Lipidomic Profiling of Kidney Cortical Tubule Segments Identifies Lipotypes with Physiological Implications.

A detailed knowledge of the lipid composition of components of nephrons is crucial for understanding physiological processes and the development of kidney diseases. However, the lipidomic composition of kidney tubular segments is unknown. We manually isolated the proximal convoluted tubule (PCT), the cortical thick ascending limb of Henle's loop, and the cortical collecting duct from 5 lean and obese mice and subjected the samples to shotgun lipidomics analysis by high-resolution mass spectrometry acquisition. Across all samples, more than 500 lipid species were identified, quantified, and compared. We observed significant compositional differences among the 3 tubular segments, which serve as true signatures. These intrinsic lipidomic features are associated with a distinct proteomic program that regulates highly specific physiological functions. The distinctive lipidomic features of each of the 3 segments are mostly based on the relative composition of neutral lipids, long-chain polyunsaturated fatty acids, sphingolipids, and ether phospholipids. These features support the hypothesis of a lipotype assigned to specific tubular segments. Obesity profoundly impacts the lipotype of PCT. In conclusion, we present a comprehensive lipidomic analysis of 3 cortical segments of mouse kidney tubules. This valuable resource provides unparalleled detail that enhances our understanding of tubular physiology and the potential impact of pathological conditions.

Improved Spatial Resolution of Metabolites in Tissue Biopsies Using High-Resolution Magic-Angle-Spinning Slice Localization NMR Spectroscopy.

High-resolution magic-angle-spinning 1H NMR spectroscopy (HR-MAS NMR) is a well-established technique for assessing the biochemical composition of intact tissue samples. In this study, we utilized a method based on HR-MAS NMR spectroscopy with slice localization (SLS) to achieve spatial resolution of metabolites. The obtained 7 slice spectra from each of the model samples (i.e., chicken thigh muscle with skin and murine renal biopsy including medulla (M) and cortex (C)) showed distinct metabolite compositions. Furthermore, we analyzed previously acquired 1H HR-MAS NMR spectra of separated cortex and medulla samples using multivariate statistical methods. Concentrations of glycerophosphocholine (GPC) were found to be significantly higher in the renal medulla compared to the cortex. Using GPC as a biomarker, we identified the tissue slices that were predominantly the cortex or medulla. This study demonstrates that HR-MAS SLS combined with multivariate statistics has the potential for identifying tissue heterogeneity and detailed biochemical characterization of complex tissue samples.

Bioimaging and Biodistribution of the Metal-Ion-Controlled Self-Assembly of PYY3-36 Studied by SPECT/CT.

The controlled self-assembly of peptide- and protein-based pharmaceuticals is of central importance for their mode of action and tuning of their properties. Peptide YY3-36 (PYY3-36 ) is a 36-residue peptide hormone that reduces food intake when peripherally administered. Herein, we describe the synthesis of a PYY3-36 analogue functionalized with a metal-ion-binding 2,2'-bipyridine ligand that enables self-assembly through metal complexation. Upon addition of CuII , the bipyridine-modified PYY3-36 peptide binds stoichiometric quantities of metal ions in solution and contributes to the organization of higher-order assemblies. In this study, we aimed to explore the size effect of the self-assembly in vivo by using non-invasive quantitative single-photon emission computed tomography/computed tomography (SPECT/CT) imaging. For this purpose, bipyridine-modified PYY3-36 was radiolabeled with a chelator holding 111 InIII , followed by the addition of CuII to the bipyridine ligand. SPECT/CT imaging and biodistribution studies showed fast renal clearance and accumulation in the kidney cortex. The radiolabeled bipyridyl-PYY3-36 conjugates with and without CuII presented a slightly slower excretion 1 h post injection compared to the unmodified-PYY3-36 , thus demonstrating that higher self-assemblies of the peptide might have an effect on the pharmacokinetics.

Microsomal and Cytosolic Scaling Factors in Dog and Human Kidney Cortex and Application for In Vitro-In Vivo Extrapolation of Renal Metabolic Clearance.

