Biomedical subjects

Peter C Harris

Publications and source records attributed to Peter C Harris.

At least 37 records · Page 2Linked to original sources

PubMed2004

A polycystin-1 multiprotein complex is disrupted in polycystic kidney disease cells.

Autosomal dominant polycystic kidney disease (ADPKD) is typified by the accumulation of fluid-filled cysts and abnormalities in renal epithelial cell function. The disease is principally caused by mutations in the gene encoding polycystin-1, a large basolateral plasma membrane protein expressed in kidney epithelial cells. Our studies reveal that, in normal kidney cells, polycystin-1 forms a complex with the adherens junction protein E-cadherin and its associated catenins, suggesting a role in cell adhesion or polarity. In primary cells from ADPKD patients, the polycystin-1/polycystin-2/E-cadherin/beta-catenin complex was disrupted and both polycystin-1 and E-cadherin were depleted from the plasma membrane as a result of the increased phosphorylation of polycystin-1. The loss of E-cadherin was compensated by the transcriptional upregulation of the normally mesenchymal N-cadherin. Increased cell surface N-cadherin in the disease cells in turn stabilized the continued plasma membrane localization of beta-catenin in the absence of E-cadherin. The results suggest that enhanced phosphorylation of polycystin-1 in ADPKD cells precipitates changes in its localization and its ability to form protein complexes that are critical for the stabilization of adherens junctions and the maintenance of a fully differentiated polarized renal epithelium.

Adherens Junctions↗

PubMed2004

Development of multiorgan pathology in the wpk rat model of polycystic kidney disease.

Rodent models of polycystic kidney disease (PKD) have provided valuable insight into the cellular changes associated with cystogenesis in humans. The present study characterizes the morphology of renal and extrarenal pathology of autosomal recessive PKD induced by the wpk gene in Wistar rats. In wpk(-/-) rats, proximal tubule and collecting duct cysts develop in utero and eventually consume the kidney. Increased apoptosis, mitosis, and extracellular tenascin deposition parallel cyst development. Extrarenal pathology occurs in the immune system (thymic and splenic hypoplasia) and central nervous system (CNS; hypoplasia to agenesis of the corpus callosum with severe hydrocephalus). Severity of hydrocephalus varied inversely with size of the corpus callosum. In wpk(-/-) rats, the corpus callosum exhibits relatively few axons that cross the midline. This CNS pathology is similar to that described in three human renal cystic syndromes: orofaciodigital, genitopatellar, and cerebrorenal-digital syndromes. Collecting duct and ventricular ependymal cilia appear morphologically normal. To determine if rodent background strain and the presence of modifier genes affect severity of the disease, we crossed the Wistar-wpk rat with Brown Norway (BN) and Long Evan (LE) rats and found the degree of renal and cerebral pathology was diminished as evidenced by lower kidney weight as a percent of body weight and serum urea nitrogen concentration in cystic rats on LE or BN strains as well as less prominent cranial enlargement. Crosses with BN rats allowed us to localize the wpk gene on chromosome 5 very close to the D5Rat73 marker. The wpk gene lies within a chromosomal region known to harbor a PKD modifier locus. In summary, the types of renal and cerebral pathology seen in the Wistar wpk rat are a unique combination seen only in this rodent model.

Agenesis of Corpus Callosum↗

PubMed2004

PKHD1 mutations in autosomal recessive polycystic kidney disease (ARPKD).

