Drug toxicity in the proximal tubule: new models, methods and mechanisms.

The proximal tubule (PT) reabsorbs most of the glomerular filtrate and plays an important role in the uptake, metabolism and excretion of xenobiotics. Some therapeutic drugs are harmful to the PT, and resulting nephrotoxicity is thought to be responsible for approximately 1 in 6 of cases of children hospitalized with acute kidney injury (AKI). Clinically, PT dysfunction leads to urinary wasting of important solutes normally reabsorbed by this nephron segment, leading to systemic complications such as bone demineralization and a clinical scenario known as the renal Fanconi syndrome (RFS). While PT defects can be diagnosed using a combination of blood and urine markers, including urinary excretion of low molecular weight proteins (LMWP), standardized definitions of what constitutes clinically significant toxicity are lacking, and identifying which patients will go on to develop progressive loss of kidney function remains a major challenge. In addition, much of our understanding of cellular mechanisms of drug toxicity is still limited, partly due to the constraints of available cell and animal models. However, advances in new and more sophisticated in vitro models of the PT, along with the application of high-content analytical methods that can provide readouts more relevant to the clinical manifestations of nephrotoxicity, are beginning to extend our knowledge. Such technical progress should help in discovering new biomarkers that can better detect nephrotoxicity earlier and predict its long-term consequences, and herald a new era of more personalized medicine.

Predicting the protein composition of human urine in normal and pathological states: Quantitative description based on Dent1 disease (CLCN5 mutation).

KEY POINTS: The presence of plasma proteins in urine is difficult to interpret quantitatively. It may be a result of impaired glomerular filtration or impaired proximal tubule (PT) reabsorption, or both. Dent1 disease (CLCN5 mutation) abolishes PT protein reabsorption leaving glomerular function intact. Using urine protein measurements from patients with Dent1 disease and normal individuals, we devised a mathematical model that incorporates two PT transport processes with distinct kinetics. This model predicts albumin, α1 -microglobulin (α1 -m), ß2 -microglobulin (ß2 -m) and retinol-binding protein 4 (RBP4) urine concentrations. Our results indicate that the urinary excretion of ß2 -m and RBP4 differs from that of albumin and α1 -m in their sensitivity to changes in the glomerular filtration rate, glomerular protein leak, tubular protein uptake via endocytosis and PT water reabsorption. The model predicts quantitatively how hyperfiltration and glomerular leak interact to promote albuminuria. Our model should contribute to improved understanding and interpretation of urine protein measurements in renal disease. ABSTRACT: To clarify the relative contributions of glomerular filtration and tubular uptake to urinary protein excretion, we developed a mathematical model of protein reabsorption in the human proximal tubule (PT) using Michaelis-Menten kinetics and molar urinary protein measurements taken from human Dent1 disease (CLCN5 loss-of-function mutation). ß2 -Microglobulin (ß2 -m) and retinol-binding protein 4 (RBP4) are normally reabsorbed with 'very high' efficiency uptake kinetics and fractional urinary excretion of 0.025%, whereas albumin and α1 -microglobulin (α1 -m) are reabsorbed by 'high' efficiency uptake kinetics and 50-fold higher fractional urinary excretion of 1.15%. Our model correctly predicts the urinary ß2 -m, RBP4 and α1 -m content in aristolochic acid nephropathy, and elevated ß2 -m excretion with increased single nephron glomerular filtration rate (SNGFR) following unilateral-nephrectomy. We explored how altered endocytic uptake, water reabsorption, SNGFR and glomerular protein filtration affect excretion. Our results help to explain why ß2 -m and RBP4 are more sensitive markers of PT dysfunction than albumin or α1 -m, and suggest that reduced PT sodium and water reabsorption in Fanconi syndrome may contribute to proteinuria. Transition of albumin excretion from normal to microalbuminuria, a 5-fold increase, corresponds to a 3.5-fold elevation in albumin glomerular filtration, supporting the use of microalbuminuria screening to detect glomerular leak in diabetes. In macroalbuminuria, small albumin permeability changes produce large changes in excretion. However, changes in SNGFR can alter protein excretion, and hyperfiltration with glomerular leak can combine to increase albuminuria. Our model provides a validated quantitative description of the transport processes underlying the protein composition of human urine in normal and pathophysiological states.

