New insights into the role of HNF-1ß in kidney (patho)physiology.

Hepatocyte nuclear factor-1ß (HNF-1ß) is an essential transcription factor that regulates the development and function of epithelia in the kidney, liver, pancreas, and genitourinary tract. Humans who carry HNF1B mutations develop heterogeneous renal abnormalities, including multicystic dysplastic kidneys, glomerulocystic kidney disease, renal agenesis, renal hypoplasia, and renal interstitial fibrosis. In the embryonic kidney, HNF-1ß is required for ureteric bud branching, initiation of nephrogenesis, and nephron segmentation. Ablation of mouse Hnf1b in nephron progenitors causes defective tubulogenesis, whereas later inactivation in elongating tubules leads to cyst formation due to downregulation of cystic disease genes, including Umod, Pkhd1, and Pkd2. In the adult kidney, HNF-1ß controls the expression of genes required for intrarenal metabolism and solute transport by tubular epithelial cells. Tubular abnormalities observed in HNF-1ß nephropathy include hyperuricemia with or without gout, hypokalemia, hypomagnesemia, and polyuria. Recent studies have identified novel post-transcriptional and post-translational regulatory mechanisms that control HNF-1ß expression and activity, including the miRNA cluster miR17 ∼ 92 and the interacting proteins PCBD1 and zyxin. Further understanding of the molecular mechanisms upstream and downstream of HNF-1ß may lead to the development of new therapeutic approaches in cystic kidney disease and other HNF1B-related renal diseases.

Tisp40 deficiency attenuates renal ischemia reperfusion injury induced apoptosis of tubular epithelial cells.

Renal ischemia reperfusion (IR) is a major cause of acute kidney injury (AKI) and no effective treatments have been established. Tisp40 is a transcription factor of the CREB/ATF family and involves in cell apoptosis, proliferation and differentiation, but its role in renal IR remains unknown. Here, we investigated the role of Tisp40 in renal IR injury. In vivo, Tisp40 knockout (KO) and wild-type (WT) mice were subjected to thirty minutes of bilateral renal ischemia and 48h reperfusion, the blood and kidneys were harvested for analysis. In vitro, Tisp40 overexpression and vector cells were subjected to hypoxia/reoxygenation (HR), the apoptosis rate and the expressions of related proteins were measured. Following IR, the expressions of Tisp40 protein, serum creatinine (sCr), blood urea nitrogen (BUN) and apoptosis of tubular cells were significantly increased in WT mice. However, Tisp40 deficiency significantly attenuated the increase of sCr, BUN and apoptosis of tubular cells. Following HR, apoptosis of tubular cells was increased in Tisp40 overexpression cells compared with vector cells. Mechanistically, Tisp40 promoted the expressions of C/EBP homologous protein (CHOP), Bax and Cleaved caspase3 and suppressed the expression of Bcl-2 in renal IR injury. In conclusion, Tisp40 aggravates tubular cells apoptosis in renal IR injury.

Impairment of Wnt11 function leads to kidney tubular abnormalities and secondary glomerular cystogenesis.

BACKGROUND: Wnt11 is a member of the Wnt family of secreted signals controlling the early steps in ureteric bud (UB) branching. Due to the reported lethality of Wnt11 knockout embryos in utero, its role in later mammalian kidney organogenesis remains open. The presence of Wnt11 in the emerging tubular system suggests that it may have certain roles later in the development of the epithelial ductal system. RESULTS: The Wnt11 knockout allele was backcrossed with the C57Bl6 strain for several generations to address possible differences in penetrance of the kidney phenotypes. Strikingly, around one third of the null mice with this inbred background survived to the postnatal stages. Many of them also reached adulthood, but urine and plasma analyses pointed out to compromised kidney function. Consistent with these data the tubules of the C57Bl6 Wnt11 (-/-) mice appeared to be enlarged, and the optical projection tomography indicated changes in tubular convolution. Moreover, the C57Bl6 Wnt11 (-/-) mice developed secondary glomerular cysts not observed in the controls. The failure of Wnt11 signaling reduced the expression of several genes implicated in kidney development, such as Wnt9b, Six2, Foxd1 and Hox10. Also Dvl2, an important PCP pathway component, was downregulated by more than 90 % due to Wnt11 deficiency in both the E16.5 and NB kidneys. Since all these genes take part in the control of UB, nephron and stromal progenitor cell differentiation, their disrupted expression may contribute to the observed anomalies in the kidney tubular system caused by Wnt11 deficiency. CONCLUSIONS: The Wnt11 signal has roles at the later stages of kidney development, namely in coordinating the development of the tubular system. The C57Bl6 Wnt11 (-/-) mouse generated here provides a model for studying the mechanisms behind tubular anomalies and glomerular cyst formation.

