Epigenetics and autosomal dominant polycystic kidney disease

The roles of epigenetic modulation of gene expression and protein functions in autosomal dominant polycystic kidney disease (ADPKD) have recently become the focus of scientific investigation. Evidence generated to date indicates that one of the epigenetic modifiers, histone deacetylases (HDACs), are important regulators of ADPKD. HDACs are involved in regulating the expression of the Pkd1 gene and are the target of fluid flow-induced calcium signal in kidney epithelial cells. Pharmacological inhibition of HDAC activity has been found to reduce the progression of cyst formation and slow the decline of kidney function in Pkd1 conditional knockout mice and Pkd2 knockout mice, respectively, implicating the potential clinical application of HDAC inhibitors on ADPKD. Since the expression of HDAC6 is upregulated in cystic epithelial cells, the potential roles of HDAC6 in regulating cilia resorption and epidermal growth factor receptor (EGFR) trafficking through deacetylating α-tubulin and regulating Wnt signaling through deacetylating β-catenin are also discussed. This article is part of a Special Issue entitled: Polycystic Kidney Disease.     

TRPP2 and autosomal dominant polycystic kidney disease

Mutations in TRPP2 (polycystin-2) cause autosomal dominant polycystic kidney disease (ADPKD), a common genetic disorder characterized by progressive development of fluid-filled cysts in the kidney and other organs. TRPP2 is a Ca(2+)-permeable nonselective cation channel that displays an amazing functional versatility at the cellular level. It has been implicated in the regulation of diverse physiological functions including mechanosensation, cell proliferation, polarity, and apoptosis. TRPP2 localizes to different subcellular compartments, such as the endoplasmic reticulum (ER), the plasma membrane and the primary cilium. The channel appears to have distinct functions in different subcellular compartments. This functional compartmentalization is thought to contribute to the observed versatility and specificity of TRPP2-mediated Ca(2+) signaling. In the primary cilium, TRPP2 has been suggested to function as a mechanosensitive channel that detects fluid flow in the renal tubule lumen, supporting the proposed role of the primary cilium as the unifying pathogenic concept for cystic kidney disease. This review summarizes the known and emerging functions of TRPP2, focusing on the question of how channel function translates into complex morphogenetic programs regulating tubular structure.     

Urine proteome of autosomal dominant polycystic kidney disease patients

Background

Autosomal dominant polycystic kidney disease (ADPKD) is responsible for 10% of cases of the end stage renal disease. Early diagnosis, especially of potential fast progressors would be of benefit for efficient planning of therapy. Urine excreted proteome has become a promising field of the search for marker patterns of renal diseases including ADPKD. Up to now however, only the low molecular weight fraction of ADPKD proteomic fingerprint was studied. The aim of our study was to characterize the higher molecular weight fraction of urinary proteome of ADPKD population in comparison to healthy controls as a part of a general effort aiming at exhaustive characterization of human urine proteome in health and disease, preceding establishment of clinically useful disease marker panel.

Results

We have analyzed the protein composition of urine retentate (>10 kDa cutoff) from 30 ADPKD patients and an appropriate healthy control group by means of a gel-free relative quantitation of a set of more than 1400 proteins. We have identified an ADPKD-characteristic footprint of 155 proteins significantly up- or downrepresented in the urine of ADPKD patients. We have found changes in proteins of complement system, apolipoproteins, serpins, several growth factors in addition to known collagens and extracellular matrix components. For a subset of these proteins we have confirmed the results using an alternative analytical technique.

Conclusions

Obtained results provide basis for further characterization of pathomechanism underlying the observed differences and establishing the proteomic prognostic marker panel.     

Bilineal disease and trans-heterozygotes in autosomal dominant polycystic kidney disease

In searching for a putative third gene for autosomal dominant polycystic kidney disease (ADPKD), we studied the genetic inheritance of a large family (NFL10) previously excluded from linkage to both the PKD1 locus and the PKD2 locus. We screened 48 members of the NFL10 pedigree, by ultrasonography, and genotyped them, with informative markers, at both the PKD1 locus and the PKD2 locus. Twenty-eight of 48 individuals assessed were affected with ADPKD. Inspection of the haplotypes of these individuals suggested the possibility of bilineal disease from independently segregating PKD1 and PKD2 mutations. Using single-stranded conformational analysis, we screened for and found a PKD2 mutation (i.e., 2152delA; L736X) in 12 affected pedigree members. Additionally, when the disease status of these individuals was coded as "unknown" in linkage analysis, we also found, with markers at the PKD1 locus, significant LOD scores (i.e., >3.0). These findings strongly support the presence of a PKD1 mutation in 15 other affected pedigree members, who lack the PKD2 mutation. Two additional affected individuals had trans-heterozygous mutations involving both genes, and they had renal disease that was more severe than that in affected individuals who had either mutation alone. This is the first documentation of bilineal disease in ADPKD. In humans, trans-heterozygous mutations involving both PKD1 and PKD2 are not necessarily embryonically lethal. However, the disease associated with the presence of both mutations appears to be more severe than the disease associated with either mutation alone. The presence of bilineal disease as a confounder needs to be considered seriously in the search for the elusive PKD3 locus.     