Multiparametric imaging reveals that mitochondria-rich intercalated cells in the kidney collecting duct have a very high glycolytic capacity.

Alpha intercalated cells (αICs) in the kidney collecting duct (CD) belong to a family of mitochondria rich cells (MRCs) and have a crucial role in acidifying the urine via apical V-ATPase pumps. The nature of metabolism in αICs and its relationship to transport was not well-understood. Here, using multiphoton live cell imaging in mouse kidney tissue, FIB-SEM, and other complementary techniques, we provide new insights into mitochondrial structure and function in αICs. We show that αIC mitochondria have a rounded structure and are not located in close proximity to V-ATPase containing vesicles. They display a bright NAD(P)H fluorescence signal and low uptake of voltage-dependent dyes, but are energized by a pH gradient. However, expression of complex V (ATP synthase) is relatively low in αICs, even when stimulated by metabolic acidosis. In contrast, anaerobic glycolytic capacity is surprisingly high, and sufficient to maintain intracellular calcium homeostasis in the presence of complete aerobic inhibition. Moreover, glycolysis is essential for V-ATPase-mediated proton pumping. Key findings were replicated in narrow/clear cells in the epididymis, also part of the MRC family. In summary, using a range of cutting-edge techniques to investigate αIC metabolism in situ, we have discovered that these mitochondria dense cells have a high glycolytic capacity.

Brain dysfunction in tubular and tubulointerstitial kidney diseases.

Kidney function has two important elements: glomerular filtration and tubular function (secretion and reabsorption). A persistent decrease in glomerular filtration rate (GFR), with or without proteinuria, is diagnostic of chronic kidney disease (CKD). While glomerular injury or disease is a major cause of CKD and usually associated with proteinuria, predominant tubular injury, with or without tubulointerstitial disease, is typically non-proteinuric. CKD has been linked with cognitive impairment, but it is unclear how much this depends on a decreased GFR, altered tubular function or the presence of proteinuria. Since CKD is often accompanied by tubular and interstitial dysfunction, we explore here for the first time the potential role of the tubular and tubulointerstitial compartments in cognitive dysfunction. To help address this issue we selected a group of primary tubular diseases with preserved GFR in which to review the evidence for any association with brain dysfunction. Cognition, mood, neurosensory and motor disturbances are not well characterized in tubular diseases, possibly because they are subclinical and less prominent than other clinical manifestations. The available literature suggests that brain dysfunction in tubular and tubulointerstitial diseases is usually mild and is more often seen in disorders of water handling. Brain dysfunction may occur when severe electrolyte and water disorders in young children persist over a long period of time before the diagnosis is made. We have chosen Bartter and Gitelman syndromes and nephrogenic diabetes insipidus as examples to highlight this topic. We discuss current published findings, some unanswered questions and propose topics for future research.

The 1918 Influenza Pandemic: Back to the Future?

BACKGROUND: It is just over a century since the 1918 flu pandemic, sometimes referred to as the "mother" of pandemics. This brief retrospective of the 1918 pandemic is taken from the viewpoint of the current SARS-CoV-2/COVID-19 pandemic and is based on a short lecture given during the 2021 Virtual Congress of the ERA-EDTA. SUMMARY: This review summarizes and highlights some of the earlier pandemic's salient features, some parallels with today, and some potential learnings, bearing in mind that the flu pandemic occurred over 100 years ago at a time of major turmoil during the climax to WWl, and with limited medical expertise and knowledge, research facilities, or well-structured and resourced healthcare services. While there is little or no information on renal complications at the time, or an effective treatment, some observations in relation to COVID-19 and vaccination are included. Key Messages: Lessons are difficult to draw from 1918 other than the importance and value of non-pharmaceutical measures to limit viral transmission. While the economic impact of the 1918 pandemic was significant, as it is now with COVID-19, subsequent economic analysis has shown that protecting public health and preserving economic activity are not mutually exclusive. Both H1N1 and SARS-CoV-2 viruses are neurotropic and may cause chronically debilitating neurological diseases, including conditions such as encephalitis lethargica (still debated) and myalgic encephalomyelitis (chronic fatigue syndrome), respectively. Although coronavirus and influenza viral infections have some similarities, they are certainly not the same, as we are realising, and future infectious pandemics may still surprise us, but being "forewarned is forearmed."