A Point Mutation in p190A RhoGAP Affects Ciliogenesis and Leads to Glomerulocystic Kidney Defects.

Rho family GTPases act as molecular switches regulating actin cytoskeleton dynamics. Attenuation of their signaling capacity is provided by GTPase-activating proteins (GAPs), including p190A, that promote the intrinsic GTPase activity of Rho proteins. In the current study we have performed a small-scale ENU mutagenesis screen and identified a novel loss of function allele of the p190A gene Arhgap35, which introduces a Leu1396 to Gln substitution in the GAP domain. This results in decreased GAP activity for the prototypical Rho-family members, RhoA and Rac1, likely due to disrupted ordering of the Rho binding surface. Consequently, Arhgap35-deficient animals exhibit hypoplastic and glomerulocystic kidneys. Investigation into the cystic phenotype shows that p190A is required for appropriate primary cilium formation in renal nephrons. P190A specifically localizes to the base of the cilia to permit axoneme elongation, which requires a functional GAP domain. Pharmacological manipulations further reveal that inhibition of either Rho kinase (ROCK) or F-actin polymerization is able to rescue the ciliogenesis defects observed upon loss of p190A activity. We propose a model in which p190A acts as a modulator of Rho GTPases in a localized area around the cilia to permit the dynamic actin rearrangement required for cilia elongation. Together, our results establish an unexpected link between Rho GTPase regulation, ciliogenesis and glomerulocystic kidney disease.

Fetal renin-angiotensin-system blockade syndrome: renal lesions.

BACKGROUND: Fetuses exposed to angiotensin-converting enzyme inhibitors or angiotensin receptor antagonists during the second and/or third trimesters of gestation are at high risk of developing severe complications. They consist in fetal hypotension, and anuria/oligohydramnios leading to Potter sequence, frequently associated with hypocalvaria. Most fetuses die during the pre- or postnatal period, whereas others recover normal or subnormal renal function. However, the secondary occurrence of renal failure or hypertension has been reported in children after apparent complete recovery. METHODS: In this context, we analyzed renal lesions in 14 fetus/neonates who died soon after exposure to renin-angiotensin-system (RAS) blockers. Our objective was to determine the causes for the persistence or the secondary occurrence of renal complications reported in some of the survivors. RESULTS: As previously described, renal tubular dysgenesis is usually observed. Additional lesions, such as thickening of the muscular wall of arterioles and interlobular arteries, glomerular cysts, and interstitial fibrosis, develop early during fetal life. CONCLUSION: We suggest that renal lesions that develop before birth may persist after withdrawal of the causative drugs and normalization of blood and renal perfusion pressure. Their persistence could explain the severe long-term outcome of some of these patients. Long-term study of children exposed to RAS blockers during fetal life is strongly recommended.

Leucine-rich repeat kinase 2 (LRRK2)-deficient rats exhibit renal tubule injury and perturbations in metabolic and immunological homeostasis.

Genetic evidence links mutations in the LRRK2 gene with an increased risk of Parkinson's disease, for which no neuroprotective or neurorestorative therapies currently exist. While the role of LRRK2 in normal cellular function has yet to be fully described, evidence suggests involvement with immune and kidney functions. A comparative study of LRRK2-deficient and wild type rats investigated the influence that this gene has on the phenotype of these rats. Significant weight gain in the LRRK2 null rats was observed and was accompanied by significant increases in insulin and insulin-like growth factors. Additionally, LRRK2-deficient rats displayed kidney morphological and histopathological alterations in the renal tubule epithelial cells of all animals assessed. These perturbations in renal morphology were accompanied by significant decreases of lipocalin-2, in both the urine and plasma of knockout animals. Significant alterations in the cellular composition of the spleen between LRRK2 knockout and wild type animals were identified by immunophenotyping and were associated with subtle differences in response to dual infection with rat-adapted influenza virus (RAIV) and Streptococcus pneumoniae. Ontological pathway analysis of LRRK2 across metabolic and kidney processes and pathological categories suggested that the thioredoxin network may play a role in perturbing these organ systems. The phenotype of the LRRK2 null rat is suggestive of a complex biology influencing metabolism, immune function and kidney homeostasis. These data need to be extended to better understand the role of the kinase domain or other biological functions of the gene to better inform the development of pharmacological inhibitors.

