of preservation. This article is protected by copyright. All rights reserved

A metabolomic study of the urine of rats with Alzheimer's disease and the efficacy of Ding‐Zhi‐Xiao‐Wan on the afflicted rats

Article

Feb 2020

As a well‐known traditional Chinese medicine formula, Ding‐Zhi‐Xiao‐Wan has long been used for the routine treatment of Alzheimer's disease. However, the mechanism of Ding‐Zhi‐Xiao‐Wan in treating Alzheimer's disease is unclear. Therefore, a non‐targeted metabolomics method based on ultra‐high performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry has been established to explore the metabolic variations in the urine of Alzheimer's disease rats and investigate the therapeutic mechanism of Ding‐Zhi‐Xiao‐Wan on Alzheimer's disease. To develop a better rat model of Alzheimer's disease, amyloid β25‐35 was injected into the bilateral hippocampus of Sprague–Dawley rats. Multivariate analysis approaches were applied to differentiate the urine components between the four groups. Thereafter, a targeted metabolomics method was used to verify the identified endogenous metabolites and determine the mechanism of action of Ding‐Zhi‐Xiao‐Wan. Altogether, 26 potential biomarkers were found, of which 15 biomarkers (10 of them are potential biomarkers found in non‐targeted metabolomics) were identified. The results show that Ding‐Zhi‐Xiao‐Wan mainly affects the pathways of taurine and hypotaurine metabolism, tryptophan metabolism and phenylalanine metabolism. Ding‐Zhi‐Xiao‐Wan might play a role in the treatment of Alzheimer's disease by mediating anti‐oxidative stress, regulation of energy metabolism, improvement of intestinal microbes, and protection of nerve cells. This article is protected by copyright. All rights reserved

Lipid Profiling Using 1H NMR Spectroscopy

Chapter

Aug 2019

Lipid profiling, which includes fatty acids, phospholipids, glycerides, and cholesterols is extremely important because of the essential role lipids play in the regulation of metabolism in animals. ¹H-NMR-based protocols for high-throughput lipid analysis in complex mixtures have been developed and applied to biological systems. Many classes of lipids can be quantitatively analyzed in many sample matrices including serum, cells, and tissues using a simple ¹H NMR experiment. In this chapter, we provide protocols for NMR-based lipid profiling including sample preparation, NMR experiments, and quantification using the LipSpin software tool.

Quantitative NMR-Based Metabolomics on Tissue Biomarkers and Its Translation into In Vivo Magnetic Resonance Spectroscopy

Chapter

May 2019

Nuclear magnetic resonance (NMR) spectroscopy is an established analytical platform for analyzing metabolic profiles of cells, tissues, and body fluids. There are several advantages in introducing an NMR-based study design into metabolomics studies, including a fast and comprehensive detection, characterization, and quantification of dozens of endogenous metabolites in a single NMR spectrum. Quantitative proton ¹H-NMR is the most useful NMR-based platform for metabolomics. The frozen tissues can be analyzed noninvasively using a high-resolution magic angle spinning (HR-MAS) ¹H-NMR spectroscopy; or several extraction techniques can be applied to detect additional metabolites using a conventional liquid-based NMR technique. In this chapter, we report on tissue collection, handling, extraction methods, and ¹H-NMR acquisition protocols developed in the past decades for a precise and quantitative NMR-metabolomics approach. The NMR acquisition protocols (both HR-MAS and conventional ¹H-NMR spectroscopy) and spectral analysis steps are also presented. Since NMR can be applied “in vivo” using horizontal bore MRI scanners, several in vivo sequences for localized ¹H-MRS (magnetic resonance spectroscopy) are presented which can be directly applied for noninvasive detection of brain metabolites.

Proton exchange in aqueous urea solutions measured by water‐exchange (WEX) NMR spectroscopy and chemical exchange saturation transfer (CEST) imaging in vitro

Article

Apr 2019

Purpose To characterize the proton exchange in aqueous urea solutions using a modified version of the WEX II filter at high magnetic field, and to assess the feasibility of performing quantitative urea CEST MRI on a 3T clinical MR system. Methods In order to study the dependence of the exchange‐rate constant of urea as a function of pH and T, the WEX‐spectra were acquired at 600 MHz from urea solutions in a pH range from 6.4 to 8.0 and a temperature range from to . The CEST experiments were performed on a 3T MRI scanner by applying a train of 50 Gaussian‐shaped pulses, each 100‐millisecond long with a spacing of 100 milliseconds, for saturation. Exchange rates of urea were calculated using the (extended) AREX metric. Results The results showed that proton exchange in aqueous urea solutions is acid and base catalyzed with the rate constants: l/(mol·s) and l/(mol·s), respectively. Since the urea protons undergo a slow exchange with water protons, the CEST effect of urea can be observed efficiently at 3T. However, in neutral solutions the exchange rate of urea is minimal and cannot be estimated using the quantitative CEST approach. Conclusions By means of the WEX‐spectroscopy, the kinetic parameters of the proton exchange in urea solutions have been determined. It was also possible to estimate the exchange rates of urea in a broad range of pH values using the CEST method at a clinical scanner.

Metabolomic Analysis Reveals a Protective Effect of Fu-Fang-Jin-Qian-Chao Herbal Granules on Oxalate-Induced Kidney Injury

Article

Full-text available

Feb 2019

Nephrolithiasis is one of the world's major public health burdens with a high incidence and a risk of persistent renal dysfunction. Fu-Fang-Jin-Qian-Chao granules (FFJQC), a traditional Chinese herb formula, is commonly used in treatment of nephrolithiasis. However, the therapeutic mechanism of FFJQC on kidney stone has still been a mystery. The objective of the present study is to explore the therapeutic mechanism of FFJQC on kidney injury and identify unique metabolomics patterns using a mouse model of kidney stone induced by a calcium oxalate (CaOx) deposition. Von Kossa staining and immuno-histopathological staining of osteopontin (OPN), cluster of differentiation 44 (CD44) and calbindin-D28k were conducted on renal sections. Biochemical analysis was performed on serum, urine, and kidney tissues. A metabolomics approach based on ultra-HPLC coupled with quadrupole-TOF-MS (UHPLC-Q-TOF/MS) was used for serum metabolic profiling. The immunohistopathological and biochemical analysis showed the therapeutic benefits of FFJQC. The expression levels of OPN and CD44 were decreased while calbindin-D28k increased after the CaOx injured mice were treated with FFJQC. In addition, total of 81 serum metabolites were identified to be associated with protective effects of FFJQC on CaOx crystal injured mice. Most of these metabolites were involved in purine, amino acid, membrane lipid and energy metabolism. Potential metabolite biomarkers were found for CaOx crystal-induced renal damage. Potential metabolite biomarkers of CaOx crystal-induced renal damage were found. FFJQC shows therapeutic benefits on CaOx crystal injured mice via regulation of multiple metabolic pathways including amino acids, purine, pyrimidine, glycerolipid, arachidonic acid (AA), sphingolipid, glycerophospholipid, and fatty acid.