Autosomal recessive polycystic kidney disease (ARPKD) is an important cause of childhood renal- and liver-related morbidity and mortality. The clinical spectrum is widely variable. About 30 to 50% of affected individuals die in the neonatal period, while others survive into adulthood. ARPKD is caused by mutations in the PKHD1 (polycystic kidney and hepatic disease 1) gene on chromosome 6p12, which is among the largest human genes, with a minimum of 86 exons assembled into a variety of alternatively spliced transcripts. The longest continuous open reading frame is predicted to yield a 4,074-aa (447-kDa) multidomain integral membrane protein (fibrocystin/polyductin) of unknown function. This update compiles all known PKHD1 mutations and polymorphisms/sequence variants. Mutations were found to be scattered throughout the gene without evidence of clustering at specific sites. Most PKHD1 mutations are unique to single families ("private mutations") hampering genotype-phenotype correlations. Correlations have been drawn for the type of mutation rather than for the site of individual mutations. All patients carrying two truncating mutations displayed a severe phenotype with perinatal or neonatal demise, while patients surviving the neonatal period bear at least one missense mutation. However, some missense changes are obviously as devastating as truncating mutations. The present article intends 1) to provide an overview of PKHD1 mutations and polymorphisms/sequence variants identified so far, 2) to discuss potential genotype-phenotype correlations, and 3) to review them in the context of their clinical implications. A constantly updated list of mutations is available online (www.humgen.rwth-aachen.de) and investigators are invited to submit their novel data to this PKHD1 mutation database.

DNA Mutational Analysis↗

PubMed2004

Molecular genetics of autosomal recessive polycystic kidney disease.

Autosomal recessive polycystic kidney disease (ARPKD) is a severe form of inherited childhood nephropathy ( approximately 1:20,000 live births) characterized by fusiform dilatation of collecting ducts and congenital hepatic fibrosis. Up to 30% die as neonates due to respiratory insufficiency and the majority of surviving infants develop hypertension. Progression to end stage renal disease occurs in 20-45% of cases within 15 years but a proportion maintain renal function into adulthood where complications of liver disease predominate. The ARPKD disease gene, PKHD1, has recently been identified through analysis of an orthologous animal model, the PCK rat. PKHD1 is a large gene ( approximately 470 kb) with 67 exons from which multiple transcripts may be generated by alternative splicing. It is highly expressed in kidney, with lower levels in liver and pancreas. The ARPKD protein, fibrocystin (4074 aa and 447 kDa), is predicted to be an integral membrane, receptor-like protein containing multiple copies of an Ig-like domain (TIG). Fibrocystin is localized to the branching ureteric bud, collecting and biliary ducts, consistent with the disease phenotype, and often absent from ARPKD tissue. In common with other PKD-related proteins, fibrocystin is localized to the primary cilia of renal epithelial cells, reinforcing the link between ciliary dysfunction and cyst development. Screens of PKHD1 have revealed 119 different mutations of various types spread throughout the gene. Several ancestral changes have been described, some localized to specific geographic populations. The majority of patients are compound heterozygotes and preliminary genotype/phenotype studies associate two truncating mutations with severe disease. The complexities of PKHD1, marked allelic heterogeneity and high level of missense changes complicate gene-based diagnostics.

Animals↗

PubMed2004

Biliary dysgenesis in the PCK rat, an orthologous model of autosomal recessive polycystic kidney disease.

Hepatic polycystic disease occurs alone or in combination with polycystic kidney disease (PKD). In autosomal recessive PKD (ARPKD), liver lesions are the major cause of morbidity and mortality in older patients. ARPKD is caused by a mutation to PKHD1 and the PCK rat is an orthologous model of disease. Recently, we showed that fibrocystin, Pkhd1 protein, is localized to primary cilia in rat cholangiocytes and that disruption of its ciliary expression results in biliary cystogenesis. This study describes biliary phenotype in the PCK rat using micro-computed tomography scanning and three-dimensional reconstruction, and light, scanning, and transmission microscopy. Our results show that the biliary tree undergoes extensive remodeling resulting in bile duct dilatation, focal budding, and formation of cysts that are initially connected to bile ducts, but throughout time separate from them. Progressive liver enlargement results from massive cyst formation while liver parenchymal volume remains unchanged. Cilia in cystic cells are abnormal consistent with the notion that the primary defect in ARPKD resulting in cystogenesis may be linked to ciliary dysfunction. Our results suggest that the PCK rat is a useful model for studies of biliary cystogenesis and treatment options of inherited cystic liver disease.

Animals↗

PubMed2004

Follow-up of intracranial aneurysms in autosomal-dominant polycystic kidney disease.