Physiological regulation of phosphate by vitamin D, parathyroid hormone (PTH) and phosphate (Pi).

Inorganic phosphate (Pi) is an abundant element in the body and is essential for a wide variety of key biological processes. It plays an essential role in cellular energy metabolism and cell signalling, e.g. adenosine and guanosine triphosphates (ATP, GTP), and in the composition of phospholipid membranes and bone, and is an integral part of DNA and RNA. It is an important buffer in blood and urine and contributes to normal acid-base balance. Given its widespread role in almost every molecular and cellular function, changes in serum Pi levels and balance can have important and untoward effects. Pi homoeostasis is maintained by a counterbalance between dietary Pi absorption by the gut, mobilisation from bone and renal excretion. Approximately 85% of total body Pi is present in bone and only 1% is present as free Pi in extracellular fluids. In humans, extracellular concentrations of inorganic Pi vary between 0.8 and 1.2 mM, and in plasma or serum Pi exists in both its monovalent and divalent forms (H2PO4- and HPO42-). In the intestine, approximately 30% of Pi absorption is vitamin D regulated and dependent. To help maintain Pi balance, reabsorption of filtered Pi along the renal proximal tubule (PT) is via the NaPi-IIa and NaPi-IIc Na+-coupled Pi cotransporters, with a smaller contribution from the PiT-2 transporters. Endocrine factors, including, vitamin D and parathyroid hormone (PTH), as well as newer factors such as fibroblast growth factor (FGF)-23 and its coreceptor α-klotho, are intimately involved in the control of Pi homeostasis. A tight regulation of Pi is critical, since hyperphosphataemia is associated with increased cardiovascular morbidity in chronic kidney disease (CKD) and hypophosphataemia with rickets and growth retardation. This short review considers the control of Pi balance by vitamin D, PTH and Pi itself, with an emphasis on the insights gained from human genetic disorders and genetically modified mouse models.

Liquorice, Liddle, Bartter or Gitelman-how to differentiate?

Hypokalaemia with alkalosis can suggest excess aldosterone. Aldosterone stimulates the collecting duct mineralocorticoid receptor (MR) to upregulate the epithelial sodium channel (ENaC) and stimulate electrogenic sodium reabsorption, with secretion of potassium and protons. Gitelman, Bartter and Liddle syndrome, and liquorice ingestion all cause hypokalaemic alkalosis. This mini-review outlines the pathophysiology of these conditions as well as how to differentiate them.

Renal Tubular Cell Mitochondrial Dysfunction Occurs Despite Preserved Renal Oxygen Delivery in Experimental Septic Acute Kidney Injury.