Increased glutamine anabolism sensitizes non-small cell lung cancer to gefitinib treatment

Article

Full-text available

Dec 2018

To better understand the resistance mechanism of non-small cell lung cancers (NSCLCs) to gefitinib, the metabolic profiles of gefitinib-resistant A549 cells and gefitinib-sensitive PC-9 cells were analyzed with a metabolomics analytical platform. A549 and PC-9 cells exhibited significant differences in the levels of glutamine-related metabolites. After gefitinib treatment, the glutamine level decreased in A549 cells but showed no change in PC-9 cells. The glutamine consumed by A549 cells was used to generate ATP and glutathione (GSH). As glutamine utilization was suppressed in gefitinib-treated PC-9 cells, the resulting ATP shortage and ROS accumulation led to cell death. The difference in glutamine metabolism was caused by differential changes in the levels of glutamine synthetase (GS, encoded by glutamate-ammonia ligase (GLUL)). GLUL expression was upregulated in gefitinib-sensitive cells, but it was either absent from gefitinib-resistant cells or no significant change was observed in the gefitinib-treated cells. GLUL overexpression in A549 cells significant sensitized them to gefitinib and decreased their invasive capacity. Conversely, knockout GS in PC-9 cells reduced gefitinib sensitivity and enhanced metastasis. Furthermore, the continuous exposure of gefitinib-sensitive HCC827 cells to gefitinib created gefitinib-resistant (GR) HCC827 cells, which exhibited a GLUL deletion and resistance to gefitinib. Thus, GLUL plays a vital role in determining the sensitivity of NSCLCs to gefitinib. Elevated GS levels mediate increased glutamine anabolism, and this novel mechanism sensitizes NSCLCs to gefitinib. The inhibition of glutamine utilization may serve as a potential therapeutic strategy to overcome gefitinib resistance in the clinic.

Biochemical mechanisms of nephrotoxicity: Application for metabolomics

Article

Aug 2007

This review describes biochemical pathways of nephrotoxicity and the application of metabolic biomarkers as they relate to nephrotoxicity. Specific and sensitive biomarkers constitute the missing link in the continuum of exposure to toxins and susceptibility, disease development and possible therapeutic intervention. Important requirements for biomarker development are a detailed understanding of biochemical pathways involved in nephrotoxicity, minimal invasiveness and capacity to screen large at-risk populations. Lastly, possible biomarker candidates should be organ specific and equally applicable in preclinical drug testing as well as in clinical care of patients. This review discusses four major metabolic pathways associated with disturbed renal homeostasis: i) direct metabolic evidence of abnormal excretion of endogenous metabolites; ii) disturbances in kidney osmolarity and renal osmolyte homeostasis; iii) impaired energy state followed by dysregulation of glucose, fatty acid and ketone body metabolism; and iv) oxidative stress in renal tissues. Each of these pathways can be monitored by specific surrogate markers in urine and blood using modern metabolornics technologies.

Nuclear Magnetic Resonance Strategies for Metabolic Analysis

Chapter

Jan 2017
ADV EXP MED BIOL

NMR spectroscopy is a powerful tool for metabolomic studies, offering highly reproducible and quantitative analyses. This burgeoning field of NMR metabolomics has been greatly aided by the development of modern spectrometers and software, allowing high-throughput analysis with near real-time feedback. Whilst one-dimensional proton (1D-1H) NMR analysis is best described and remains most widely used, a plethora of alternative NMR techniques are now available that offer additional chemical and structural information and resolve many of the limitations of conventional 1D-1H NMR such as spectral overlay. In this book chapter, we review the principal concepts of practical NMR spectroscopy, from common sample preparation protocols to the benefits and theoretical concepts underpinning the commonly used pulse sequences. Finally, as a case study to highlight the utility of NMR as a method for metabolomic investigation, we have detailed how NMR has been used to gain valuable insight into the metabolism occurring in kidneys prior to transplantation and the potential implications of this.

Urinary Metabolomics for Noninvasive Detection of Antibody-Mediated Rejection in Children After Kidney Transplantation

Article

Jan 2017

Background: Biomarkers are needed that identify patients with antibody-mediated rejection (ABMR). The goal of this study was to evaluate the utility of urinary metabolomics for early noninvasive detection of ABMR in pediatric kidney transplant recipients. Methods: Urine samples (n=396) from a prospective, observational cohort of 59 renal transplant patients with surveillance or indication biopsies were assayed for 133 unique metabolites by quantitative mass spectrometry. Samples were classified according to Banff criteria for ABMR and partial least squares discriminant analysis was used to identify associated changes in metabolite patterns by creating a composite index based on all 133 metabolites. Results: Urine samples of patients with (n=40) and without ABMR (n=278) were analyzed and a classifier for ABMR was identified (AUC=0.84; 95% CI 0.77-0.91; p=0.006). Application of the classifier to "indeterminate" samples (samples that partially fulfilled Banff criteria for ABMR; n=65) yielded an ABMR score of 0.19±0.15, intermediate between scores for ABMR and no ABMR (0.28±0.14 and 0.10±0.13 respectively, p≤0.001). The ABMR score was associated with the presence of DSA, biopsy indication, Banff ct, t, ah and cg scores, and retained accuracy when applied to subclinical cases (creatinine <25% increase from baseline) or had minimal or no transplant glomerulopathy (Banff cg0-1). Exploratory classifiers that segregated samples based on concurrent T cell-mediated rejection (TCMR) identified overlapping metabolite signatures between ABMR and TCMR, suggesting similar pathophysiology of tissue injury. Conclusions: These preliminary findings identify a urine metabolic classifier for ABMR. Independent validation is needed to verify its utility for accurate, non-invasive ABMR detection.

The Role of Metabolomics in the Study of Kidney Diseases and in the Development of Diagnostic Tools

Chapter

Jan 2017

Metabolomics, or biochemical profiling, is closely integrated with genomics and proteomics. The metabonome is the complete collection of small molecule metabolites or chemicals (<1500 Da) that can be found at any given moment within a cell, organ, or organism. The metabonome is dynamic, constantly changing. Processes, such as diseases or drug effects, can affect cell biochemistry in a variety of ways. Alterations in the metabolome will precede the histopathological and pathophysiological changes that result from these altered states.Renal cells in particular are known to have a widely fluctuating process of metabolism, and renal disease in particular seems amenable to elucidation via metabolomics. Tubular cells and cells of the collecting ducts for instance, are known to function differently and display altered cell metabolism in response to factors, such as renal osmolarity and oxygen gradients. As the cell is constantly exchanging metabolites with the various fluid compartments of the body, changes in cell metabolism are often reflected by changes of the metabolite patterns in bodily fluids, such as plasma and urine. The analysis of metabolite patterns in urine and plasma is an attractive concept as these fluids are easily accessible, and could allow for the detection and monitoring of a disease or drug effect with high sensitivity and specificity. It is felt that with the more detailed information provided by metabolomics, the location of the injury also could be gleaned through analysis of the altered metabolic profile. Although there is no doubt that metabolomics has great potential to advance our understanding of kidney diseases and drug effects, to advance clinical diagnostics, and to be an important tool in the individualization of treatment, as of today the impact of metabolomics on clinical practice has been relatively small. Reasons include the complexity of developing molecular markers into diagnostic tools ranging from discovery, mechanistic, and clinical qualification to regulatory approval.