BACKGROUND: Approximately 8% of autosomal-dominant polycystic kidney disease (ADPKD) patients have intracranial aneurysms. The risk of growth and rupture of those discovered by presymptomatic screening is key to the feasibility and success of a screening program. This study was initiated to ascertain this risk. METHODS: ADPKD patients were offered screening with magnetic resonance (MR) imaging that included three-dimensional time-of-flight MR angiographic and three-dimensional phase-contrast sequences. Patients with aneurysms were recommended periodic surveillance, initially at 6 months and yearly, and less frequently after demonstration of their stability. RESULTS: Twenty-two saccular and one fusiform aneurysms were detected at the initial screening in 21 patients from 19 families (seven men and 14 women, 47.9 +/- 10.6 years old). All the saccular aneurysms were small (mean diameter 3.5 mm, range 2.0 to 6.5 mm) and the majority (77%) in the anterior circulation. Two patients died from unrelated causes without further follow-up. One patient was lost to follow-up. A new 2 mm middle cerebral artery aneurysm developed in one patient. One aneurysm increased from a size of 4 mm to 5 mm after a follow-up of 105 months. No aneurysmal development or growth occurred in the remaining 16 patients. No aneurysmal rupture occurred during a mean imaging follow-up of 81 months and a mean clinical follow-up of 92 months. During the period of the study, two additional ADPKD patients, with three intracranial aneurysms detected elsewhere by presymptomatic MR angiographic screening, were referred for surgical treatment. The larger size of these aneurysms (10, 8, and 8 mm) probably reflects referral bias. CONCLUSION: Most intracranial aneurysms detected by presymptomatic screening in ADPKD patients are small and in the anterior circulation. The follow-up results do not suggest an increased risk for growth and rupture, compared to those of intracranial aneurysms in the general population. These data do not support widespread screening for intracranial aneurysms in the ADPKD population.

Adult↗

PubMed2004

Epidermal growth factor receptor tyrosine kinase inhibition is not protective in PCK rats.

BACKGROUND: Advances in the understanding of cystogenesis, identification of the PKHD1 gene and availability of a rat model (the PCK rat) caused by a Pkhd1 mutation facilitate testing of therapies for autosomal-recessive polycystic kidney disease (ARPKD). Considerable support exists for the importance of the epidermal growth factor (EGF)/transforming growth factor-alpha (TGF-alpha)/EGF receptor (EGFR) axis and of the adenylyl cyclase-adenosine 3',5'-cyclic monophosphate (cAMP) pathway in the pathogenesis of cyst formation and progressive enlargement. METHODS: To determine whether EGFR tyrosine kinase inhibition is protective in the PCK rat, male and female animals were treated with EKI-785 or EKB-569 or with vehicle alone between 3 and 10 weeks of age. Biochemical and histomorphometric analysis, immunohistochemistry, immunoblotting, enzyme immunoassay, and quantitative reverse transcription-polymerase chain reaction (RT-PCR) were used to ascertain the effects of treatment. RESULTS: Contrary to other murine models of ARPKD, overexpression and apical mislocalization of EGFR were not detected in the PCK rats. Consistent with these expression results, EKI-785 or EKB-569 administration had no effect or worsened PKD, and had no effect on the development of fibrocystic liver disease. Increased renal cAMP and vasopressin V2 receptor expression were observed in the EKI-785-treated animals. CONCLUSION: EGFR tyrosine kinase inhibition did not protect PCK rats from the development of PKD. This may be due to effects on collecting duct cAMP that counteract possible beneficial effects on the extracellular-regulated protein kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway, particularly in the absence of EGFR overexpression or mislocalization. The relevance of these observations to the treatment of human cystic kidney diseases deserves further study.

Aminoquinolines↗

PubMed2003

Inhibition of renal cystic disease development and progression by a vasopressin V2 receptor antagonist.