In vitro-in vivo extrapolation of drug metabolism data obtained in enriched preparations of subcellular fractions rely on robust estimates of physiologically relevant scaling factors for the prediction of clearance in vivo. The purpose of the current study was to measure the microsomal and cytosolic protein per gram of kidney (MPPGK and CPPGK) in dog and human kidney cortex using appropriate protein recovery marker and evaluate functional activity of human cortex microsomes. Cytochrome P450 (CYP) content and glucose-6-phosphatase (G6Pase) activity were used as microsomal protein markers, whereas glutathione-S-transferase activity was a cytosolic marker. Functional activity of human microsomal samples was assessed by measuring mycophenolic acid glucuronidation. MPPGK was 33.9 and 44.0 mg/g in dog kidney cortex, and 41.1 and 63.6 mg/g in dog liver (n = 17), using P450 content and G6Pase activity, respectively. No trends were noted between kidney, liver, and intestinal scalars from the same animals. Species differences were evident, as human MPPGK and CPPGK were 26.2 and 53.3 mg/g in kidney cortex (n = 38), respectively. MPPGK was 2-fold greater than the commonly used in vitro-in vivo extrapolation scalar; this difference was attributed mainly to tissue source (mixed kidney regions versus cortex). Robust human MPPGK and CPPGK scalars were measured for the first time. The work emphasized the importance of regional differences (cortex versus whole kidney-specific MPPGK, tissue weight, and blood flow) and a need to account for these to improve assessment of renal metabolic clearance and its extrapolation to in vivo.

Relationship between mercury in kidney, blood, and urine in environmentally exposed individuals, and implications for biomonitoring.

BACKGROUND: Individuals without occupational exposure are exposed to mercury (Hg) from diet and dental amalgam. The kidney is a critical organ, but there is limited information regarding the relationship between Hg in kidney (K-Hg), urine (U-Hg), blood (B-Hg), and plasma (P-Hg). OBJECTIVES: The aim was to determine the relationship between K-Hg, U-Hg, B-Hg, and P-Hg among environmentally exposed individuals, estimate the biological half-time of K-Hg, and provide information useful for biomonitoring of Hg. METHODS: Kidney cortex biopsies and urine and blood samples were collected from 109 living kidney donors. Total Hg concentrations were determined and the relationships between K-Hg, U-Hg, P-Hg, and B-Hg were investigated in regression models. The half-time of K-Hg was estimated from the elimination constant. RESULTS: There were strong associations between K-Hg and all measures of U-Hg and P-Hg (rp=0.65-0.84, p<0.001), while the association with B-Hg was weaker (rp=0.29, p=0.002). Mean ratios between K-Hg (in µg/g) and U-Hg/24h (in µg) and B-Hg (in µg/L) were 0.22 and 0.19 respectively. Estimates of the biological half-time varied between 30 and 92days, with significantly slower elimination in women. Adjusting overnight urine samples for dilution using urinary creatinine resulted in less bias in relation to K-Hg or U-Hg/24h, compared with other adjustment techniques. CONCLUSIONS: The relationship between K-Hg and U-Hg is approximately linear. K-Hg can be estimated using U-Hg and gender. Women have longer half-time of Hg in kidney compared to men. Adjusting overnight urine samples for creatinine concentration resulted in less bias.

Proteome analysis of acute kidney injury - Discovery of new predominantly renal candidates for biomarker of kidney disease.