OBJECTIVE: To explain the paradigm of significant renal functional impairment despite preserved hemodynamics and histology in sepsis-induced acute kidney injury. DESIGN: Prospective observational animal study. SETTING: University research laboratory. SUBJECTS: Male Wistar rats. INTERVENTION: Using a fluid-resuscitated sublethal rat model of fecal peritonitis, changes in renal function were characterized in relation to global and renal hemodynamics, and histology at 6 and 24 hours (n = 6-10). Sham-operated animals were used as comparison (n = 8). Tubular cell mitochondrial function was assessed using multiphoton confocal imaging of live kidney slices incubated in septic serum. MEASUREMENTS AND MAIN RESULTS: By 24 hours, serum creatinine was significantly elevated with a concurrent decrease in renal lactate clearance in septic animals compared with sham-operated and 6-hour septic animals. Renal uncoupling protein-2 was elevated in septic animals at 24 hours although tubular cell injury was minimal and mitochondrial ultrastructure in renal proximal tubular cells preserved. There was no significant change in global or renal hemodynamics and oxygen delivery/consumption between sham-operated and septic animals at both 6- and 24-hour timepoints. In the live kidney slice model, mitochondrial dysfunction was seen in proximal tubular epithelial cells incubated with septic serum with increased production of reactive oxygen species, and decreases in nicotinamide adenine dinucleotide and mitochondrial membrane potential. These effects were prevented by coincubation with the reactive oxygen species scavenger, 4-hydroxy-2,2,6,6-tetramethyl-piperidin-1-oxyl. CONCLUSIONS: Renal dysfunction in sepsis occurs independently of hemodynamic instability or structural damage. Mitochondrial dysfunction mediated by circulating mediators that induce local oxidative stress may represent an important pathophysiologic mechanism.

Purinergic signaling in kidney disease.

Nucleotides are key subunits for nucleic acids and provide energy for intracellular metabolism. They can also be released from cells to act physiologically as extracellular messengers or pathologically as danger signals. Extracellular nucleotides stimulate membrane receptors in the P2 and P1 family. P2X are ATP-activated cation channels; P2Y and P1 are G-protein coupled receptors activated by ATP, ADP, UTP, and UDP in the case of P2 or adenosine for P1. Renal P2 receptors influence both vascular contractility and tubular function. Renal cells also express ectonucleotidases that rapidly hydrolyze extracellular nucleotides. These enzymes integrate this multireceptor purinergic-signaling complex by determining the nucleotide milieu to titrate receptor activation. Purinergic signaling also regulates immune cell function by modulating the synthesis and release of various cytokines such as IL1-ß and IL-18 as part of inflammasome activation. Abnormal or excessive stimulation of this intricate paracrine system can be pro- or anti-inflammatory, and is also linked to necrosis and apoptosis. Kidney tissue injury causes a localized increase in ATP concentration, and sustained activation of P2 receptors can lead to renal glomerular, tubular, and vascular cell damage. Purinergic receptors also regulate the activity and proliferation of fibroblasts, promoting both inflammation and fibrosis in chronic disease. In this short review we summarize some of the recent findings related to purinergic signaling in the kidney. We focus predominantly on the P2X7 receptor, discussing why antagonists have so far disappointed in clinical trials and how advances in our understanding of purinergic signaling might help to reposition these compounds as potential treatments for renal disease.

Mistargeting of peroxisomal EHHADH and inherited renal Fanconi's syndrome.

BACKGROUND: In renal Fanconi's syndrome, dysfunction in proximal tubular cells leads to renal losses of water, electrolytes, and low-molecular-weight nutrients. For most types of isolated Fanconi's syndrome, the genetic cause and underlying defect remain unknown. METHODS: We clinically and genetically characterized members of a five-generation black family with isolated autosomal dominant Fanconi's syndrome. We performed genomewide linkage analysis, gene sequencing, biochemical and cell-biologic investigations of renal proximal tubular cells, studies in knockout mice, and functional evaluations of mitochondria. Urine was studied with the use of proton nuclear magnetic resonance ((1)H-NMR) spectroscopy. RESULTS: We linked the phenotype of this family's Fanconi's syndrome to a single locus on chromosome 3q27, where a heterozygous missense mutation in EHHADH segregated with the disease. The p.E3K mutation created a new mitochondrial targeting motif in the N-terminal portion of EHHADH, an enzyme that is involved in peroxisomal oxidation of fatty acids and is expressed in the proximal tubule. Immunocytofluorescence studies showed mistargeting of the mutant EHHADH to mitochondria. Studies of proximal tubular cells revealed impaired mitochondrial oxidative phosphorylation and defects in the transport of fluids and a glucose analogue across the epithelium. (1)H-NMR spectroscopy showed elevated levels of mitochondrial metabolites in urine from affected family members. Ehhadh knockout mice showed no abnormalities in renal tubular cells, a finding that indicates a dominant negative nature of the mutation rather than haploinsufficiency. CONCLUSIONS: Mistargeting of peroxisomal EHHADH disrupts mitochondrial metabolism and leads to renal Fanconi's syndrome; this indicates a central role of mitochondria in proximal tubular function. The dominant negative effect of the mistargeted protein adds to the spectrum of monogenic mechanisms of Fanconi's syndrome. (Funded by the European Commission Seventh Framework Programme and others.).