The polycystic kidney diseases (PKDs) are a group of genetic disorders causing significant renal failure and death in children and adults. There are no effective treatments. Two childhood forms, autosomal recessive PKD (ARPKD) and nephronophthisis (NPH), are characterized by collecting-duct cysts. We used animal models orthologous to the human disorders to test whether a vasopressin V2 receptor (VPV2R) antagonist, OPC31260, would be effective against early or established disease. Adenosine-3',5'-cyclic monophosphate (cAMP) has a major role in cystogenesis, and the VPV2R is the major cAMP agonist in the collecting duct. OPC31260 administration lowered renal cAMP, inhibited disease development and either halted progression or caused regression of established disease. These results indicate that OPC31260 may be an effective treatment for these disorders and that clinical trials should be considered.

Animals↗

PubMed2003

Cellular and subcellular localization of the ARPKD protein; fibrocystin is expressed on primary cilia.

Autosomal recessive polycystic kidney disease (ARPKD) is an infantile form of PKD characterized by fusiform dilation of collecting ducts and congenital hepatic fibrosis. The ARPKD gene, PKHD1, is large (approximately 470 kb; 67 exons) with a 12222 bp longest open reading frame, although multiple different splice forms may be generated. The predicted full-length ARPKD protein, fibrocystin, is membrane bound with 4074 amino acids (447 kDa molecular weight). To characterize the pattern of fibrocystin expression we have generated four monoclonal antibodies (mAb) to the cytoplasmic tail of the protein. Western analysis of human kidney membrane protein showed an identical pattern with each mAb; a strongly expressing large product (>450 kDa), consistent with the predicted protein size, and a weaker approximately 220 kDa band. The same large product was detected in rat and mouse kidney with lower level expression in liver. To further show that these mAbs recognize fibrocystin, tissue from ARPKD patients was analyzed and no fibrocystin products were detected. Immunohistochemical analysis of the developing kidney showed expression in the branching ureteric bud and collecting ducts, expression that persisted into adulthood. Biliary duct staining was found in the liver, plus staining in the pancreas and developing testis. Immunofluorescence analysis of MDCK cells showed a major site of expression in the primary cilia. Recent studies have associated the disease protein in various human and animal forms of PKD with cilia. The localization of fibrocystin to cilia further strengthens that correlation and indicates that the primary defect in ARPKD may be linked to ciliary dysfunction.

Animals↗

PubMed2003

Pkd2 haploinsufficiency alters intracellular calcium regulation in vascular smooth muscle cells.

Autosomal-dominant polycystic kidney disease is a multiorgan disease and its vascular manifestations are common and life-threatening. Despite this, little is known about their pathogenesis. Somatic mutations to the normal PKD allele in cystic epithelia and cyst development associated with the unstable Pkd2(WS25) allele suggest a two-hit model of cystogenesis. However, it is unclear if this model can account for the cardiovascular pathology or if haploinsufficiency alone is disease-associated. In the present study, we found a decreased polycystin-2 (PC2, protein encoded by Pkd2 gene) expression in Pkd2( +/-) vessels, roughly half the wild-type level, and an enhanced level of intracranial vascular abnormalities in Pkd2 (+/-) mice when induced to develop hypertension. Consistent with these observations, freshly dissociated Pkd2 (+/-) vascular smooth muscle cells have significantly altered intracellular Ca(2+) homeostasis. The resting [Ca(2+)](i) is 17.1% lower in Pkd2 (+/-) compared with wild-type cells (P=0.0003) and the total sarcoplasmic reticulum Ca(2+) store (emptied by caffeine plus thapsigargin) is decreased (P<0.0001). The store operated Ca(2+) (SOC) channel activity is also decreased in Pkd2 (+/-) cells (P=0.008). These results indicate that inactivation of just one Pkd2 allele is sufficient to significantly alter intracellular Ca(2+) homeostasis, and that PC2 is necessary to maintain normal SOC activity and the SR Ca(2+) store in VSMCs. Based on these findings, and the fact that [Ca(2+)](i) signaling is essential to the regulation of contraction, production and secretion of extracellular matrix, cellular proliferation and apoptosis, we propose that the abnormal intracellular Ca(2+) regulation associated with Pkd2 haploinsufficiency is directly related to the vascular phenotype.

Alleles↗

PubMed2003

Association of mutation position in polycystic kidney disease 1 (PKD1) gene and development of a vascular phenotype.