Progesterone induced mesenchymal differentiation and rescued cystic dilation of renal tubules of Pkd1(-/-) mice.

Autosomal dominant polycystic kidney disease (ADPKD), the most common hereditary disease affecting the kidneys, is caused in 85% of cases by mutations in the PKD1 gene. The protein encoded by this gene, polycystin-1, is a renal epithelial cell membrane mechanoreceptor, sensing morphogenetic cues in the extracellular environment, which regulate the tissue architecture and differentiation. However, how such mutations result in the formation of cysts is still unclear. We performed a precise characterization of mesenchymal differentiation using PAX2, WNT4 and WT1 as a marker, which revealed that impairment of the differentiation process preceded the development of cysts in Pkd1(-/-) mice. We performed an in vitro organ culture and found that progesterone and a derivative thereof facilitated mesenchymal differentiation, and partially prevented the formation of cysts in Pkd1(-/-) kidneys. An injection of progesterone or this derivative into the intraperitoneal space of pregnant females also improved the survival of Pkd1(-/-) embryos. Our findings suggest that compounds which enhance mesenchymal differentiation in the nephrogenesis might be useful for the therapeutic approach to prevent the formation of cysts in ADPKD patients.

Renal tubular dysgenesis and tubulointerstitial nephritis antigen in juvenile nephronophthisis.

AIM: The relationship between abnormalities of tubular architecture and tubulointerstitial nephritis antigen (TIN-ag) in juvenile nephronophthisis (J-NPH) was evaluated. METHODS: Sixteen J-NPH patients were examined. Nephrocystin-1, TIN-ag, type IV collagen, Fas antigen and the C5b-9 complement complex were stained by immunohistochemical methods. RESULTS: Renal tubules of patients with J-NPH showed morphological abnormalities of tubular basement membranes (TBM) and frequent apoptosis of tubular epithelial cells. Additionally, the C5b-9 complement complex was deposited within the TBM in the absence of immunoglobulin deposition, suggesting complement-dependent TBM injury. Localization of TIN-ag in the TBM of J-NPH patients disclosed a partial defect or discontinuity in 14 of the 16 patients, while type IV collagen immunoreactivity was relatively preserved. These findings suggest that tubulogenesis is disturbed during nephronogenesis in J-NPH patients because of a defect in nephrocystin, an NPHP gene product. TBM defects induce further morphological abnormalities such as cystic dilation of tubules; as tubular function impairment advances, the incomplete tubules may be injured by C5b-9 complement complexes, followed by apoptotic cell death. CONCLUSION: TIN-ag, which is important in early nephrogenesis, lacks normal activity, and vulnerable and incomplete tubules with deficient TIN-ag expression are formed. Removal of these defective tubules by apoptosis combined with the C5b-9 complement complex could be the primary reason for progression to end-stage renal disease in J-NPH patients.

Deficiency in Six2 during prenatal development is associated with reduced nephron number, chronic renal failure, and hypertension in Br/+ adult mice.