Targeting chloride transport in autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) is the most frequent inherited kidney disease. Transepithelial fluid secretion is one of the key factors of cystogenesis in ADPKD. Multiple studies have suggested that fluid secretion across ADPKD cyst-lining cells is driven by the secretion of chloride, essentially mediated by the CFTR channel and stimulated by increased intracellular levels of 3′,5′-cyclic adenosine monophosphate. This review focuses on the pathophysiology of fluid secretion in ADPKD based on the pioneering studies of Jared Grantham and colleagues, and on the follow-up investigations from the molecular level to the potential applications in ADPKD patients. Altogether, the studies of fluid and chloride transport in ADPKD paved the way for innovative therapeutic targets to prevent cyst volume expansion and thus, kidney disease progression.     

Regional localization of the autosomal dominant polycystic kidney disease locus

The localization of the autosomal dominant polycystic kidney disease locus (PKD1) within an array of anonymous polymorphic DNA sequences on chromosome 16 band p13 was determined by multipoint mapping. Nine polymorphic DNA markers, including two hypervariable sequences, were used to study 19 PKD1 and 21 reference families. PKD1 was found to lie proximal to the 3' and 5' hypervariable regions of alpha-globin and distal to the anonymous sequence CRI-0327. Somatic cell hybrid mapping places PKD1 within the region 16p13.11-16pter. The availability of an array of linked markers which bracket the PKD1 locus provides a framework for further attempts to identify the PKD1 gene and offers an improved method of presymptomatic diagnosis of the disease.     

Fluid transport and cystogenesis in autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) is the most frequent inherited nephropathy. The development and enlargement of cysts in ADPKD requires tubular cell proliferation, abnormalities in the extracellular matrix and transepithelial fluid secretion. Multiple studies have suggested that fluid secretion across ADPKD cyst-lining cells is driven by the transepithelial secretion of chloride, mediated by the apical CFTR channel and specific basolateral transporters. The whole secretory process is stimulated by increased levels of cAMP in the cells, probably reflecting modifications in the intracellular calcium homeostasis and abnormal stimulation of the vasopressin V2 receptor. This review will focus on the pathophysiology of fluid secretion in ADPKD cysts, starting with classic, morphological and physiological studies that were followed by investigations of the molecular mechanisms involved and therapeutic trials targeting these pathways in cellular and animal models and ADPKD patients. This article is part of a Special Issue entitled: Polycystic Kidney Disease.     

Generation of c-Myc transgenic pigs for autosomal dominant polycystic kidney disease

After several decades of research, autosomal dominant polycystic kidney disease (ADPKD) is still incurable and imposes enormous physical, psychological, and economic burdens on patients and their families. Murine models of ADPKD represent invaluable tools for studying this disease. These murine forms of ADPKD can arise spontaneously, or they can be induced via chemical or genetic manipulations. Although these models have improved our understanding of the etiology and pathogenesis of ADPKD, they have not led to effective treatment strategies. The mini-pig represents an effective biomedical model for studying human diseases, as the pig’s human-like physiological processes help to understand disease mechanisms and to develop novel therapies. Here, we tried to generate a transgenic model of ADPKD in pigs by overexpressing c-Myc in kidney tissue. Western-blot analysis showed that c-Myc was overexpressed in the kidney, brain, heart, and liver of transgenic pigs. Immunohistochemical staining of kidney tissue showed that exogenous c-Myc predominantly localized to renal tubules. Slightly elevated blood urea nitrogen levels were observed in transgenic pigs 1 month after birth, but no obvious abnormalities were detected after that time. In the future, we plan to subject this model to renal injury in an effort to promote ADPKD progression.     

A tumor necrosis factor-alpha-mediated pathway promoting autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) is caused by heterozygous mutations in either PKD1 or PKD2, genes that encode polycystin-1 and polycystin-2, respectively. We show here that tumor necrosis factor-alpha (TNF-alpha), an inflammatory cytokine present in the cystic fluid of humans with ADPKD, disrupts the localization of polycystin-2 to the plasma membrane and primary cilia through a scaffold protein, FIP2, which is induced by TNF-alpha. Treatment of mouse embryonic kidney organ cultures with TNF-alpha resulted in formation of cysts, and this effect was exacerbated in the Pkd2(+/-) kidneys. TNF-alpha also stimulated cyst formation in vivo in Pkd2(+/-) mice. In contrast, treatment of Pkd2(+/-) mice with the TNF-alpha inhibitor etanercept prevented cyst formation. These data reveal a pathway connecting TNF-alpha signaling, polycystins and cystogenesis, the activation of which may reduce functional polycystin-2 below a critical threshold, precipitating the ADPKD cellular phenotype.     

