Renal ischemia alters the transcriptomic and epigenetic profile of inflammatory genes in swine scattered tubular-like cells.

BACKGROUND: Scattered tubular-like cells (STCs) are differentiated renal tubular cells that during recovery from ischemic injury dedifferentiate to repair other injured renal cells. Renal artery stenosis (RAS), often associated with chronic inflammatory injury, compromises the integrity and function of STCs, but the underlying mechanisms remain unknown. We hypothesized that RAS alters the transcriptomic and epigenetic profile of inflammatory genes in swine STCs. METHODS: STCs were harvested from pig kidneys after 10 weeks of RAS or sham (n=6 each). STC mRNA profiles of inflammatory genes were analyzed using high-throughput mRNA-sequencing (seq) and their DNA methylation (5mC) and hydroxymethylation (5hmC) profiles by DNA immunoprecipitation and next-generation sequencing (MeDIP-seq) (n=3 each), followed by an integrated (mRNA-seq/MeDIP-seq) analysis. STC protein expression of candidate differentially expressed (DE) genes and common proinflammatory proteins were subsequently assessed in vitro before and after epigenetic (Bobcat339) modulation. RESULTS: mRNA-seq identified 57 inflammatory genes up-regulated in RAS-STCs versus Normal-STCs (>1.4 or <0.7-fold, P<0.05), of which 14% exhibited lower 5mC and 5% higher 5hmC levels in RAS-STCs versus Normal-STCs, respectively. Inflammatory gene and protein expression was higher in RAS-STCs compared with Normal-STCs but normalized after epigenetic modulation. CONCLUSIONS: These observations highlight a novel modulatory mechanism of this renal endogenous repair system and support development of epigenetic or anti-inflammatory therapies to preserve the reparative capacity of STCs in individuals with RAS.

Congenital Heart Disease in Adults with Autosomal Dominant Polycystic Kidney Disease.

INTRODUCTION: Autosomal dominant polycystic kidney disease (ADPKD) is caused mainly by pathogenic variants in PKD1 or PKD2 encoding the polycystin-1 and -2 proteins. Polycystins have shown to have an essential role in cardiac development and function in animal models. In the current study, we describe the clinical association between ADPKD and congenital heart disease (CHD). METHODS: Medical records from Mayo Clinic were queried for all patients with confirmed ADPKD and CHD between 1993 and 2020. CHD was categorized into left-to-right shunt, obstructive, and complex lesions. Patent foramen ovale, mitral valve prolapse, and bicuspid aortic valve anomalies were excluded. RESULTS: Twenty-five out of 1,359 (1.84%) ADPKD patients were identified to have CHD. Of these, 84% were Caucasians and 44% were males. The median (Q1-Q3) age (years) at CHD diagnosis was 12.0 (2.0-43.5). Fourteen patients (56%) had left-to-right shunt lesions, 6 (24%) had obstructive lesions and 5 (20%) complex lesions. Seventeen patients (68%) had their defects surgically corrected at a median age (Q1-Q3) of 5.5 (2.0-24.7). Among 13 patients with available genetic testing, 12 (92.3%) had PKD1 pathogenic variants, and none had PKD2. The median (Q1-Q3) age at last follow-up visit was 47.0 (32.0-62.0) and median (Q1-Q3) eGFR was 35.8 (11.4-79.0) mL/min/1.73 m2. Three patients (12%) died; all of them had left-to-right shunt lesions. DISCUSSION/CONCLUSION: We observed a higher CHD frequency in ADPKD than the general population (1.84 vs. 0.4%). While only PKD1 pathogenic variants were identified in this cohort, further studies are needed to confirm this novel finding and understand the role of polycystins in the development of the heart and vessels.

Renal mitochondrial injury in the pathogenesis of CKD: mtDNA and mitomiRs.

Chronic kidney disease (CKD) is a public health concern that affects over 200 million people worldwide and is associated with a tremendous economic burden. Therefore, deciphering the mechanisms underpinning CKD is crucial to decelerate its progression towards end-stage renal disease (ESRD). Renal tubular cells are populated with a high number of mitochondria, which produce cellular energy and modulate several important cellular processes, including generation of reactive oxygen species (ROS), calcium homeostasis, proliferation, and apoptosis. Over the past few years, increasing evidence has implicated renal mitochondrial damage in the pathogenesis of common etiologies of CKD, such as diabetes, hypertension, metabolic syndrome (MetS), chronic renal ischemia, and polycystic kidney disease (PKD). However, most compelling evidence is based on preclinical studies because renal biopsies are not routinely performed in many patients with CKD. Previous studies have shown that urinary mitochondrial DNA (mtDNA) copy numbers may serve as non-invasive biomarkers of renal mitochondrial dysfunction. Emerging data also suggest that CKD is associated with altered expression of mitochondria-related microRNAs (mitomiRs), which localize in mitochondria and regulate the expression of mtDNA and nucleus-encoded mitochondrial genes. This review summarizes relevant evidence regarding the involvement of renal mitochondrial injury and dysfunction in frequent forms of CKD. We further provide an overview of non-invasive biomarkers and potential mechanisms of renal mitochondrial damage, especially focusing on mtDNA and mitomiRs.