The Br/+ mutant mouse displays decreased embryological expression of the homeobox transcription factor Six2, resulting in hertitable renal hypoplasia. The purpose of this study was to characterize the renal physiological consequences of embryonic haploinsuffiency of Six2 by analyzing renal morphology and function in the adult Br heterozygous mutant. Adult Br/+ kidneys weighed 50% less than those from wild-type mice and displayed glomerulopathy. Stereological analysis of renal glomeruli showed that Br/+ kidneys had an average of 88% fewer glomeruli than +/+ kidneys, whereas individual glomeruli in Br/+ mice maintained an average volume increase of 180% compared with normal nephrons. Immunostaining revealed increased levels of endothelin-1 (ET-1), endothelin receptors A (ET(A)) and B (ET(B)), and Na-K-ATPase were present in the dilated renal tubules of mutant mice. Physiological features of chronic renal failure (CRF) including elevated mean arterial pressure, increased plasma creatinine, and dilute urine excretion were measured in Br/+ mutant mice. Electron microscopy of the Br/+ glomeruli revealed pathological alterations such as hypercellularity, extracellular matrix accumulation, and a thick irregular glomerular basement membrane. These results indicate that adult Br/+ mice suffer from CRF associated with reduced nephron number and renal hypoplasia, as well as glomerulopathy. Defects are associated with embryological deficiencies of Six2, suggesting that proper levels of this protein during nephrogenesis are critical for normal glomerular development and adult renal function.

Inherited renal tubular dysgenesis: the first patients surviving the neonatal period.

Renal tubular dysgenesis (RTD) is a clinical disorder either acquired during fetal development or inherited as an autosomal recessive condition. Inherited RTD is caused by mutations in the genes encoding the components of the renin-angiotensin system angiotensinogen, renin, angiotensin-converting enzyme and angiotensin II receptor type 1. Inherited RTD is characterized by early onset oligohydramnios, skull ossification defects, preterm birth and neonatal pulmonary and renal failure. The histological hallmark is the absence or poor development of proximal tubules. So far, all patients died either in utero or shortly after birth. We report the first patients with inherited RTD surviving the neonatal period and still being alive. Genetic and functional analysis of the renin-angiotensin system contributes to the diagnosis of RTD. In conclusion, the clinical diagnosis of inherited RTD is easily missed after birth without renal biopsy or information on affected family members. Genetic and functional analysis of the renin-angiotensin system contributes to correct diagnosis.

Andersen-Tawil syndrome: new potassium channel mutations and possible phenotypic variation.

OBJECTIVE: To evaluate clinical, genetic, and electrophysiologic features of patients with Andersen-Tawil syndrome (ATS) in the United Kingdom. METHODS: Clinical and neurophysiologic evaluation was conducted of 11 families suspected to have ATS. Molecular genetic analysis of each proband was performed by direct DNA sequencing of the entire coding region of KCNJ2. Control samples were screened by direct DNA sequencing. The electrophysiologic consequences of several new mutations were studied in an oocyte expression system. RESULTS: All 11 ATS families harbored pathogenic mutations in KCNJ2 with six mutations not previously reported. Some unusual clinical features including renal tubular defect, CNS involvement, and dental and phonation abnormalities were observed. Five mutations (T75M, D78G, R82Q, L217P, and G300D) were expressed, all of which resulted in nonfunctional channels when expressed alone, and co-expression with wild-type (WT) KCNJ2 demonstrated a dominant negative effect. CONCLUSION: Six new disease-causing mutations in KCNJ2 were identified, one of which was in a PIP2 binding site. Molecular expression studies indicated that five of the mutations exerted a dominant negative effect on the wild-type allele. KCNJ2 mutations are an important cause of ATS in the UK.

Fish mesonephric model of polycystic kidney disease in medaka (Oryzias latipes) pc mutant.

BACKGROUND: Polycystic kidney disease (PKD) is a common hereditary disease. A number of murine and zebrafish mutants have been generated and used for the study of PKD as metanephric and pronephric models, respectively. Here, we report a medaka (Oryzias latipes) mutant that develops numerous cysts in the kidney in adulthood fish in an autosomal-recessive manner as a mesonephric model of PKD. METHODS: The phenotypes of the medaka pc mutant were described in terms of morphologic, histologic, and ultrastructural features. The pc see-through stock was produced by crossing a pc mutant and a fish from the see-through stock and used for observing the kidney through the transparent body wall of a live fish. RESULTS: The mutant developed bilateral massive enlargement of the kidney in adulthood. They sexually matured normally within 2 months of age and died within 6 months of age. The affected kidney was occupied by numerous, fluid-filled cysts, which were lined by attenuated squamous epithelial cells. Developmentally, cystic formation began in the pronephros in 10-day-old fry and in the mesonephros in 20-day-old fry at the microscopic level. The pc see-through stock was useful in observing disease progression in live fish. CONCLUSION: The kidney disorder that develops in the medaka pc mutant is a mesonephric counterpart of PKD, particularly an autosomal-dominant PKD, based on its morphologic, histologic, and ultrastructural features, and slow progression.