Sequential Analysis of a Panel of Biomarkers and Pathologic Findings in a Resuscitated Rat Model of Sepsis and Recovery.

OBJECTIVES: To characterize the temporal pattern of a panel of blood and urinary biomarkers in an animal model of fecal peritonitis and recovery. DESIGN: Prospective observational animal study. SETTING: University research laboratory. SUBJECTS: Male Wistar rats. INTERVENTIONS: A fluid-resuscitated, long-term (3 d) rat model of sepsis (fecal peritonitis) and recovery was used to understand the temporal association of sepsis biomarkers in relation to systemic hemodynamics, inflammation, and renal function. At predefined time points (3, 6, 12, 24, 48, 72 hr), animals (≥ 6 per group) underwent echocardiography, blood and urine sampling, and had kidneys taken for histological analysis. Comparison was made against sham-operated controls and naïve animals. MEASUREMENTS AND MAIN RESULTS: The systemic proinflammatory response was maximal at 6 hours, corresponding with the nadir of stroke volume. Serum creatinine peaked late (24 hr), when clinical recovery was imminent. Histological evidence of tubular injury and cell death was minimal. After a recovery period, all biomarkers returned to levels approaching those observed in sham animals. Apart from urine clusterin and interleukin-18, all other urinary biomarkers were elevated at earlier time points compared with serum creatinine. Urine neutrophil gelatinase-associated lipocalin was the most sensitive marker among those studied, rising from 3 hours. While serum creatinine fell at 12 hours, serum cystatin C increased, suggestive of decreased creatinine production. CONCLUSIONS: Novel information is reported on the temporal profile of a panel of renal biomarkers in sepsis in the context of systemic and renal inflammation and recovery. Insight into the pathophysiology of acute kidney injury is gleaned from the temporal change markers of renal injury (urine neutrophil gelatinase-associated lipocalin, kidney injury molecule-1, calbindin), followed by a marker of cell cycle arrest (urine insulin-like growth factor-binding protein 7) and, finally, by functional markers of filtration (serum creatinine and cystatin C). These clinically relevant findings should have significant influence on future clinical testing.

Putative tissue location and function of the SLC5 family member SGLT3.

NEW FINDINGS: What is the topic of this review? This review summarizes the evidence on the localization, electrophysiological properties, agonist specificity and putative physiological role of sodium-glucose transporter 3 (SGLT3). What advances does it highlight? Published information is reviewed in some detail by comparing human and rodent isoforms, as well as advances in testing hypotheses for the physiological role of SGLT3 as a glucose sensor or incretin release mediator. We provide a critical overview of available published data and discuss a putative functional role for SGLT3 in human and mouse physiology. Sodium-glucose transporter 3 (SGLT3) has attracted interest because of its putative role as a glucose sensor, rather than a sugar transporter, in contrast to its co-family members SGLT1 and SGLT2. Significant progress has been made in characterizing the electrophysiological properties in vitro of the single human SGLT3 isoform and the two mouse isoforms, SGLT3a and SGLT3b. Although early reports indicated SGLT3 expression in the small intestinal myenteric and submucosal neurones, hypothalamic neurones, portal vein and kidney, a lack of reliable antibodies has left unanswered its exact tissue and cellular localization. Several hypotheses for a role of SGLT3 in glucose sensing, gastric emptying, glucagon-like peptide-1 release and post-Roux-en-Y gastric bypass remodelling have been explored, but so far there is only limited and indirect supportive evidence using non-specific agonists/antagonists, with no firm conclusions. There are no published or available data in knockout animals, and translation is difficult because of its different isoforms in human versus rodent, as well as a lack of selective agonists or antagonists, all of which make SGLT3 challenging to study. However, its unique electrophysiological properties, ubiquitous expression at the mRNA level, enrichment in the small intestine and potential, but uncertain, physiological role demand more attention. The purpose of this overview and review of SGLT3 biology is to provide an update, highlight the gaps in our knowledge and try to signpost potential ways forward to define its likely function in vivo.