The main bottleneck in studies aiming to identify novel biomarkers in acute kidney injury (AKI) has been the identification of markers that are organ and process specific. Here, we have used different tissues from a controlled porcine renal ischemia/reperfusion (I/R) model to identify new, predominantly renal biomarker candidates for kidney disease. Urine and serum samples were analyzed in pre-ischemia, ischemia (60min) and 4, 11 and 16h post-reperfusion, and renal cortex samples after 24h of reperfusion. Peptides were analyzed on the Q-Exactive™. In renal cortex proteome, we observed an increase in the synthesis of proteins in the ischemic kidney compared to the contralateral, highlighted by transcription factors and epithelial adherens junction proteins. Intersecting the set of proteins up- or down-regulated in the ischemic tissue with both serum and urine proteomes, we identified 6 proteins in the serum that may provide a set of targets for kidney injury. Additionally, we identified 49, being 4 predominantly renal, proteins in urine. As prove of concept, we validated one of the identified biomarkers, dipeptidyl peptidase IV, in a set of patients with diabetic nephropathy. In conclusion, we identified 55 systemic proteins, some of them predominantly renal, candidates for biomarkers of renal disease. BIOLOGICAL SIGNIFICANCE: The main bottleneck in studies aiming to identify novel biomarkers in acute kidney injury (AKI) has been the identification of markers that are predominantly renal. In fact, putative biomarkers for this condition have also been identified in a number of other clinical scenarios, such as cardiovascular diseases, chronic kidney failure or in patients being treated in intensive care units from a number of conditions. Here we propose a comprehensive, sequential screening procedure able to identify and validate potential biomarkers for kidney disease, using kidney ischemia/reperfusion as a paradigm for a kidney pathological event.

Cadmium, lead and mercury concentrations in pathologically altered human kidneys.

Heavy metals, including cadmium (Cd), lead (Pb) and mercury (Hg) act as nephrotoxic agents, particularly in the renal cortex. The aim of the study was to determine the concentrations of Cd, Pb and Hg in kidneys removed from patients due to lesions of various etiologies and from patients after the rejection of transplanted kidneys. Additionally, we determined the influence of selected biological and environmental factors on the concentrations of toxic metals. The study material consisted of kidneys with tumor lesions (n = 27), without tumors (n = 7) and its extracted grafts (n = 10) obtained from patients belongs to the north-western areas of Poland. The determined metal concentrations in the renal cortex and medulla may be arranged in the following descending order: Cd > Pb > Hg. The highest concentrations of Cd and Hg were found in the cortex, while the maximum content Pb was observed in the medulla. Significant correlations were found in the concentrations of the same metals between cortex and medulla and between Pb and Hg in the renal medulla. Pb content was higher in the renal medulla of men than in the cortex of the elderly (above 60 years of age). The highest concentrations of Pb and Hg were found in the cortex and medulla, of the kidneys had not neoplastic changes, and lower content of these metals were found in the extracted kidney grafts. In summary, renal grafts accumulate less heavy metals than cancerous kidneys, what could have been caused by immunosuppressors taken by the graft recipients. Moreover, sex, age and smoking are key factors responsible for xenobiotics concentrations.

Set of Novel Automated Quantitative Microproteomics Protocols for Small Sample Amounts and Its Application to Kidney Tissue Substructures.

Here we assessed the ability of an automated sample preparation device equipped with disposable microcolumns to prepare mass-limited samples for high-sensitivity quantitative proteomics, using both label-free and isobaric labeling approaches. First, we compared peptide label-free quantification reproducibility for 1.5-150 µg of cell lysates and found that labware preconditioning was essential for reproducible quantification of <7.5 µg digest. Second, in-solution and on-column tandem mass tag (TMT) labeling protocols were compared and optimized for 1 µg of sample. Surprisingly, standard methods for in-solution and on-column labeling showed poor TMT labeling (50-85%); however, novel optimized and automated protocols restored efficient labeling to >98%. Third, compared with a single long gradient experiment, a simple robotized high-pH fractionation protocol using only 6 µg of starting material doubled the number of unique peptides and increased proteome coverage 1.43-fold. To facilitate the analysis of heterogeneous tissue samples, such as those obtained from laser capture microdissection, a modified BCA protein assay was developed that consumes and detects down to 15 ng of protein. As a proof-of-principle, the modular automated workflow was applied to 0.5 and 1 mm2 mouse kidney cortex and medulla microdissections to show the method's potential for real-life small sample sources and to create kidney substructure-specific proteomes.

The first investigation of Wilms' tumour atomic structure-nitrogen and carbon isotopic composition as a novel biomarker for the most individual approach in cancer disease.