Berberine slows cell growth in autosomal dominant polycystic kidney disease cells

Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary monogenic disorder characterized by development and enlargement of kidney cysts that lead to loss of renal function. It is caused by mutations in two genes (PKD1 and PKD2) encoding for polycystin-1 and polycystin-2 proteins which regulate different signals including cAMP, mTOR and EGFR pathways. Abnormal activation of these signals following PC1 or PC2 loss of function causes an increased cell proliferation which is a typical hallmark of this disease. Despite the promising findings obtained in animal models with targeted inhibitors able to reduce cystic cell growth, currently, no specific approved therapy for ADPKD is available. Therefore, the research of new more effective molecules could be crucial for the treatment of this severe pathology. In this regard, we have studied the effect of berberine, an isoquinoline quaternary alkaloid, on cell proliferation and apoptosis in human and mouse ADPKD cystic cell lines. Berberine treatment slows cell proliferation of ADPKD cystic cells in a dose-dependent manner and at high doses (100 μg/mL) it induces cell death in cystic cells as well as in normal kidney tubule cells. However, at 10 μg/mL, berberine reduces cell growth in ADPKD cystic cells only enhancing G₀/G₁ phase of cell cycle and inhibiting ERK and p70-S6 kinases. Our results indicate that berberine shows a selected antiproliferative activity in cellular models for ADPKD, suggesting that this molecule and similar natural compounds could open new opportunities for the therapy of ADPKD patients.     

Effective treatment of an orthologous model of autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) is a leading cause of end-stage renal disease. The vasopressin V2 receptor (VPV2R) antagonist OPC31260 has been effective in two animal models of PKD with pathologies that are probably related. Here we show, in a mouse model of ADPKD (Pkd2(-/tm1Som)), a similar cellular phenotype and response to OPC31260 treatment, with reduction of renal cyclic AMP (cAMP) levels, prevention of renal enlargement, marked inhibition of cystogenesis and protection of renal function.     

Role of endothelin and nitric oxide in the pathogenesis of arterial hypertension in autosomal dominant polycystic kidney disease

The pathogenesis of arterial hypertension in autosomal dominant polycystic kidney disease (ADPKD) is complex and likely dependent on interaction of hemodynamic, endocrine and neurogenic factors. We decided to evaluate the role of endothelin (ET1) and nitric oxide (NO) in the regulation of arterial blood pressure (BP) and to determine plasma levels of ET1 and NO in the group of patients with ADPKD. The ADPKD group (18 patients, 6 men + 12 women, mean age 44.6+/-11.7 years, with creatinine clearancecorrig > 1.1 ml/s) was compared with a control group of 27 healthy volunteers of comparable age. Plasma levels of ET1 assessed by direct RIA determination in the group of ADPKD patients (11.03+/-1.8 fmol/ml) were significantly increased (p<0.001) in comparison with the control group (2.66+/-0.58 fmol/ml), while no significant differences were observed between normotensive and hypertensive patients in the ADPKD group. Serum levels of NO were evaluated according to the determination of serum levels of their metabolites - nitrites/nitrates. Serum levels of NO in the group of ADPKD patients (39.85+/-.38 micro mol/l) were significantly higher (p<0.05) in comparison with the control group (22.7+/-1.20 micro mol/l), whereas in the ADPKD group no significant differences were observed between normotensive and hypertensive patients. Thus, our study supports the concept of complex alteration of both vasoconstrictor and vasodilator systems in the pathogenesis of arterial hypertension in ADPKD.     

Compromised cytoarchitecture and polarized trafficking in autosomal dominant polycystic kidney disease cells

Cystogenesis associated with autosomal dominant polycystic kidney disease (ADPKD) is characterized by perturbations in the polarized phenotype and function of cyst-lining epithelial cells. The polycystins, the protein products of the genes mutated in the majority of ADPKD cases, have been described recently, but the pathological mechanism by which causal mutations result in the mislocalization of cell membrane proteins has remained unclear. This report documents the dissociation from the ADPKD cell basolateral membrane of three molecules essential for spatial organization and exocytosis. The adherens junction protein E-cadherin, the subcellular disposition of which governs intercellular and intracellular architecture, was discovered sequestered in an internal ADPKD cell compartment. At the same time, sec6 and sec8, components of a complex critical for basolateral cargo delivery normally arrayed at the apico-lateral apex, were depleted from the ADPKD cell plasma membrane. An analysis of membrane transport revealed that basolateral trafficking of proteins and lipids was impaired as a result of delayed cargo exit from the ADPKD cell Golgi apparatus. Apical transport proceeded normally. Taken together with recent documentation of an association between polycystin-1 and E-cadherin (Huan and van Adelsberg 1999), the data suggest that causal mutations disrupt E-cadherin-dependent cytoarchitecture, adversely affecting protein assemblies crucial for basolateral trafficking.     