Changes in urinary metabolomic profile during relapsing renal vasculitis

Article

Full-text available

Dec 2016

Current biomarkers of renal disease in systemic vasculitis lack predictive value and are insensitive to early damage. To identify novel biomarkers of renal vasculitis flare, we analysed the longitudinal urinary metabolomic profile of a rat model of anti-neutrophil cytoplasmic antibody (ANCA) vasculitis. Wistar-Kyoto (WKY) rats were immunised with human myeloperoxidase (MPO). Urine was obtained at regular intervals for 181 days, after which relapse was induced by re-challenge with MPO. Urinary metabolites were assessed in an unbiased fashion using nuclear magnetic resonance (NMR) spectroscopy, and analysed using partial least squares discriminant analysis (PLS-DA) and partial least squares regression (PLS-R). At 56 days post-immunisation, we found that rats with vasculitis had a significantly different urinary metabolite profile than control animals; the observed PLS-DA clusters dissipated between 56 and 181 days, and re-emerged with relapse. The metabolites most altered in rats with active or relapsing vasculitis were trimethylamine N-oxide (TMAO), citrate and 2-oxoglutarate. Myo-inositol was also moderately predictive. The key urine metabolites identified in rats were confirmed in a large cohort of patients using liquid chromatography–mass spectrometry (LC-MS). Hypocitraturia and elevated urinary myo-inositol remained associated with active disease, with the urine myo-inositol:citrate ratio being tightly correlated with active renal vasculitis.

Trimethylamine-N-Oxide: Friend, Foe, or Simply Caught in the Cross-Fire?

Article

Nov 2016

Trimethylamine-N-oxide (TMAO), a gut-derived metabolite, has recently emerged as a candidate risk factor for cardiovascular disease and other adverse health outcomes. However, the relation between TMAO and chronic disease can be confounded by several factors, including kidney function, the gut microbiome, and flavin-containing monooxygenase 3 (FMO3) genotype. Thus, whether TMAO is a causative agent in human disease development and progression, or simply a marker of an underlying pathology, remains inconclusive. Importantly, dietary sources of TMAO have beneficial health effects and provide nutrients that have critical roles in many biological functions. Pre-emptive dietary strategies to restrict TMAO-generating nutrients as a means to improve human health warrant careful consideration and may not be justified at this time.

NMR spectroscopy as a novel approach to the monitoring of renal transplant function

Article

Full-text available

Jan 1993

High field 1H NMR spectroscopy was used for the rapid multicomponent analysis of low molecular wt compounds in urine in order to investigate the patterns of metabolic changes associated with early renal allograft dysfunction. Urine samples were collected daily for 14 days from 33 patients who underwent primary renal allograft transplantation, and analyzed by 500 and/or 600 MHz 1H NMR spectroscopy. All patients received 20 mg prednisolone and 5 mg/kg b.d. oral cyclosporin A (CsA) solution. In this study no patient showed clinical or histopathological evidence of CsA nephrotoxicity. For each patient the NMR-generated metabolite data were correlated with the clinical observations, graft biopsy pathology, and data from conventional laboratory techniques for assessing renal function. The NMR spectra of urine from patients with immediate functioning grafts were similar with respect to their patterns of amino acids, organic acids and organic amines, whereas the patients with delayed or non-functioning grafts showed significantly different metabolite excretion patterns. In longitudinal studies on individual patients there were increased urinary levels of trimethylamine-N-oxide (TMAO), dimethylamine (DMA), lactate, acetate, succinate, glycine and alanine during episodes of graft dysfunction. However, only the urinary concentration of TMAO was statistically significantly higher (P < 0.025) in the urine collected from patients during episodes of graft dysfunction (410 +/- 102 microM TMAO/mM creatinine) than in patients with good graft function (91 +/- 18 microM TMAO/mM creatinine) or healthy control subjects (100 +/- 50 microM TMAO/mM creatinine). These findings suggest that graft dysfunction is associated with damage to the renal medulla which causes the release of TMAO into the urine from the damaged renal medullary cells. This provides a possible novel urinary marker for post-transplant graft dysfunction. This study shows that NMR spectroscopy of biofluids, when used in combination with conventional laboratory techniques, is a valuable aid to renal transplant monitoring.

Donor age and delayed graft function as predictors of renal allograft survival in rejection-free patients

Article

Full-text available

Apr 1999

Transplant recipients of kidneys harvested from old donors have a high incidence of delayed graft function (DGF) and a poor graft outcome. This result is partly explained by the increased incidence of acute rejection in patients suffering from DGF. However, the long-term impact of donor age and DGF in rejection free renal transplants is not well established. The aim of the present work is to evaluate the impact of donor age and DGF on long-term outcome in renal transplants with or without acute rejection. We review all cadaveric kidney transplants performed in our centre between April 1984 and December 1995 treated with a cyclosporin-based immunosuppression. Five hundred and ninety-five patients were included. The overall incidence of DGF was 29.1%, and this event was associated with an increased donor age and cold ischaemia time. Univariate and multivariate analysis showed that graft loss was associated with acute rejection (relative risk (RR) 2.24, 95% confidence interval (CI) 1.62-3.01); DGF (RR 1.83, 95% CI 1.32-2.54); donors >50 years (RR 1.65, 95% CI 1.13-2.38); and retransplantation (RR 1.52, 95% CI 1.01-2.31). In rejection-free patients there were two independent predictors of graft failure: donor >50 years (RR 2.40, 95% CI 1.45-4.01); and DGF (RR 2.42, 95% CI 1.53-3.84). Regardless of the presence of acute rejection, delayed graft function amplifies the detrimental effect of advanced donor age on long-term graft outcome.

‘Metabonomics’: Understanding the Metabolic Responses of Living Systems to Pathophysiological Stimuli Via Multivariate Statistical Analysis of Biological Nmr Spectroscopic Data

Article

Full-text available

Dec 1999

Opinion: Understanding 'Global' Systems Biology: Metabonomics and the Continuum of Metabolism

Article

Full-text available

Sep 2003

To apply genomic knowledge effectively in drug discovery, mechanistic connectivities between genetic variation and disease processes need to be established via systems biology approaches. Humans have hundreds of functionally specialized cell types that interact differentially with environmental factors to influence disease development and to modulate the effects of drugs. Metabonomics can provide a means of modelling these interactions, but the relationships between 'endogenous' metabolic processes (coded in the genome and intrinsic to cellular function) and 'xenobiotic' (foreign compound) metabolism are poorly understood, especially with respect to environmental factors. We present an overview of 'global' mammalian metabolic conversions that should be accounted for in human systems biology models and propose a new probabilistic approach to help understand gene-disease relationships and vexed issues of idiosyncratic drug toxicity.

The effect of ferric-nitrilotriacetic acid on the profile of PUFA in the kidney and liver of rats

Article

Sep 2001

Intraperitoneal injection of the iron-complex, ferric-nitrilotriacetate (Fe-NTA), induces renal proximal tubular damage associated with oxidative damage in vivo. Fe-NTA induced a time-dependent decrease of several polyunsaturated fatty acids (PUFA), together with increased conjugated diene values and decreased cellular levels of α-tocopherol and glutathione. At the time of maximum detectable oxidation (3 h), after the injection of a sublethal dose of Fe-NTA there were clear reductions in the peak values over the controls for several fatty acids notably, 20:5 (eicosapentaenoic acid) (36%), 22:6 (docosahexanoic acid) (30%), 20:3 n6 (eicosatrienoic acid) (30%) and 20:4 (arachidonic acid) (28%) in the kidney. Fewer fatty acids showed a reduction in their residual values in the liver. 20:5 was reduced by 45% and for the 18:3 n3 and 18:3 n6, reductions of 35%, respectively. The profile of PUFAs is sensitive to the oxidative damage due to Fe-NTA and this may find applications as oxidative biomarker model.