Postprandial adjustments in renal phosphate excretion do not involve a gut-derived phosphaturic factor.

NEW FINDINGS: What is the central question of this study? Does a previously hypothesized signalling mechanism, believed to detect postprandial increases in intestinal phosphate and that can stimulate the kidneys to rapidly excrete phosphate, operate under physiological conditions? What is the main finding and its importance? Contrary to earlier reports, rapid signalling between the small intestine and kidney mediated by a gut-derived phosphaturic factor in response to a physiological intestinal phosphate load is not supported by the present findings; moreover, hyperphosphataemia and increased parathyroid hormone concentrations are likely to be the underlying factors responsible for the phosphaturia following a supraphysiological intestinal phosphate load. To date, the role of the small intestine in the regulation of postprandial phosphate homeostasis has remained unclear and controversial. Previous studies have proposed the presence of a gut-derived phosphaturic factor that acts independently of changes in plasma phosphate concentration or parathyroid hormone (PTH) concentration; however, these early studies used duodenal luminal phosphate concentrations in the molar range, and therefore, the physiological relevance of this is uncertain. In the present study, we used both in vivo and in vitro approaches to investigate the presence of this putative 'intestinal phosphatonin'. Instillation of 1.3 m phosphate into the duodenum rapidly induced phosphaturia, but in contrast to previous reports, this was associated with significant hyperphosphataemia and elevated PTH concentration; however, there was not the expected decrease in abundance of the renal sodium-phosphate cotransporter NaPi-IIa. Instillation of a physiological (10 mm) phosphate load had no effect on plasma phosphate concentration, PTH concentration or phosphate excretion. Moreover, phosphate uptake by opossum kidney cells was unaffected after incubation with serosal fluid collected from intestinal segments perfused with different concentrations of phosphate. Taken together, these findings do not support the concept of a gut-derived phosphaturic factor that can mediate rapid signalling between the gut and kidney, leading to increased urinary phosphate excretion, as part of normal phosphate homeostasis.

Experimental type II diabetes and related models of impaired glucose metabolism differentially regulate glucose transporters at the proximal tubule brush border membrane.

What is the central question of this study? Although SGLT2 inhibitors represent a promising treatment for patients suffering from diabetic nephropathy, the influence of metabolic disruption on the expression and function of glucose transporters is largely unknown. What is the main finding and its importance? In vivo models of metabolic disruption (Goto-Kakizaki type II diabetic rat and junk-food diet) demonstrate increased expression of SGLT1, SGLT2 and GLUT2 in the proximal tubule brush border. In the type II diabetic model, this is accompanied by increased SGLT- and GLUT-mediated glucose uptake. A fasted model of metabolic disruption (high-fat diet) demonstrated increased GLUT2 expression only. The differential alterations of glucose transporters in response to varying metabolic stress offer insight into the therapeutic value of inhibitors. SGLT2 inhibitors are now in clinical use to reduce hyperglycaemia in type II diabetes. However, renal glucose reabsorption across the brush border membrane (BBM) is not completely understood in diabetes. Increased consumption of a Western diet is strongly linked to type II diabetes. This study aimed to investigate the adaptations that occur in renal glucose transporters in response to experimental models of diet-induced insulin resistance. The study used Goto-Kakizaki type II diabetic rats and normal rats rendered insulin resistant using junk-food or high-fat diets. Levels of protein kinase C-ßI (PKC-ßI), GLUT2, SGLT1 and SGLT2 were determined by Western blotting of purified renal BBM. GLUT- and SGLT-mediated d-[(3) H]glucose uptake by BBM vesicles was measured in the presence and absence of the SGLT inhibitor phlorizin. GLUT- and SGLT-mediated glucose transport was elevated in type II diabetic rats, accompanied by increased expression of GLUT2, its upstream regulator PKC-ßI and SGLT1 protein. Junk-food and high-fat diet feeding also caused higher membrane expression of GLUT2 and its upstream regulator PKC-ßI. However, the junk-food diet also increased SGLT1 and SGLT2 levels at the proximal tubule BBM. Glucose reabsorption across the proximal tubule BBM, via GLUT2, SGLT1 and SGLT2, is not solely dependent on glycaemic status, but is also influenced by diet-induced changes in glucose metabolism. We conclude that different metabolic disturbances result in complex adaptations in renal glucose transporter protein levels and function.