Autosomal dominant polycystic kidney disease: molecular genetics and molecular pathogenesis

Mutations in three different genes, PKD1, PKD2 and PKD3, can cause a very similar clinical picture of the autosomal dominant form of polycystic kidney disease (ADPKD). Apparently, mutations in the PKD3 gene, which is still unmapped, are very rare, whereas PKD1 defects account for about 85% of cases. Although ADPKD is a frequent monogenic disorder affecting approximately 1:1000 individuals in the Caucasian population, progress in understanding its pathology was somewhat slow until relatively recently when the PKD1 and PKD2 genes were mapped and cloned. They are both large, being approximately 52 kb and 68 kb in length respectively, and in addition, PKD1 is fairly complex, thus complicating mutation detection. The gene products, polycystin-1 and polycystin-2, are trans-membranous glycoproteins and are considered to be involved in signalling pathways, in cooperation with additional partners. Immunostaining studies in both humans and mice have revealed information regarding the localization of polycystins and their role in the development and maintenance of nephrons. Recent experimentation from various laboratories has shown that loss of heterozygosity and acquired somatic second hits may account, at least partly, for the inter- and intrafamilial phenotypic heterogeneity of the disease, while at the same time, the existence of other modifying loci is also hypothesized. The two-hit hypothesis is admittedly a very attractive one in that it can explain many of the features of the disease, whereas recent data regarding a trains-heterozygous model for cystogenesis adds to the complexity of the molecular mechanisms that can lead to pathogenesis.     

Mutations for the autosomal recessive and autosomal dominant forms of polycystic kidney disease are not allelic

Summary The autosomal dominant form of polycystic kidney disease (ADPKD) has been linked to the -globin gene locus on 16p. Linkage studies between the autosomal recessive type (ARPKD) and the 3 HVR of the -globin gene cluster showed that the ARPKD and ADPKD are not allelic.     

Contribution of renal innervation to hypertension in rat autosomal dominant polycystic kidney disease

The kidney has both afferent (sensory) and efferent (sympathetic) nerves that can influence renal function. Renal innervation has been shown to play a role in the pathogenesis of many forms of hypertension. Hypertension and flank pain are common clinical manifestations of autosomal dominant (AD) polycystic kidney disease (PKD). We hypothesize that renal innervation contributes to the hypertension and progression of cystic change in rodent PKD. In the present study, the contribution of renal innervation to hypertension and progression of renal histopathology and dysfunction was assessed in male Han:SPRD-Cy/+ rats with ADPKD. At 4 weeks of age, male offspring from crosses of heterozygotes (Cy/+) were randomized into either 1) bilateral surgical renal denervation, 2) surgical sham denervation control, or 3) nonoperated control groups. A midline laparotomy was performed to allow the renal denervation (i.e., physical stripping of the nerves and painting the artery with phenol/alcohol). Blood pressure (tail cuff method), renal function (BUN) and histology were assessed at 8 weeks of age. Bilateral renal denervation reduced the cystic kidney size, cyst volume density, systolic blood pressure, and improved renal function (BUN) as compared with nonoperated controls. Operated control cystic rats had kidney weights, cyst volume densities, systolic blood pressures, and plasma BUN levels that were intermediate between those in the denervated animals and the nonoperated controls. The denervated group had a reduced systolic blood pressure compared with the operated control animals, indicating that the renal innervations was a major contributor to the hypertension in this model of ADPKD. Renal denervation was efficacious in reducing some pathology, including hypertension, renal enlargement, and cystic pathology. However, sham operation also affected the cystic disease but to a lesser extent. We hypothesize that the amelioration of hypertension in Cy/+ rats was due to the effects of renal denervation on the renin angiotensin system.     

A transducin-like gene maps to the autosomal dominant polycystic kidney disease gene region

A novel human gene (sazD) that maps to the autosomaldominant polycystic kidney disease region shares sequence similarity with members of the β-transducin superfamily. The cDNA sazD-c predicts an 58-kDa protein (sazD) with seven internal repeats, similar to the WD-40 motif of the transducin family. The size of this protein family has been expanding rapidly; however, neither the structure nor the function of this repeated motif is known. Preliminary data do not suggest that sazD is mutated in patients with polycystic kidney disease.