Rapid and noninvasive diagnosis of the presence and severity of coronary heart disease using 1H-NMR-based metabonomics

Article

Nov 2002

Although a wide range of risk factors for coronary heart disease have been identified from population studies, these measures, singly or in combination, are insufficiently powerful to provide a reliable, noninvasive diagnosis of the presence of coronary heart disease. Here we show that pattern-recognition techniques applied to proton nuclear magnetic resonance (1H-NMR) spectra of human serum can correctly diagnose not only the presence, but also the severity, of coronary heart disease. Application of supervised partial least squares-discriminant analysis to orthogonal signal-corrected data sets allows >90% of subjects with stenosis of all three major coronary vessels to be distinguished from subjects with angiographically normal coronary arteries, with a specificity of >90%. Our studies show for the first time a technique capable of providing an accurate, noninvasive and rapid diagnosis of coronary heart disease that can be used clinically, either in population screening or to allow effective targeting of treatments such as statins.

Sirolimus, but not the structurally related RAD (everolimus), enhances the negative effects of cyclosporine on mitochondrial metabolism in the rat brain

Article

Jul 2001
BRIT J PHARMACOL

Clinical studies have shown enhancement of cyclosporine toxicity when co-administered with the immunosuppressant sirolimus. We evaluated the biochemical mechanisms underlying the sirolimus/cyclosporine interaction on rat brain metabolism using magnetic resonance spectroscopy (MRS) and compared the effects of sirolimus with those of the structurally related RAD. Two-week-old rats (25 g) were allocated to the following treatment groups (all n=6): I. control, II. cyclosporine (10 mg kg−1 d−1), III. sirolimus (3 mg kg−1 d−1), IV. RAD (3 mg kg−1 d−1), V. cyclosporine+sirolimus and VI. cyclosporine+RAD. Drugs were administered by oral gavage for 6 days. Twelve hours after the last dose, metabolic changes were assessed in brain tissue extracts using multinuclear MRS. Cyclosporine significantly inhibited mitochondrial glucose metabolism (glutamate: 78±6% of control; GABA: 67±12%; NAD+: 76±3%; P<0.05), but increased lactate production. Sirolimus and RAD inhibited cytosolic glucose metabolism via lactate production (sirolimus: 81±3% of control, RAD: 69±2%; P<0.02). Sirolimus enhanced cyclosporine-induced inhibition of mitochondrial glucose metabolism (glutamate: 60±4%; GABA: 59±8%; NAD+: 45±5%; P<0.02 versus cyclosporine alone). Lactate production was significantly reduced. In contrast, RAD antagonized the effects of cyclosporine (glutamate, GABA, and NAD+, not significantly different from controls). The results can partially be explained by pharmacokinetic interactions: co-administration increased the distribution of cyclosporine and sirolimus into brain tissue, while co-administration with RAD decreased cyclosporine brain tissue concentrations. In addition RAD, but not sirolimus, distributed into brain mitochondria. The combination of cyclosporine/RAD compares favourably to cyclosporine/sirolimus in regards to their effects on brain high-energy metabolism and tissue distribution in the rat. British Journal of Pharmacology (2001) 133, 875–885; doi:10.1038/sj.bjp.0704142

Simultaneous Measurement of Allantoin and Urate in Plasma: Analytical Evaluation and Potential Clinical Application in Oxidant:Antioxidant Balance Studies

Article

Jul 1999

Evaluation of renal medulla injury after cold preservation and transplantation: Noninvasive determination of medullar damage by proton nuclear magnetic resonance spectroscopy of urine and plasma

Article

Mar 2000
TRANSPL P

Is allantoin in serum and urine a useful indicator of exercise-induced oxidative stress in humans?

Article

Apr 2000

To assess whether allantoin levels in serum and urine are influenced by exhaustive and moderate exercise and whether allantoin is a useful indicator of exercise-induced oxidative stress in humans, we made subjects perform exhaustive and moderate (100% and 40% VO2max) cycling exercise and examined the levels of allantoin, thiobarbituric acid reactive substances (TBARS) and urate in serum and urine. Immediately after exercise at 100% VO2max, the serum allantoin/urate ratio was significantly elevated compared with the resting levels while the serum urate levels was significantly elevated 30 min after exercise. The serum TBARS levels did not increase significantly compared with the resting levels. Urinary allantoin excretion significantly increased during 60 min of recovery after exercise, however, urinary urate excretion decreased significantly during the same period. The urinary allantoin/urate ratio also rapidly increased during 60 min of recovery after exercise. Urinary TBARS excretion decreased during the first 60 min of the recovery period and thereafter significantly increased during the latter half of the recovery period. On the contrary, after 40% VO2max of exercise, no significant changes in the levels of urate, allantoin and TBARS in serum or urine were observed. These findings suggest that allantoin levels in serum and urine may reflect the extent of oxidative stress in vivo and that the allantoin which appeared following exercise may have originated not from urate formed as a result of exercise but from urate that previously existed in the body. Furthermore, these findings support the view that allantoin in serum and urine is a more sensitive and reliable indicator of in vivo oxidative stress than lipid peroxidation products measured as TBARS.

Influence of Retrieval Conditions on Renal Medulla Injury: Evaluation by Proton NMR Spectroscopy in an Isolated Perfused Pig Kidney Model

Article

Oct 2000

Delayed graft function (DGF) has remained an important complication after renal transplantation. The exact causes of DGF remain to be clarified, particularly the impact of retrieval conditions and preservation factors. In the present investigation, (1)HNMR spectroscopy of urine was assessed in order to detect the influence of retrieval condition on renal medulla damage. The isolated perfused pig kidney (IPK) was used to assess initial renal function from multiorgan donors (MOD) or single organ donors (SOD) after in situ cold flush and 24-h cold storage (CS) preservation with two standard preservation solutions: Euro-Collins (EC) and University of Wisconsin (UW) solutions. Kidneys flushed with cold heparinized saline and immediately perfused were used as the control group. Kidneys were perfused for 90 min at 37.5 degrees C for functional evaluation. During reperfusion, renal perfusion flow rate (PF) was measured. Glomerular filtration rate (GFR), tubular reabsorption of Na(+), and lactate dehydrogenase (LDH) and N-acetyl-beta-d-glucosaminidase (NAG) excretions were determined. Ischemia-reperfusion impairment was also determined by histological techniques and (1)HNMR spectroscopy. PF, GFR, and tubular reabsorption of Na(+) were significantly decreased in experimental groups when compared to the control group but there was no significant difference between experimental SOD groups. GFR was significantly greater in UW-MOD than in EC-MOD and tubular reabsorption of Na(+) was significantly greater in UW-MOD than in EC-MOD after 45 min of reperfusion. The release of LDH in the effluent and the urinary excretion of NAG were not significantly different after 24-h CS in the various experimental groups. The most relevant resonances determined by (1)HNMR spectroscopy were citrate, trimethylamine-N-oxide, lactate, acetate, and amino acids. Excretion of these markers was significantly different when compared to biochemical markers. A resonance (P) detected particularly in EC-MOD after 24-h CS was identified and well correlated to renal dysfunction. Histological study showed that ultrastructural damage and mitochondrial injury were more pronounced in the EC-MOD group. These results show that retrieval condition influences renal medullary damage. NMR spectroscopy, which is a noninvasive and nondestructive technique, is more efficient in assessing renal damage than conventional histology and biochemical analysis.