The paper describes a novel approach to investigating Wilms' tumour (nephroblastoma) biology at the atomic level. Isotope Ratio Mass Spectrometry (IRMS) was used to directly assess the isotope ratios of nitrogen and carbon in 84 Wilms' tumour tissue samples from 28 cases representing the histological spectrum of nephroblastoma. Marked differences in nitrogen and carbon isotope ratios were found between nephroblastoma histological types and along the course of cancer disease, with a breakout in isotope ratio of the examined elements in tumour tissue found between stages 2 and 3. Different isotopic compositions with regard to nitrogen and carbon content were observed in blastemal Wilms' tumour, with and without focal anaplasia, and in poorly- and well-differentiated epithelial nephroblastoma. This first assessment of nitrogen and carbon isotope ratio reveals the previously unknown part of Wilms' tumour biology and represents a potential novel biomarker, allowing for a highly individual approach to treating cancer. Furthermore, this method of estimating isotopic composition appears to be the most sensitive tool yet for cancer tissue evaluation, and a valuable complement to established cancer study methods with prospective clinical impact.

Decellularized Human Kidney Cortex Hydrogels Enhance Kidney Microvascular Endothelial Cell Maturation and Quiescence.

The kidney peritubular microvasculature is highly susceptible to injury from drugs and toxins, often resulting in acute kidney injury and progressive chronic kidney disease. Little is known about the process of injury and regeneration of human kidney microvasculature, resulting from the lack of appropriate kidney microvascular models that can incorporate the proper cells, extracellular matrices (ECMs), and architectures needed to understand the response and contribution of individual vascular components in these processes. In this study, we present methods to recreate the human kidney ECM (kECM) microenvironment by fabricating kECM hydrogels derived from decellularized human kidney cortex. The majority of native matrix proteins, such as collagen-IV, laminin, and heparan sulfate proteoglycan, and their isoforms were preserved in similar proportions as found in normal kidneys. Human kidney peritubular microvascular endothelial cells (HKMECs) became more quiescent when cultured on this kECM gel compared with culture on collagen-I-assessed using phenotypic, genotypic, and functional assays; whereas human umbilical vein endothelial cells became stimulated on kECM gels. We demonstrate for the first time that human kidney cortex can form a hydrogel suitable for use in flow-directed microphysiological systems. Our findings strongly suggest that selecting the proper ECM is a critical consideration in the development of vascularized organs on a chip and carries important implications for tissue engineering of all vascularized organs.

The Feasibility of Magnetic Resonance Imaging for Quantification of Liver, Pancreas, Spleen, Vertebral Bone Marrow, and Renal Cortex R2* and Proton Density Fat Fraction in Transfusion-Related Iron Overload.

OBJECTIVE: We aimed to evaluate the feasibility of quantification of liver, pancreas, spleen, vertebral bone marrow, and renal cortex R2* and magnetic resonance imaging-proton density fat fraction (MRI-PDFF) and to evaluate the correlations among them in patients with transfusion-related iron overload. MATERIALS AND METHODS: A total of 9 patients (5 boys, 4 girls) who were referred to our clinic with suspicion of hepatic iron overload were included in this study. All patients underwent T1-independent volumetric multi-echo gradient-echo imaging with T2* correction and spectral fat modeling. MRI examinations were performed on a 1.5 T MRI system. RESULTS: All patients had hepatic iron overload. Severe hepatic iron overload was recorded in 5/9 patients (56%), and when we evaluated the PDFF maps of these patients, we observed an extensive patchy artifact in the liver in 4 of 5 patients (R2* greater than 671 Hz). When we performed MRI-PDFF measurements despite these artifacts, we observed artifactual high MRI-PDFF values. There was a close correlation between average pancreas R2* and average pancreas MRI-PDFF (p=0.003, r=0.860). There was a significant correlation between liver R2* and average pancreas R2* (p=0.021, r=0.747), liver R2* and renal cortex R2* (p=0.020, r=0.750), and average pancreas R2* and renal cortex R2* (p=0.003, r=0.858). There was a significant negative correlation between vertebral bone marrow R2* and age (p=0.018, r=-0.759). CONCLUSION: High iron content of the liver, especially with a T2* value shorter than the first echo time can spoil the efficacy of PDFF calculation. Fat deposition in the pancreas is accompanied by pancreatic iron overload. There is a significant correlation between hepatic siderosis and pancreatic siderosis. Renal cortical and pancreatic siderosis are correlated, too.