What is nephrocalcinosis?

The available publications on nephrocalcinosis are wide-ranging and have documented multiple causes and associations of macroscopic or radiological nephrocalcinosis, most often located in the renal medulla, with various metabolic and genetic disorders; in fact, so many and various are these that it is difficult to define a common underlying mechanism. We have reviewed nephrocalcinosis in relation to its definition, genetic associations, animal models, and putative mechanisms. We have concluded, and hypothesized, that nephrocalcinosis is primarily a renal interstitial process, resembling metastatic calcification, and that it may have some features in common with, and pathogenic links to, vascular calcification.

Inhibition of the purinergic P2X7 receptor improves renal perfusion in angiotensin-II-infused rats.

Chronic activation of the renin-angiotensin system promotes hypertension, renal microvascular dysfunction, tissue hypoxia, and inflammation. Despite similar hypertension, an injurious response to excess angiotensin II is greater in F344 than in Lewis rats; the latter displaying renoprotection. Here we studied whether p2rx7, encoding the P2X7 receptor (P2X7R), is a candidate gene for the differential susceptibility to vascular dysfunction under high angiotensin II tone. A 14-day infusion of angiotensin II into F344 rats increased blood pressure by about 15 mm Hg without inducing fibrosis or albuminuria. In vivo pressure natriuresis was suppressed, medullary perfusion reduced by half, and the corticomedullary oxygenation gradient disrupted. Selective P2X7R antagonism restored pressure natriuresis, promoting a significant leftward shift in the intercept and increasing the slope. Sodium excretion was increased sixfold and blood pressure normalized. The specific P2X7R antagonist AZ11657312 increased renal medullary perfusion, but only in angiotensin II-treated rats. Tissue oxygenation was improved by P2X7R blockade, particularly in poorly oxygenated regions of the kidney. Thus, activation of P2X7R induces microvascular dysfunction and regional hypoxia when angiotensin II is elevated and these effects may contribute to progression of renal injury induced by chronic angiotensin II.

The urinary proteome and metabonome differ from normal in adults with mitochondrial disease.

We studied the extent and nature of renal involvement in a cohort of 117 adult patients with mitochondrial disease, by measuring urinary retinol-binding protein (RBP) and albumin; established markers of tubular and glomerular dysfunction, respectively. Seventy-five patients had the m.3243A>G mutation and the most frequent phenotypes within the entire cohort were 14 with MELAS, 33 with MIDD, and 17 with MERRF. Urinary RBP was increased in 29 of 75 of m.3243A>G patients, whereas albumin was increased in 23 of the 75. The corresponding numbers were 16 and 14, respectively, in the 42 non-m.3243A>G patients. RBP and albumin were higher in diabetic m.3243A>G patients than in nondiabetics, but there were no significant differences across the three major clinical phenotypes. The urine proteome (mass spectrometry) and metabonome (nuclear magnetic resonance) in a subset of the m.3243A>G patients were markedly different from controls, with the most significant alterations occurring in lysosomal proteins, calcium-binding proteins, and antioxidant defenses. Differences were also found between asymptomatic m.3243A>G carriers and controls. No patients had an elevated serum creatinine level, but 14% had hyponatremia, 10% had hypophosphatemia, and 14% had hypomagnesemia. Thus, abnormalities in kidney function are common in adults with mitochondrial disease, exist in the absence of elevated serum creatinine, and are not solely explained by diabetes.