Kidney Retrieval Conditions Influence Damage to Renal Medulla: Evaluation by Proton Nuclear Magnetic Resonance (NMR) Spectroscopy

Article

Dec 2000

In the present investigation, the influence of retrieval condition on medullary damage in kidneys was assessed. The isolated perfused pig kidney was used to assess initial renal function from multiorgan donors or single organ donors after cold flush and 24 h cold storage preservation with two preservation solutions: Euro-Collins and University of Wisconsin solutions. Kidneys flushed with cold heparinized saline and immediately perfused were used as a control group. Kidneys were perfused for 90 min at 37.5 degrees C and renal perfusion flow rate, glomerular filtration rate, tubular reabsorption of Na+ and lactate dehydrogenase and N-acetyl-beta-D-glucosaminidase excretion were determined. Ischaemia reperfusion impairment was also determined by 1H NMR (proton nuclear magnetic resonance) spectroscopy. Renal function was significantly decreased in experimental groups when compared to the control group, but there was no significant difference between experimental groups after 24 h cold storage. The release of lactate dehydrogenase in the effluent and the urinary excretion of N-acetyl-beta-D-glucosaminidase were not significantly different after 24 h cold storage. The most relevant resonances determined by 1H NMR spectroscopy were citrate, trimethylamine-N-oxide, lactate, acetate and amino acids. Excretion of these markers was significantly different when compared to biochemical markers. A resonance P (Peak) detected particularly in Euro-Collins solution multiorgan donors after 24 h cold storage was identified and well correlated to renal dysfunction. N-acetyl-beta-D-glucosaminidase spectroscopy, which is a non-invasive and non-destructive technique, is more efficient to assess renal damage than conventional histology and biochemical analysis.

Delayed graft function influences renal function but not survival

Article

Aug 2000
TRANSPL P

Delayed graft function influences renal function, but not survival.Background In renal transplantation, the impact of delayed graft function (DGF) on prognosis is controversial. We analyzed the risk factors of DGF and its impact on graft function and prognosis.Methods Seven hundred thirty-four cadaveric renal transplants performed between 1983 and 1997 were analyzed. DGF was diagnosed when serum creatinine levels increased, remained unchanged, or decreased less than 10% per day in three consecutive days in the first week after transplantation. Creatinine clearances of more or less than 50 or 30 mL/min at one year were used as cut-off points for optimal and suboptimal graft function, respectively. The logistic regression model was used to identify independent risk factor related to DGF and renal function one year after transplantation. The Cox regression model was used to examine the influence of DGF on long-term graft survival.ResultsMultivariate analysis revealed the following risk factors for DGF: recipient pretransplantation mean arterial blood pressure of less than 100 mm Hg (OR = 2.08, 95% CI, 1.43 to 3.03), female donor to male recipient combination (OR = 1.55, 95% CI, 1.02 to 2.35), donor age of more than 50 years (OR = 2.21, 95% CI, 1.49 to 3.26), cold ischemia time of more than 28 hours (OR = 1.78, 95% CI, 1.19 to 2.63), and peak panel reactive antibodies of more than 50% (OR = 1.7, 95% CI, 1.15 to 2.55). The incidence of DGF was one of the independent risk factors for suboptimal graft function at one year (OR = 1.68, 95% CI, 1.14 to 2.48), together with donor age of more than 50 years (OR = 2.39, 95% CI, 1.61 to 3.57), female donor gender (OR = 1.99, 95% CI, 1.42 to 2.78), the occurrence of acute rejection episodes (OR = 2.66, 95% CI, 1.87 to 3.78), peak panel-reactive antibodies of more than 50% (OR = 1.67, 95% CI, 1.15 to 2.47), and sharing of 1 to 3 versus 4 to 8 cross-reactive antigens groups (OR = 1.65, 95% CI, 1.09 to 2.49). Moreover, DGF was one of the two independent risk factors for acute rejection episodes, but it had no independent effect on graft survival.Conclusion Several risk factors for DGF were identified, of which a low recipient pretransplant mean arterial blood pressure, the transplantation of kidneys from female donors to male recipients, and a prolonged cold ischemia time are potentially avoidable. Although DGF is one of the several risk factors of acute rejection and suboptimal function at one year, it is not independently associated with an increased rate of graft loss.

Oxidant stress and antioxidant capacity in urine of renal transplant recipients predict early graft function

Article

Feb 2001
TRANSPL P

Prolonged cold preservation augments vascular injury independent of renal transplant immunogenicity and function

Article

Sep 2001

While prolonged cold ischemia has detrimental effects on graft survival, the mechanisms remain unclear. We tested whether or not cold preservation enhances intragraft inflammatory responses and vascular injury. Rat renal grafts were cold preserved in University of Wisconsin solution for 2, 4, 6, 12, 24, and 48 hours, and then transplanted into syngeneic recipients and harvested after 24 hours. Frozen sections were examined histologically and stained for vascular cellular adhesion molecule-1 (VCAM-1), platelet-endothelial cell adhesion molecule-1 (PECAM-1), major histocompatibility complex (MHC) class II, tissue factor, leukocyte function associated molecule-1 (LFA-1), very late antigen-4 (VLA-4), as well as for inflammatory cells. Function did not differ between isografts preserved for shorter (2 to 6 hours) or longer times (12 to 24 hours). Neutrophil influx and that of LFA-1-positive cells showed similar increases in all groups. Compared with short preservation groups, the long preserved grafts had more VLA-4-positive ED-1+ monocytic infiltrates adjacent to vessels expressing VCAM-1 (P < or = 0.001). Increased preservation duration had no effect on infiltration with recipient ED-2+ macrophages, MHC class II-positive cells, or dendritic cells. Decreased color intensity and continuity of PECAM-1 staining indicated loss of endothelial integrity in grafts preserved for longer than six hours. Intensity in VCAM-1 staining increased progressively in grafts preserved for more than six hours and was localized predominantly on the endothelium of elastic vessels. Endothelial cells, vascular smooth muscle cells, and monocytes expressed increasingly more tissue factor in grafts preserved for more than six hours, revealing enhanced intragraft procoagulant capacity. Furthermore, grafts with preservation times of more than six hours developed more severe vascular endothelial injury and worse tubular necrosis scores (P < or = 0.001) compared with grafts with shorter preservation times. Because of the prominent vascular injury, strategies for endothelial protection should be attempted in grafts with long preservation times in clinical renal transplantation.