BACKGROUND: Patients with autosomal dominant polycystic kidney disease (ADPKD) are at risk of developing intracranial aneurysms, and subarachnoid haemorrhage is a major cause of death and disability. Familial clustering of intracranial aneurysms suggests that genetic factors are important in the aetiology. We tested whether the germline mutation predisposes to this vascular phenotype. METHODS: DNA samples from patients with ADPKD and vascular complications were screened for mutations throughout the PKD1 and PKD2 genes. Comparisons were made between the PKD1 and PKD2 populations and with a control PKD1 cohort (without the vascular phenotype). FINDINGS: Mutations were characterised in 58 ADPKD families with vascular complications; 51 were PKD1 (88%) and seven PKD2 (12%). The median position of the PKD1 mutation was significantly further 59 in the vascular population than in the 87 control pedigrees (aminoacid position 2163 vs 2773, p=0.0034). Subsets of the vascular population with aneurysmal rupture, early rupture, or families with more than one vascular case had median mutation locations further 59 (aminoacid position 1811, p=0.0018; 1671, p=0.0052; and 1587, p=0.0003). INTERPRETATION: Patients with PKD2, as well as those with PKD1, are at risk of intracranial aneurysm. The position of the mutation in PKD1 is predictive for development of intracranial aneurysms (59 mutations are more commonly associated with vascular disease) and is therefore of prognostic importance. Since the PKD1 phenotype is associated with mutation position, the disease is not simply due to loss of all disease allele products.

Adult↗

PubMed2003

PKHDL1, a homolog of the autosomal recessive polycystic kidney disease gene, encodes a receptor with inducible T lymphocyte expression.

Autosomal-recessive polycystic kidney disease (ARPKD) is caused by mutation to a large gene, PKHD1, encoding a putative receptor protein, fibrocystin. We have identified, through analysis of human genomic sequence, a PKHD1 homolog, PKHDL1, in chromosome region 8q23. The PKHDL1 transcript of 13081 bp was amplified as 16 fragments and sequenced; the sequence of the murine ortholog, Pkhdl1 (chromosome region 15B3) was also determined. PKHDL1 contains 78 exons, covers a genomic region of approximately 168 kb and encodes a large protein, fibrocystin-L. Screening PKHDL1 in ARPKD patients with no PKHD1 mutations revealed several sequence variants but no clear mutations, making it unlikely that it is ARPKD-associated. Human fibrocystin-L is predicted to be a large receptor protein (4243 aa; 466 kDa) with a signal peptide, single transmembrane domain and short cytoplasmic tail. Fibrocystin-L is homologous to fibrocystin throughout most of the extracellular region with overall identity of 25.0% and similarity of 41.5%. Fibrocystin-L has extracellular domains similar to fibrocystin with 14 copies of the TIG domain and two regions of significant homology to the protein TMEM2. Genomic sequence analysis identified no other full-length fibrocystin homologs in humans, mice or other sequenced organisms. The Fugu fish has a fibrocystin-L ortholog but no fibrocystin, suggesting that the newly identified protein may be the ancestral form. PKHDL1 and Pkhdl1 are widely expressed at a low level in most tissues but only detected in blood-derived cell-lines. Low level expression was detected in many primary immune cell subtypes but up-regulated specifically in T lymphocytes, following activation signals, suggesting a role in cellular immunity.

Amino Acid Sequence↗

PubMed2003

Defects in cholangiocyte fibrocystin expression and ciliary structure in the PCK rat.