Fibromyxoid nephrogenic adenoma protruding in a renal cortical cyst. A rare morphological variant in an outstanding location.

Nephrogenic adenoma (NA) is an unusual, benign lesion of the urinary tract, generally presenting in the bladder and with less frequency in the renal pelvis, urethra or ureter. It consists of tubules, microcysts and papillae lined by a single layer of low cuboidal epithelium without atypia. Recently, a fibromyxoid variant mimicking an infiltrating mucinous adenocarcinoma has been described. We report hereby the case of a 70-year-old female with a fibromixoid NA protruding in a renal cortical cyst. Only one case of NA in a renal cortical cyst has been found in the literature and it was of the classical type. The development of a NA in a renal cortical cyst lends support to the theory that the NA results from proliferation of secondarily implanted exfoliated renal epithelial cells.

Mitochondrial proteomes of porcine kidney cortex and medulla: foundation for translational proteomics.

BACKGROUND: Emerging evidence has linked mitochondrial dysfunction to the pathogenesis of many renal disorders, including acute kidney injury, sepsis and even chronic kidney disease. Proteomics is a powerful tool in elucidating the role of mitochondria in renal pathologies. Since the pig is increasingly recognized as a major mammalian model for translational research, the lack of physiological proteome data of large mammals prompted us to examine renal mitochondrial proteome in porcine kidney cortex and medulla METHODS: Kidneys were obtained from six healthy pigs. Mitochondria from cortex and medulla were isolated using differential centrifugation and proteome maps of cortical and medullar mitochondria were constructed using two-dimensional gel electrophoresis (2DE). Protein spots with significant difference between mitochondrial fraction of renal cortex and medulla were identified by mass spectrometry. RESULTS: Proteomic analysis identified 81 protein spots. Of these spots, 41 mitochondrial proteins were statistically different between renal cortex and medulla (p < 0.05). Protein spots containing enzymes of beta oxidation, amino acid metabolism, and gluconeogenesis were predominant in kidney cortex mitochondria. Spots containing tricarboxylic acid cycle enzymes and electron transport system proteins, proteins maintaining metabolite transport and mitochondrial translation were more abundant in medullar mitochondria. CONCLUSION: This study provides the first proteomic profile of porcine kidney cortex and medullar mitochondrial proteome. Different protein expression pattern reflects divergent functional metabolic role of mitochondria in various kidney compartments. Our study could serve as a useful reference for further porcine experiments investigating renal mitochondrial physiology under various pathological states.

[PREPARATION OF HUMAN TISSUE PROTEIN EXTRACTS ENRICHED WITH THE SPHINGOMYELIN SYNTHASE 1].

Sphingomyelin synthase 1 (SMS 1) catalyzes sphingomyelin biosynthesis in eukaryotic cells. We previously studied the structure of the human SGMS1 gene, which encodes the enzyme and its numerous transcripts. The tissue-specific expression of the transcripts was also described. Analysis of the SMS1 protein expression in human tissues using immunoblotting of tissue extracts prepared in the RIPA (Radio Immuno-Precipitation Assay) buffer revealed a weak signal in renal cortex, testis, lung, and no signal in placenta and lymphatic node. In this work, a new method of preparation of the tissue protein extracts enriched with SMS1 was suggested. The method based on the consecutive extraction with a buffer containing 0.05 and 1 mg/ml of the Quillaja saponaria saponin allowed SMS1 to be detected in all tissues tested. The SMS1 content in the saponin extract of kidney cortex is about 12-fold higher compared to the RIPA extraction procedure.

Differential Effect of Renal Cortical and Medullary Interstitial Fluid Calcium on Blood Pressure Regulation in Salt-Sensitive Hypertension.