Long-term assessment of posttransplant renal prognosis with P-31 magnetic resonance spectroscopy

Article

Aug 2001

31P-magnetic resonance spectroscopy (MRS) has been widely used to study pretransplantation renal viability, and although some had discussed posttransplant renal viability, no one has examined long-term posttransplant renal prognosis. We discuss the use of 31P-MRS to assess the long-term prognosis from the time when MRS was performed. We studied 20 patients with renal allografts. 1.5 Tesla clinical magnetic resonance imaging (MRI) and 15 cm surface coil was used for 31P-MRS. Localized 31P-MRS was done using image selected in vivo spectroscopy (ISIS) method. Individual peaks were fitted by Lorenzian line-shapes with a least square method and peak area ratios were calculated. A beta-adenosine triphosphate/inorganic phosphate (beta-ATP/Pi) ratio >1.2 had sensitivity of 92.8%, specificity of 100%, and accuracy of 95% for predicting 3-year renal survival; a beta-ATP/Pi ratio >1.2 had sensitivity of 90.9%, specificity of 66.7%, and accuracy of 76.9% for predicting 5-year renal survival. We compared 31P-MRS spectra data between the survived group and failed group. The survived group had significantly higher beta-ATP/Pi, alpha-ATP/Pi, and phosphodiester (PDE)/Pi ratios than the failed group. We discussed the beta-ATP/Pi value as a parameter for predicting long-term survival of a transplanted kidney from the time when MRS was performed. A value above 1.2 suggests a high probability of 3-year renal survival, whereas a value over 2.5 indicates that the transplanted kidney could survive over 5 years. 31P-MRS may be useful for predicting long-term survival of transplanted kidneys, but additional studies are needed.

Risk factors for delayed graft function after renal transplantation and their significance for long- term clinical outcome

Article

Feb 2002

Delayed graft function (DGF) remains a grieving complication after renal transplantation. In this study, we examined various factors related to organ donation, transport, and transplantation for their influence on the incidence of DGF and on long-term prognosis. The incidence of DGF, renal function after 5 years, and allograft survival were analyzed in 200 kidneys transplanted in Düsseldorf as well as in 193 partner kidneys transplanted at 43 other centers. The main risk factors for DGF were donor age, cold ischemia time (CIT) and organ shipment. DGF itself, as well as donor age, influenced the long term prognosis. A significant relationship between the partner organs regarding clinical outcome was demonstrated. Non-immunological factors strongly influence the clinical results after renal transplantation. Organs of older donors have a limited long-term prognosis. To minimize additional risks, prevention of DGF, especially by reducing CIT, should be regarded as of paramount importance.

Measurement of free radicals and NO by chemiluminescence to identify the reperfusion injury in renal transplantation

Article

Mar 2002

Oxygen free radicals are generated during the reperfusion of ischaemic organs. Several experimental studies have demonstrated that the damage produced by reperfusion can be prevented by a scavenger of free radicals. Furthermore, a significantly improved 5 year graft survival rate after cadaveric renal transplantation has been reported in patients treated with scavengers of free radicals (Land et al., 1993). Therefore, a question remains to be answered: whether a routine monitoring of the radical-mediated reperfusion injury with renal transplantation is useful, and whether there is a necessity for a generalized protective treatment in transplant patients. In a prospective trial, we evaluated a group of eight patients during and after renal cadaveric transplantation (three men, five women), using the chemiluminometric measurement of serum free radicals and NO. The serum quantities of free radicals and NO were significantly increased after reperfusion of the transplant kidney (p < 0.02). The mean time of noticeably increased levels of serum free radicals was 4.8 +/- 1.2 h after reperfusion. The results thus showed an increased liberation of free radicals in the peripheral blood of transplant recipients as possible evidence of free radicals-mediated reperfusion injury in renal transplantation. The generation of free radicals measured by chemiluminescence allow a controlled therapy to decrease the generation of free radicals with antioxidants during the early transplantation period e.g. in older recipients.

The Inflammatory Aspect of the Microcirculation in Hypertension: Oxidative Stress, Leukocytes/Endothelial Interaction, Apoptosis

Article

Feb 2002

Evidence is increasing in hypertensive models for an inflammatory reaction in the microcirculation with abnormal leukocyte counts and adhesion to the endothelium, enhanced arteriolar tone, and microvascular and tissue apoptosis. The spontaneous form of hypertension (SHR) is accompanied by a glucocorticoid-dependent increase in circulating leukocyte count with elevated levels of activation and at the same time depressed leukocyte-endothelial interaction and endothelial P-selectin function. The SHR exhibits immune suppression with lymphocyte apoptosis in the thymus. Generation of reactive oxygen species (ROS) in and around microvascular endothelial cells may regulate signal transduction pathways responsible for controlling gene expression and protein modification and thereby cause an elevation of vascular tone and, in excess, may form an injury mechanism for cells and tissue. A series of enzyme systems such as xanthine oxidase, reduced nicotinamide adenine dinucleotide phosphate/reduced nicotinamide adenine dinucleotide oxidase, and cytochrome P450 monooxygenases in conjunction with suppression of ROS scavengers seem to be involved in the oxidative stress responses in hypertension. The increase in ROS generation contributes to vascular remodeling, apoptosis, and proliferation of vascular smooth muscle, whereas gaseous monoxides such as nitric oxide and carbon monoxide have the ability to modulate elevated vascular tone and proliferative cell responses. Such biological actions of gases not only regulate activation of soluble guanylate cyclase but could also be attributable to inhibition of cytochrome P450 monooxygenases. We examine here the molecular basis of signal transduction by ROS, NO, and CO and functional alterations in their sensor molecules. An inflammatory reaction may underlie the pathogenesis of hypertension and its associated lesion formation and organ dysfunction.

Antioxidant enzyme determination and a study of lipid peroxidation in renal transplantation

Article

Jun 2002

The aim of this study was to evaluate the antioxidant enzymes, catalase (CAT) and superoxide dismutase (SOD), in renal transplant patients and at the same time to report on the degree of lipid peroxidation observed in these patients. In order to do so we measured erythrocyte malondialdehyde (MDA), a product of lipid peroxidation. These measurements were made at different times: before the transplant, 48 h after the transplant, a week after the transplant and two weeks after the transplant. The values reported were compared with a control group. The results showed that there was a higher MDA level in the transplant group than in the control group one week after the transplant. In addition, two weeks after the transplant, the activities of CAT and SOD were higher in the transplant group than in the control group.

Close Association Between the Reduction in Myocardial Energy Metabolism and Infarct Size: Dose-Response Assessment of Cyclosporine

Article

Sep 2002

Cyclosporine protects the heart against ischemia/reperfusion injury, but its effect on cardiac metabolism is largely unknown. We assessed cyclosporine-induced metabolic changes in the rat heart prior to occlusion using magnetic resonance spectroscopy (MRS) and correlated effects with infarct size in a coronary occlusion/reperfusion model. The two study groups were cyclosporine and cyclosporine + coronary occlusion (n = 20/group). Rats were pretreated with cyclosporine (5, 10, 15, and 25 mg/kg/day) or the vehicle by oral gavage for 3 days (n = 4/dose). On day 4, hearts of rats in the cyclosporine group were excised, and extracted cell metabolites were measured using (1)H and (31)P MRS. The second group was subjected to 30 min of coronary artery occlusion followed by 24 h of reperfusion. Infarct size and area at risk were measured using a double staining method. In the cyclosporine group, cyclosporine reduced cardiac energy metabolism (ATP: r = -0.89, P < 0.001) via depression of oxidative phosphorylation and the Krebs' cycle in a dose-dependent manner. The decrease of ATP levels was positively correlated with changes of NAD(+) (r = 0.89), glutamate (r = 0.95), glutamine (r = 0.84), and glucose concentrations (r = 0.92, all P < 0.002). It was inversely correlated with lactate (r = -0.93, P < 0.001). In the coronary occlusion group, cyclosporine dose dependently reduced the ratio [area of infarct/area of the left ventricle] (r = -0.86, P < 0.01), with 15 mg/kg/day being the most effective cyclosporine dose. The reduction in infarct size correlated with the reduction in oxidative phosphorylation (ATP: r = 0.97; NAD(+): r = 0.82, P < 0.01). The reduction in cardiac energy metabolism before occlusion may be the cause of myocardial preservation during ischemia/reperfusion.