Mode of action: oxalate crystal-induced renal tubule degeneration and glycolic acid-induced dysmorphogenesis--renal and developmental effects of ethylene glycol.

Ethylene glycol can cause both renal and developmental toxicity, with metabolism playing a key role in the mode of action (MOA) for each form of toxicity. Renal toxicity is ascribed to the terminal metabolite oxalic acid, which precipitates in the kidney in the form of calcium oxalate crystals and is believed to cause physical damage to the renal tubules. The human relevance of the renal toxicity of ethylene glycol is indicated by the similarity between animals and humans of metabolic pathways, the observation of renal oxalate crystals in toxicity studies in experimental animals and human poisonings, and cases of human kidney and bladder stones related to dietary oxalates and oxalate precursors. High-dose gavage exposures to ethylene glycol also cause axial skeletal defects in rodents (but not rabbits), with the intermediary metabolite, glycolic acid, identified as the causative agent. However, the mechanism by which glycolic acid perturbs development has not been investigated sufficiently to develop a plausible hypothesis of mode of action, nor have any cases of ethylene glycol-induced developmental effects been reported in humans. Given this, and the variations in sensitivity between animal species in response, the relevance to humans of ethylene glycol-induced developmental toxicity in animals is unknown at this time.

Renal tubular dysgenesis and neonatal hemochromatosis without pulmonary hypoplasia.

Renal tubular dysgenesis is a rare disorder of differentiation of the fetal kidney. The condition has previously been reported as a postmortem diagnosis in infants who have had oligohydramnios commencing after 20 weeks gestation and have died of renal or respiratory failure shortly after birth with a clinical description of Potter sequence. The absence of clinically significant pulmonary hypoplasia in our case serves to emphasize that renal tubular dysgenesis, fetal anuria and long-standing oligohydramnios can occur without pulmonary insufficiency. The coexistence of renal tubular dysgenesis with neonatal hemochromatosis has been previously described in four published cases. The link between these two rare conditions is clinically important if dialysis or liver transplantation is considered in infants with hepatic and renal failure. Antemortem diagnosis by renal biopsy in our case enabled parental counseling and avoided the inappropriate use of peritoneal dialysis.

Renal tubular dysgenesis-a case presentation.

Renal tubular dysgenesis (RTD) is a lethal, developmental anomaly of the fetal kidney characterized by a defect in differentiation of the proximal and distal convoluted tubules. It is usually associated with oligohydramnios in later pregnancy and Potter's syndrome. A neonate with typical features who presented with mild respiratory distress, dysmorphic appearance and anuria is described. At the age of seven days, peritoneal dialysis was started and was continued until the death of the baby at the age of three months. The diagnosis was made on the bases of clinical and ultrasonographic findings confirmed by renal biopsy. A review of the literature showed that this is the first case of RTD reported in Turkey.

Perturbed medullary tubulogenesis in neonatal rat exposed to renin-angiotensin system inhibition.