BACKGROUND & AIMS: Recent studies have showed that proteins associated with polycystic kidney disease (PKD) are expressed in cilia, linking this organelle and cyst formation in the kidney, but involvement of cilia in PKD-related biliary cystogenesis has not been shown. We investigated: (1) the expression of fibrocystin (a product of PKHD1, the autosomal-recessive PKD [ARPKD] gene) in cholangiocyte cilia; (2) biliary cyst formation in an orthologous rat model, PCK; and (3) the effect of Pkhd1 mutation on ciliary structure. METHODS: Biliary cystogenesis was assessed by microcomputed tomography. Fibrocystin expression in cholangiocytes of isolated intrahepatic bile ducts (IBDUs) and liver cysts was analyzed by confocal and immunoelectron microscopy, and ciliary structure and length by scanning and transmission electron microscopy. Small interfering RNAs (siRNA) were used to examine the effect of fibrocystin loss on ciliary structure. RESULTS: The biliary tree in the PCK rat was distorted markedly, showing multiple bile duct dilatation and focal budding. In normal IBDUs, each cholangiocyte had a single cilium that expressed fibrocystin. In contrast, cilia in the PCK rat were abnormal with bulbous extensions and diminished length, and were devoid of fibrocystin. In cholangiocytes of normal IBDUs, specific siRNA reduced Pkhd1 messenger RNA by 80%, the length of cilia by 41%, and fibrocystin ciliary expression to an undetectable level. CONCLUSIONS: Our results indicate that fibrocystin is expressed in cholangiocyte cilia and that disruption of Pkhd1 by a germ line mutation in the PCK rat or by siRNA in IBDUs results in abnormalities in ciliary morphology and possibly biliary cystogenesis.

Animals↗

PubMed2003

A complete mutation screen of PKHD1 in autosomal-recessive polycystic kidney disease (ARPKD) pedigrees.

BACKGROUND: Autosomal-recessive polycystic kidney disease (ARPKD) is an important neonatal nephropathy characterized by fusiform dilation of collecting ducts, congenital hepatic fibrosis, and in some cases Caroli's disease. The ARPKD gene, PKHD1, has recently been identified. Herein we describe an effective method for PKHD1 mutation screening and the results from analysis of a novel ARPKD cohort. METHODS: The coding region of PKHD1 was amplified as 79 fragments and analyzed for base pair changes by denaturing high-performance liquid chromatography (DHPLC). Forty-seven ARPKD and 14 pedigrees with congenital hepatic fibrosis and/or Caroli's disease, were screened for PKHD1 mutations. RESULTS: Thirty-three different mutations were detected on 57 alleles (51.1% ARPKD, 32.1% congenital hepatic fibrosis/Caroli's disease). In the 22 pedigrees where both mutations were identified, two were homozygous for 9689delA and the remainder were compound heterozygotes; a combination of truncating, missense and splicing changes. Patients with two truncating mutations all died in the perinatal period. Two frequent truncating mutations were identified: 9689delA (9 alleles) and 5896insA (8 alleles) plus some more common missense changes; haplotype analysis indicated most were ancestral mutations. CONCLUSION: DHPLC has been established as a rapid mutation screening method for ARPKD. The mutation detection rate was high in severely affected patients (85%), lower in those with moderate ARPKD (41.9%), and low, but significant, in adults with congenital hepatic fibrosis/Caroli's disease (32.1%). The prospects for gene-based diagnostics are complicated by the large gene size, marked allelic heterogeneity, and clinical diversity of the ARPKD phenotype. Identification of some common mutations, especially in specific populations, will aid mutation screening.

Adolescent↗

PubMed2003

EGF receptor tyrosine kinase inhibition attenuates the development of PKD in Han:SPRD rats.

BACKGROUND: Increasing evidence supports an important role for the epidermal growth factor (EGF)/transforming growth factor-alpha (TGF-alpha)/EGF receptor (EGFR) axis in promoting tubular epithelial cell proliferation and cyst formation in polycystic kidney disease (PKD). METHODS: To determine whether the inhibition of EGFR tyrosine kinase activity can attenuate the development of PKD in the Han:SPRD rat, a frequently used animal model of autosomal-dominant slowly progressive PKD (ADPKD), wild-type and cy/+ rats were treated with EKI-785 or EKB-569 or with vehicle alone. Western analysis, immunoprecipitation, and immunohistochemistry were used to ascertain the expression, activation, and localization of EGFR. RESULTS: Overexpression, activation and apical mislocalization of EGFR were observed in the cy/+ rats. The intraperitoneal administration of EKI-785 reversed the activation of the EGFR to the level observed in wild-type animals. The intraperitoneal administration of EKI-785 (90 mg/kg body weight every third day) or of EKB-569 (20 mg/kg body weight every third day) to cy/+ rats resulted in lower kidney weights, serum concentrations of blood urea nitrogen (BUN), cyst volumes, and fibrosis scores. The administration of EKB-569 by gavage was less effective probably because of lower bioavailability. CONCLUSION: These results support a significant role for the EGF/TGF-alpha/EGFR axis in the development of PKD in the Han:SPRD rat and the therapeutic potential of EGFR tyrosine kinase inhibition in ADPKD.