NMR-Based Metabonomic Studies on the Biochemical Effects of Commonly Used Drug Carrier Vehicles in the Rat

Article

Oct 2002

The biochemical effects of a series of commonly used drug carrier vehicles were investigated using (1)H NMR spectroscopic and pattern recognition based metabonomic analysis. Animals were treated by oral gavage with six dosage vehicles: 0.5% (w/v) sodium carboxymethylcellulose/0.2% (v/v)tween; microemulsion (consisting of propylene glycol, ethanol, cremophor, and corn oil glycerides); labrafil [consisting of poly(ethylene glycol) 300 esterified with oleic acid] (30%)/corn oil (70%); 0.1 M sodium phosphate buffered water; poly(ethylene glycol) 300 and 0.5% methocel. Urine samples (n = 7) collected over a 96 h period post administration were analyzed using 600 MHz (1)H NMR spectroscopy, and principal components analysis of the spectral data was used to analyze these data. Of the six vehicles studied, three (labrafil/corn oil, PEG 300 and microemulsion) gave rise to strong vehicle-related signals in the (1)H NMR spectra of urine and were, therefore, deemed to be less suitable for NMR-based toxicity studies. To investigate any biochemical consequences of vehicle dosing, PCA was used to analyze spectral regions that did not contain vehicle-related signals, i.e., the NMR-detectable endogenous metabolite profile. PEG 300 and labrafil/corn oil induced changes in the biochemical composition of urine including increased concentrations of dicarboxylic acids, creatinine, taurine, and sugars, indicating that these vehicles were bioactive in their own right and that this might confound interpretation of biochemical effects of weakly toxic drugs dosed in these carriers. This study shows the importance of selecting appropriate vehicles for NMR-based metabonomic studies with a view to minimizing the possibility of vehicle resonances obscuring endogenous compound peaks. Furthermore, we have shown that at least two of the commonly used drug carrier vehicles caused metabolic perturbations in the urine profile. These alterations in the biochemical profile reflect vehicle-induced changes in the physiological status of the organism that may obscure the pharmacologic or toxicologic effects of drugs.

Predicting the outcome of prostate biopsy in a racially diverse population: A prospective study

Article

Nov 2002

To develop a mathematical model to predict prostate biopsy outcome using readily available clinical variables. A total of 319 men (78% African American) undergoing transrectal ultrasound-guided prostate biopsy were prospectively studied. The parameters collected included age, race, prostate-specific antigen (PSA) level, PSA density (PSAD), digital rectal examination findings, biopsy history, prostate volume (by transrectal ultrasound), and ultrasound findings. Models were constructed using multivariate logistic regression (LR) analysis and back-propagation artificial neural networks (ANNs). Patient data were randomly split into five cross-validation sets and used to develop and validate the LR and ANN models. Of the 319 men, 39% had a positive biopsy. The mean patient age was 65.1 +/- 8.3 years, with a mean PSA level of 12.6 +/- 24.9 ng/mL and a mean PSAD of 0.31 +/- 0.66 ng/mL/cm(3). Univariate analysis indicated a significant difference in age, PSA level, PSAD, free PSA, digital rectal examination findings, TRUS lesion, and biopsy history between the positive and negative biopsy groups (P <0.01). The mean area under the receiver operating characteristic curve (AUROC) for the five LR models was 0.76 +/- 0.04 (range 0.71 to 0.81). The median LR AUROC was 0.76, with a corresponding specificity of 0.13 at a sensitivity of 0.95. The mean AUROC for the five ANN models was 0.76 +/- 0.04 (range 0.71 to 0.83). The median ANN AUROC was 0.76, with a corresponding specificity of 0.21 at a sensitivity of 0.95. Two models (LR and ANN) that predict outcome with high efficiency (AUROC = 0.76) were constructed from a contemporary, prospective database. Such models may be useful to patients and physicians alike when assessing the diagnostic strategies available to detect prostate cancer.

Rapid and Noninvasive Diagnosis of the Presence and Severity of Coronary Heart Disease Using 1H-NMR-Based Metabonomics

Article

Dec 2002

Prognostic significance of free radicals: Mediated injury occurring in the kidney donor

Article

Apr 2003

Brain death is associated with hemodynamic disturbances in systemic circulation and metabolic storm, and, thus, free radical-mediated injury to donor tissues was hypothesized. An assessment of oxidative stress in the donor and its effect on posttransplant kidney graft function comprised the scope of the study. A prospective study was performed in 27 donors and 50 kidney transplant recipients. Sera from 27 brain-dead organ donors and preservation media were tested for malondialdehyde (MDA) and for total antioxidant status (TAS). Kidneys were preserved in University of Wisconsin-gluconate solution with machine perfusion. Mean ischemia time was 36.7+/-8 hours. Organs were transplanted to recipients on the Polish National Waiting List and posttransplant kidney function was monitored periodically. Posttransplant delayed graft function (DF) was diagnosed when a patient required at least one dialysis within first week after transplantation. Acute rejection was diagnosed clinically and confirmed with fine-needle biopsy if necessary. Thirty-two recipients had immediate graft function (IF), and 18 suffered from DF. MDA level in preservation solution at the end of machine perfusion was significantly higher in the DF group (52.6+/-31 vs. 25.3+/-19 micromol/L) whereas donor TAS activity was lower (1.14+/-0.2 vs. 0.97+/-0.3 mmol/mL). Patients who suffered from acute rejection received kidneys from donors with significantly higher serum MDA (66+/-73 micromol/ml vs. 23+/-49 for patients without rejection). Serum creatinine 12 to 48 months after transplantation correlated to donor- and preservation-solution MDA (P<0.006). Free-radical mediated injury occurring in the donor and during preservation is strictly correlated with immediate and long-term kidney function. It may also cause grafts to be prone to acute rejection.