BACKGROUND: Hypercalciuria is a frequent characteristic of hypertension. In this report we extend our earlier studies investigating the role of renal interstitial fluid calcium (ISF(Ca))(2+) as a link between urinary calcium excretion and blood pressure in the Dahl salt-sensitive (DS) hypertensive model. METHODS: Dahl salt-sensitive and salt-resistant (DR) rats were placed on control (0.45%) and high (8%) salt diets to determine if changes in renal cortical and medullary ISF(Ca)(2+)correlated with changes in urinary calcium excretion and blood pressure. RESULTS: We observed that renal ISFCa(2+) was predicted by urinary calcium excretion (P < 0.05) in DS rats but not DR rats. Renal cortical ISF(Ca)(2+) was negatively associated with blood pressure (P < 0.03) while renal medullary ISF(Ca)(2+) was positively associated with blood pressure in DS rats (P < 0.04). In contrast, neither urinary calcium excretion nor renal ISF(Ca)(2+) was associated with blood pressure in the DR rats under the conditions of this study. CONCLUSION: We interpret these findings to suggest that decreased renal cortical ISF(Ca)(2+) plays a role in the increase in blood pressure following a high salt diet in salt hypertension perhaps by mediating renal vasoconstriction; the role of medullary calcium remains to be fully understood. Further studies are needed to determine the mechanism of the altered renal ISF(Ca)(2+) and its role in blood pressure regulation.

Eicosanoid post-mortem induction in kidney tissue is prevented by microwave irradiation.

Previously, we, and others, have demonstrated a rapid and significant post-mortem increase in brain prostanoid (PG) levels analyzed without microwave fixation, and this is not the result of PG trapping or destruction in microwave-irradiated brain tissue. In the present study, we demonstrate a dramatic increase in kidney eicosanoid levels when analyzed without microwave fixation which was mainly accounted for by the 142-, 81-, and 62-fold increase in medullary 6-ketoPGF1α, PGE2, and PGF2α, levels, respectively, while PGD2 and TXB2 levels were increased ~7-fold. Whole kidney and cortex PG were also significantly increased in non-microwaved tissue, but at lesser extent. Arachidonic acid and the lipoxygenase products hydroxyeicosatetraenoic acids (HETE) were also induced in whole kidney, cortex, and medulla 1.5- to 5.5-fold depending upon tissue and metabolite. Cyclooxygenase inhibition with indomethacin decreased PG mass in non-microwaved tissue to basal levels, however HETE and arachidonic acid were not decreased. These data demonstrate the critical importance of kidney tissue fixation to limiting artifacts during kidney eicosanoid analysis.

The relationship between cadmium in kidney and cadmium in urine and blood in an environmentally exposed population.

INTRODUCTION: Cadmium (Cd) is toxic to the kidney and a major part of the body burden occurs here. Cd in urine (U-Cd) and blood (B-Cd) are widely-used biomarkers for assessing Cd exposure or body burden. However, empirical general population data on the relationship between Cd in kidney (K-Cd), urine, and blood are scarce. Our objectives were to determine the relationship between cadmium in kidney, urine, and blood, and calculate the elimination half-time of Cd from the kidney. METHODS: Kidney cortex biopsies, urine, and blood samples were collected from 109 living kidney donors. Cd concentrations were determined and the relationships between K-Cd, U-Cd, and B-Cd were investigated in regression models. The half-time of K-Cd was estimated from the elimination constant. RESULTS: There was a strong association between K-Cd and U-Cd adjusted for creatinine (rp=0.70, p<0.001), while the association with B-Cd was weaker (rp=0.44, p<0.001). The relationship between K-Cd and U-Cd was nonlinear, with slower elimination of Cd at high K-Cd. Estimates of the K-Cd half-time varied between 18 and 44years. A K-Cd of 25µg/g corresponds to U-Cd of 0.42µg/g creatinine in overnight urine (U-Cd/K-Cd ratio: about 1:60). Multivariate models showed Cd in blood and urinary albumin as determinants for U-Cd excretion. DISCUSSION: In healthy individuals with low-level Cd exposure, there was a strong correlation between Cd in kidney and urine, especially after adjustment for creatinine. Urinary Cd was also affected by Cd in blood and urinary albumin. Previous estimates of the U-Cd/K-Cd ratio may underestimate K-Cd at low U-Cd.