BACKGROUND: Pharmacological interruption of the angiotensin II type-1 receptor (AT(1)) signalling during nephrogenesis in rats induces irreversible abnormalities in kidney morphology, comprising papillary atrophy and tubulointerstitial damage, which are characterized by tubular dilatation/atrophy and interstitial inflammation/fibrosis. This study determined the time course for development of tubular structural and inflammatory changes and possible cytokine production in the renal medulla of newborn rats exposed to angiotensin-converting enzyme (ACE) inhibition. Additionally, medullary expression of E-cadherin, a marker for tubular formation, was investigated in ACE-inhibited rats. METHODS: Newborn rats were exposed (postnatal days 0-12) to ACE inhibitor enalapril and killed at days 1, 2, 4, 9 and 13. One kidney was used for morphological evaluation and the other for immunohistochemistry, using antibodies directed against monocytes/macrophages, T cells and E-cadherin on frozen sections. In a separate experiment, rats were treated for 9 days and had their kidneys processed for western immunoblot and immunohistochemistry, where antibodies directed against monocyte chemoattractant protein-1 (MCP-1) and tumour necrosis factor-alpha (TNF-alpha) were used on paraffin sections. RESULTS: In renal medulla from enalapril-treated rats, volume fractions of tubular lumens and interstitium were increased from postnatal days 2 and 4, respectively, while that of tubular cells was decreased from 4 days of age. Concomitant loss and/or reduction in E-cadherin expression (from day 2) was observed in dilated medullary tubules of enalapril-treated rats. Furthermore, in the medulla of enalapril-treated rats, the increased number of ED2+ (resident macrophages) cells, followed by the increase in ED1+ (monocytes/macrophages) and CD4+ T cells, was observed at days 9 and 13, respectively. This was accompanied by increased medullary expression of TNF-alpha at day 9. CONCLUSIONS: Neonatal ACE inhibition perturbs medullary tubulogenesis, as indicated by tubular dilatation and a lack of E-cadherin expression in these tubules. Macrophage/monocyte-mediated immune response is a secondary event, coincidentally associated with the up-regulation of TNF-alpha.

The Rho family of small GTPases is involved in epithelial cystogenesis and tubulogenesis.

BACKGROUND: Epithelial cyst and tubule formation represent critical processes for the development of many mammalian organs and involve transient, highly choreographed changes in cell polarity. The Rho family of small GTPases, whose prototypes are RhoA, Rac1, and Cdc42, regulate many biologic processes, including cell polarization and morphogenesis. The exocyst is a conserved eight-subunit protein complex involved in the biogenesis of polarity; in yeast, it is a downstream effector for several Rho family proteins, and, in mammals, plays a central role in cystogenesis and tubulogenesis. METHODS: Inducible cell lines expressing mutant forms of RhoA, Rac1, and Cdc42 and an in vitro model of cystogenesis and tubulogenesis were used to examine the effects of Rho family proteins on cyst and tubule formation. A series of pulse-chase assays, using basolateral, apical, and secretory proteins, were performed to examine the synthesis and membrane trafficking profile of the various Rho family mutant proteins. RESULTS: We show that expression of mutant RhoA, Rac1, and Cdc42 proteins all result in abnormal cyst and tubule formation. Furthermore, with respect to cystogenesis and tubulogenesis, the phenotypic effects of expressing each mutant Rho family protein are different. Specifically, cyst and, therefore, tubule formation is completely inhibited in the presence of constitutively active RhoA and tubulogenesis is inhibited in the presence of dominant negative Rac1. Reversal of cyst polarity is seen in the presence of dominant negative RhoA, dominant negative Rac1, and both dominant negative and constitutively active Cdc42. The series of synthesis and delivery assays, using basolateral, apical, and secretory proteins, revealed that Rho family mutant proteins display an exocyst-like trafficking profile. CONCLUSION: The differential effects suggest that RhoA, Rac1, and Cdc42 all act to control cyst and tubule formation and may act in concert to control these higher-order processes. The exocyst-like membrane trafficking profile displayed by the Rho family mutant proteins raises the possibility that Rho family proteins interact, either directly or indirectly, with the exocyst to control cyst and tubule formation.

Congenital nephrosis of the Finnish type: overview of placental pathology and literature review.

Congenital nephrosis of the Finnish type (CNF) is a rare, autosomal recessive disorder of glomerular filtration that results in massive proteinuria, edema, and ascites. Although previous studies describe the classic renal lesions characterizing this disorder, there are few documenting in detail the associated placental alterations. In this context, we present a case of CNF with emphasis on the placental pathology and compare our findings to what has been previously reported in the literature. A 36-year-old G2P1 with no significant medical history developed persistently elevated amniotic fluid alpha-fetoprotein in the absence of neural tube defects. Because of a clinical suspicion of CNF, she electively terminated the pregnancy at 19 weeks. Postmortem examination revealed characteristic renal changes, confirmed by electron microscopy, as well as significant placental villous edema. Although the placenta was not enlarged, the villi appeared profoundly hydropic. Extensive cystic vacuolar change was documented in both stem villi and tertiary villi, affecting 95% of the villi present. Since the fetus was not grossly edematous, the placental findings may represent the first sign of systemic hypoproteinemia.