Organ-specific response to inhibition of mitochondrial metabolism by cyclosporine in the rat

Article

Oct 2003

To evaluate organ-specific metabolic changes after in vivo cyclosporine (CyA) treatment, male Wistar rats were treated with 10 mg/kg per day CyA orally for 6 days. Blood, kidney, liver, and heart tissues were extracted and analyzed by magnetic resonance spectroscopy (MRS). CyA decreased the energy balance [adenosine triphosphate (ATP)/adenosine diphosphate (ADP)] in all organs (kidney [control]: 50%, liver: 64%, and heart: 62%, all P<0.01) due to decreased activity of the mitochondrial Krebs cycle and oxidative phosphorylation. As a compensatory effect, anaerobic glycolysis (lactate) was increased. This was reflected in the low glucose level in the kidney and heart, but not in the liver where a significant decrease in glycogen was seen. Only in the kidney was mitochondrial inhibition accompanied by decreased polyunsaturated fatty acid (PUFA) concentrations and elevated lipid peroxidation. The metabolic marker for nephrotoxicity, trimethylamine-N-oxide (TMAO), was elevated. While CyA decreased mitochondrial homeostasis in all organ systems, cellular adaptation was different and most efficient in the liver.

Sirolimus delays recovery of rat kidney transplants after ischemia-reperfusion injury

Article

Dec 2003

Sirolimus (SRL) seems to impair renal recovery from ischemic injury in animal models and delayed graft function after clinical renal transplantation. This study determined the impact of SRL on renal recovery after ischemia-reperfusion injury in a rat kidney transplant model. Syngeneic kidneys were preserved in University of Wisconsin solution before transplantation into bilaterally nephrectomized rats. Recipients received vehicle or SRL targeting for whole-blood trough levels of 10 to 20 ng/mL as measured by high-performance liquid chromatography. Renal function was assessed by animal survival or daily serum creatinine. Tissue samples were collected for histologic examination. Median SRL whole-blood concentrations were 16.6 +/- 1.6 ng/mL on postoperative day (POD) 1 and 12.0 +/- 0.9 ng/mL on POD 3. Transplantation of kidneys after 39 hr of cold storage resulted in 30% survival in the SRL-treated group compared with 100% survival in the control group (P=0.002). Transplantation of kidneys after 24 hr of cold storage resulted in no survival differences, but there were significant differences in renal function. Daily serum creatinine (PODs 1-4) was higher in the SRL-treated group compared with the control group (P<0.05 at all time points). Grafts from SRL-treated animals showed more severe tubular necrosis compared with control animals. When given at therapeutic immunosuppressive doses, SRL compromises renal function after ischemia-reperfusion injury in a rat kidney transplant model. The antiproliferative effect of SRL may translate into impairment of tubular repair and regeneration necessary for recovery after such injury.

Influence of Donor Pretreatment With N-Acetylcysteine on Ischemia/Reperfusion Injury in Rat Kidney Grafts

Article

Apr 2004

N-acetylcysteine (NAC) has been shown to ameliorate ischemic acute renal failure. We determined the effect of donor pretreatment with NAC on ischemia reperfusion (I/R) injury in rat kidney grafts. Lewis rats were divided into 3 groups (8 per group) and treated with saline, mannitol (1 gm/kg) or NAC (1 gm/kg intravenously) prior to donor nephrectomy. Cold stored kidneys (24 hours in UW solution) were transplanted into bilaterally nephrectomized recipients. Blood and graft tissue samples were taken 24 hours after transplantation for assessment of metabolic changes, histological damage and renal function. Metabolites associated with renal I/R injury were quantified in blood and renal tissue by magnetic resonance spectroscopy. The degree of histological damage was similar between the treatment groups. Of the counted tubules 60%were mildly damaged, whereas 40% showed moderate damage. Measurement of the metabolites allantoin and trimethylamine-N-oxide (TMAO) indicated a beneficial effect of NAC treatment. In graft tissue and recipient blood allantoin, a uric acid metabolite, was significantly lower in the NAC group vs the mannitol and saline groups (p <0.05). In recipient blood TMAO, an established marker of renal medullary injury, was significantly decreased in the NAC group vs mannitol and saline (p <0.05). Serum creatinine levels were not different between treatment groups. Donor pretreatment with NAC preserves renal metabolism and may improve outcomes of I/R injured kidney transplants. Allantoin and TMAO are sensitive metabolic markers of renal I/R injury that can be detected before the onset of functional and morphological changes.

Predicting the out-come of prostate biopsy in a racially diverse population: A prospec-tive study NMR-based metabonomic studies on the biochemical effects of commonly used drug carrier vehicles in the rat

Jan 2002
831-8351136

Crawford Ed

PORTER C, O'DONNELL C, CRAWFORD ED, et al: Predicting the out-come of prostate biopsy in a racially diverse population: A prospec-tive study. Urology 60:831–835, 2002 17. BECKWITH-HALL B, HOLMES E, LINDON JC, et al: NMR-based metabonomic studies on the biochemical effects of commonly used drug carrier vehicles in the rat. Chem Res Toxicol 15:1136–1141, 2002

Opinion: Understanding 'global' systems biology: Metabonomics and the continuum of metabolism Rapid and noninvasive diagnosis of the presence and severity of coronary heart disease using 1H-NMR-based metabonomics

Jan 2002
668-6761439

Wilson Id
Antti H Brinde J

NICHOLSON J, WILSON ID: Opinion: Understanding 'global' systems biology: Metabonomics and the continuum of metabolism. Nat Rev Drug Discov 2:668–676, 2003 10. BRINDE J, ANTTI H, HOLMES E, et al: Rapid and noninvasive diagnosis of the presence and severity of coronary heart disease using 1H-NMR-based metabonomics. Nature Med 8:1439–1444, 2002

Delayed graft function influences renal function, but not survival

Jan 2000
KIDNEY INT
859-866

Mallat Boom H
Mj
De Fijter
Jw

BOOM H, MALLAT MJ, DE FIJTER JW, et al: Delayed graft function influences renal function, but not survival. Kidney Int 58:859-866, 2000

Evaluation of renal medulla injury after cold preservation and transplantation: noninvasive determination of medullar damage by proton nuclear magnetic resonance spectroscopy of urine and plasma

Jan 2000
47-48

Baumer Richer J
Th
Gibelin H

RICHER J, BAUMER TH, GIBELIN H, et al: Evaluation of renal medulla injury after cold preservation and transplantation: noninvasive determination of medullar damage by proton nuclear magnetic resonance spectroscopy of urine and plasma. Transplant Proc 32:47-48, 2000

Rapid and noninvasive diagnosis of the presence and severity of coronary heart disease using 1H-NMR-based metabonomics

Jan 2002
1439-1444

Brinde J
H Antti
Holmes E

BRINDE J, ANTTI H, HOLMES E, et al: Rapid and noninvasive diagnosis of the presence and severity of coronary heart disease using 1H-NMR-based metabonomics. Nature Med 8:1439-1444, 2002

Sirolimus, but not the structurally related RAD (everolimus), enhances the negative effects of cyclosporine on mitochondrial metabolism in the rat brain

Jan 2001
875-885

Serkova N
Niemann Jacobsen W
Cu

SERKOVA N, JACOBSEN W, NIEMANN CU, et al: Sirolimus, but not the structurally related RAD (everolimus), enhances the negative effects of cyclosporine on mitochondrial metabolism in the rat brain. Br J Pharmacol 133:875-885, 2001

Risk factors for delayed graft function after renal transplantation and their significance for long-term clinical outcome

Jan 2002
10

Hetzel

Organ-specific response to inhibition of mitochondrial metabolism by cyclosporine in the rat

Jan 2003
748

Serkova

1H-NMR-based metabolic signatures of mild and severe ischemia/reperfusion injury in rat kidney transplants

Abstract

No full-text available

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