Correlation of histopathology, urinary biomarkers, and gene expression responses following hexachloro-1:3-butadiene-induced acute nephrotoxicity in male Hanover Wistar rats: a 28-day time course study.

Hexachloro-1:3-butadiene (HCBD) causes segment-specific injury to the proximal renal tubule. A time course study of traditional and more recently proposed urinary biomarkers was performed in male Hanover Wistar rats receiving a single intraperitoneal (ip) injection of 45 mg/kg HCBD. Animals were killed on days 1, 2, 3, 4, 5, 6, 7, 10, 14, and 28 postdosing and the temporal response of renal biomarkers was characterized using kidney histopathology, urinary and serum biochemistry, and gene expression. Histopathologic evidence of tubular degeneration was seen from day 1 until day 3 postdosing and correlated with increased urinary levels of α-glutathione S-transferase (α-GST), albumin, glucose, and kidney injury molecule-1 (KIM-1), and increased gene expression of KIM-1, NAD(P)H dehydrogenase, quinone 1, and heme oxygenase (decycling) 1. Histopathologic evidence of tubular regeneration was seen from day 2 postdosing and correlated with raised levels of urinary KIM-1 and osteopontin and increased gene expression of KIM-1 and annexin A7. Traditional renal biomarkers generally demonstrated low sensitivity. It is concluded that in rat proximal tubular injury, measurement of a range of renal biomarkers, in conjunction with gene expression analysis, provides an understanding of the extent of degenerative changes induced in the kidney and the process of regeneration.

Aristolactam-DNA adducts are a biomarker of environmental exposure to aristolochic acid.

Endemic (Balkan) nephropathy is a chronic tubulointerstitial disease frequently accompanied by urothelial cell carcinomas of the upper urinary tract. This disorder has recently been linked to exposure to aristolochic acid, a powerful nephrotoxin and human carcinogen. Following metabolic activation, aristolochic acid reacts with genomic DNA to form aristolactam-DNA adducts that generate a unique TP53 mutational spectrum in the urothelium. The aristolactam-DNA adducts are concentrated in the renal cortex, thus serving as biomarkers of internal exposure to aristolochic acid. Here, we present molecular epidemiologic evidence relating carcinomas of the upper urinary tract to dietary exposure to aristolochic acid. DNA was extracted from the renal cortex and urothelial tumor tissue of 67 patients that underwent nephroureterectomy for carcinomas of the upper urinary tract and resided in regions of known endemic nephropathy. Ten patients from nonendemic regions with carcinomas of the upper urinary tract served as controls. Aristolactam-DNA adducts were quantified by (32)P-postlabeling, the adduct was confirmed by mass spectrometry, and TP53 mutations in tumor tissues were identified by chip sequencing. Adducts were present in 70% of the endemic cohort and in 94% of patients with specific A:T to T:A mutations in TP53. In contrast, neither aristolactam-DNA adducts nor specific mutations were detected in tissues of patients residing in nonendemic regions. Thus, in genetically susceptible individuals, dietary exposure to aristolochic acid is causally related to endemic nephropathy and carcinomas of the upper urinary tract.

Effects of lead and/or cadmium on the distribution patterns of some essential trace elements in immature female rats.

Lead acetate (300 mg/L) and/or cadmium chloride (50 mg/L) were administered as drinking water to Sprague-Dawley rats for 9 weeks to investigate the effects of concurrent exposure to lead and cadmium on the distribution patterns of five essential trace elements. Inductively coupled plasma mass spectrometry was used to determine the concentrations of zinc, copper, manganese, selenium and iron in the urine at different exposure times, as well as their levels in the renal cortex and serum at the end of treatment. Compared with the control group, exposure to lead and/or cadmium resulted in a significant increase in the urinary excretion of these five elements during the experiment, whereas significant decreased levels of these elements were found in kidney and serum. In conclusion, increased urinary loss of antioxidant trace elements due to lead and/or cadmium exposure induced the deficiency of antioxidants in the body, which could result in further oxidative damage. Moreover, there was an obvious synergistic effect of lead combined with cadmium on the distribution patterns of these essential trace elements, which may be related to the severity of co-exposure to these two metals.