Renal Fanconi syndrome: taking a proximal look at the nephron.

Renal Fanconi syndrome (RFS) refers to the generalized dysfunction of the proximal tubule (PT) (Kleta R. Fanconi or not Fanconi? Lowe syndrome revisited. Clin J Am Soc Nephrol 2008; 3: 1244-1245). In its isolated form, RFS only affects the PT, but not the other nephron segments. The study of isolated RFS can thus provide specific insights into the function of the PT. In a recent paper, Klootwijk et al. investigated one such form of isolated RFS and revealed the underlying molecular basis (Klootwijk ED, Reichold M, Helip-Wooley A et al. Mistargeting of peroxisomal EHHADH and inherited renal Fanconi's syndrome. N Engl J Med 2014; 370: 129-138). The affected family had been described previously, demonstrating the typical features of RFS, such as low-molecular weight proteinuria, aminoaciduria, glycosuria and phosphaturia with consequent rickets; yet, importantly, patients had no evidence of impaired glomerular filtration (Tolaymat A, Sakarcan A, Neiberger R. Idiopathic Fanconi syndrome in a family. Part I. Clinical aspects. J Am Soc Nephrol 1992; 2: 1310-1317). Inheritance was consistent with an autosomal dominant mode. Klootwijk et al. discovered a surprising explanation: a heterozygous missense mutation causing partial mistargeting of the peroxisomal enzyme EHHADH to the mitochondria. Notably, disease causing was not the absence of the enzyme in the peroxisome, but its interference with mitochondrial function. The discovery of this novel disease mechanism not only confirmed the importance of mitochondrial function for PT transport, but also demonstrated the critical dependence of PT on fatty acid metabolism for energy generation.

Exaggerated renal fibrosis in P2X4 receptor-deficient mice following unilateral ureteric obstruction.

BACKGROUND: The ATP-sensitive P2X7 receptor (P2X7R) has been shown to contribute to renal injury in nephrotoxic nephritis, a rodent model of acute glomerulonephritis, and in unilateral ureteric obstruction (UUO), a rodent model of chronic interstitial inflammation and fibrosis. Renal tubular cells, endothelial cells and macrophages also express the closely related P2X4 receptor (P2X4R), which is chromosomally co-located with P2X7R and has 40% homology; it is also pro-inflammatory and has been shown to interact with P2X7R to modulate its pro-apoptotic and pro-inflammatory effects. Therefore, we chose to explore the function of P2X4R in the UUO model of renal injury using knockout mice. We hypothesized that UUO-induced tubulointerstitial damage and fibrosis would also be attenuated in P2X4R(-/-) mice. METHOD: P2X4R(-/-) and wild-type (WT) mice were subjected to either UUO or sham operation. Kidney samples taken on Days 7 and 14 were evaluated for renal inflammation and fibrosis, and expression of pro-fibrotic factors. RESULTS: To our surprise, the obstructed kidney in P2X4R(-/-) mice showed more severe renal injury, more collagen deposition (picrosirius red staining, increase of 53%; P < 0.05) and more type I collagen staining (increase of 107%; P < 0.01), as well as increased mRNA for TGF-ß (increase of 102%, P < 0.0005) and CTGF (increase of 157%; P < 0.05) by Day 14, compared with the UUO WT mice. CONCLUSION: These findings showed that lack of P2X4R expression leads to increased renal fibrosis, and increased expression of TGF-ß and CTGF in the UUO model.