Animals↗

PubMed2003

The ENOS polymorphism is not associated with severity of renal disease in polycystic kidney disease 1.

BACKGROUND: The renal phenotype in autosomal dominant polycystic kidney disease (ADPKD) is highly variable. Although genetic and allelic heterogeneity explain part of this variability, significant intrafamilial differences in time to end-stage renal disease (ESRD) indicate that genetic modifiers and the environment influence renal phenotype. Previously, a glutamic acid to aspartic acid polymorphism at residue 298 (E/D298) of the endothelial nitric oxide synthase (eNOS) gene ENOS was associated with disease severity in males with ADPKD. METHODS: We typed the E/D298 polymorphism in 215 mutation-defined polycystic kidney disease 1 (PKD1) patients from 80 families. In this population, 96 patients had ESRD, with a median time to renal failure of 53 years. RESULTS: Distribution of ENOS genotypes was 86 (40%), 106 (49.3%), and 23 (10.7%) for EE, ED, and DD, respectively. The occurrence of hypertension was not significantly different between genotypes. Kaplan-Meier renal survival analysis showed no significant difference between genotypes, with a median age to ESRD of 53 years for all genotypes in the total population and 52, 52, and 51 years (men) and 57, 53, and 55 years (women) for DD, DE, and EE, respectively. CONCLUSION: Although the D298 ENOS allele may be associated with lower vascular activity of eNOS, this did not correlate with severity of renal disease in this PKD1 population. An important difference between this study and one finding a modifying role for ENOS was the rigor in defining the PKD1 population. This study shows the importance of using mutation-characterized populations for association studies in ADPKD.

Age Factors↗

PubMed2003

Analysis of the polycystins in aortic vascular smooth muscle cells.

The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD.

Animals↗

PubMed2002

Pioglitazone improves the phenotype and molecular defects of a targeted Pkd1 mutant.

Mutations of either PKD1 or PKD2 are associated with autosomal dominant polycystic kidney disease (ADPKD). The molecular function of the gene product of PKD1, polycystin-1, in vitro has been elucidated recently, but the molecular pathological consequences of the loss of polycystin-1 in vivo have remained unclear. We have generated a mouse with a targeted deletion of exons 2-6 of Pkd1 to study the molecular defects in Pkd1 mutants. Homozygote embryos (Pkd1(-/-)) developed hydrops, cardiac conotruncal defects and renal cystogenesis. Total protein levels of beta-catenin in heart and kidney and c-MYC in heart were decreased in Pkd1(-/-) embryos. In the kidneys of Pkd1(-/-), the expression of E-cadherin and PECAM in basolateral membranes of renal tubules was attenuated, and tyrosine phosphorylation of epidermal growth factor receptor and Gab1 were constitutively enhanced when cystogenesis started on embryonic day (E) 15.5-16.5. Maternally administered pioglitazone, a thiazolidinedione compound, resolved these molecular defects of Pkd1(-/-). Treatment with pioglitazone improved survival of Pkd1(-/-) embryos and ameliorated the cardiac defects and the degree of renal cystogenesis. Long-term treatment with pioglitazone improved the endothelial function of adult Pkd1(+/-). These data indicated that molecular defects observed in Pkd1(-/-) embryos contributed to the pathogenesis of ADPKD and that thiazolidinediones had a compensatory effect on the pathway affected by the loss of polycystin-1. Pathways activated by thiazolidinediones may provide new therapeutic targets in ADPKD.

Animals↗