The genetic background significantly impacts the severity of kidney cystic disease in the Pkd1RC/RC mouse model of autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD), primarily due to PKD1 or PKD2 mutations, causes progressive kidney cyst development and kidney failure. There is significant intrafamilial variability likely due to the genetic background and environmental/lifestyle factors; variability that can be modeled in PKD mice. Here, we characterized mice homozygous for the PKD1 hypomorphic allele, p.Arg3277Cys (Pkd1RC/RC), inbred into the BALB/cJ (BC) or the 129S6/SvEvTac (129) strains, plus F1 progeny bred with the previously characterized C57BL/6J (B6) model; F1(BC/B6) or F1(129/B6). By one-month cystic disease in both the BC and 129 Pkd1RC/RC mice was more severe than in B6 and continued with more rapid progression to six to nine months. Thereafter, the expansive disease stage plateaued/declined, coinciding with increased fibrosis and a clear decline in kidney function. Greater severity correlated with more inter-animal and inter-kidney disease variability, especially in the 129-line. Both F1 combinations had intermediate disease severity, more similar to B6 but progressive from one-month of age. Mild biliary dysgenesis, and an early switch from proximal tubule to collecting duct cysts, was seen in all backgrounds. Preclinical testing with a positive control, tolvaptan, employed the F1(129/B6)-Pkd1RC/RC line, which has moderately progressive disease and limited isogenic variability. Magnetic resonance imaging was utilized to randomize animals and provide total kidney volume endpoints; complementing more traditional data. Thus, we show how genetic background can tailor the Pkd1RC/RC model to address different aspects of pathogenesis and disease modification, and describe a possible standardized protocol for preclinical testing.

Efficacy of Rituximab and Plasma Exchange in Antineutrophil Cytoplasmic Antibody-Associated Vasculitis with Severe Kidney Disease.

BACKGROUND: Treatment of patients with ANCA-associated vasculitis (AAV) and severe renal involvement is not established. We describe outcomes in response to rituximab (RTX) versus cyclophosphamide (CYC) and plasma exchange (PLEX). METHODS: A retrospective cohort study of MPO- or PR3-ANCA-positive patients with AAV (MPA and GPA) and severe kidney disease (eGFR <30 ml/min per 1.73 m2). Remission, relapse, ESKD and death after remission-induction with CYC or RTX, with or without the use of PLEX, were compared. RESULTS: Of 467 patients with active renal involvement, 251 had severe kidney disease. Patients received CYC (n=161) or RTX (n=64) for remission-induction, and 51 were also treated with PLEX. Predictors for ESKD and/or death at 18 months were eGFR <15 ml/min per 1.73 m2 at diagnosis (IRR 3.09 [95% CI 1.49 to 6.40], P=0.002), renal recovery (IRR 0.27 [95% CI 0.12 to 0.64], P=0.003) and renal remission at 6 months (IRR 0.40 [95% CI 0.18 to 0.90], P=0.027). RTX was comparable to CYC in remission-induction (BVAS/WG=0) at 6 months (IRR 1.37 [95% CI 0.91 to 2.08], P=0.132). Addition of PLEX showed no benefit on remission-induction at 6 months (IRR 0.73 [95% CI 0.44 to 1.22], P=0.230), the rate of ESKD and/or death at 18 months (IRR 1.05 [95% CI 0.51 to 2.18], P=0.891), progression to ESKD (IRR 1.06 [95% CI 0.50 to 2.25], P=0.887), and survival at 24 months (IRR 0.54 [95% CI 0.16 to 1.85], P=0.330). CONCLUSIONS: The apparent benefits and risks of using CYC or RTX for the treatment of patients with AAV and severe kidney disease are balanced. The addition of PLEX to standard remission-induction therapy showed no benefit in our cohort. A randomized controlled trial is the only satisfactory means to evaluate efficacy of remission-induction treatments in AAV with severe renal involvement.

Expanded Imaging Classification of Autosomal Dominant Polycystic Kidney Disease.

BACKGROUND: The Mayo Clinic imaging classification of autosomal dominant polycystic kidney disease (ADPKD) uses height-adjusted total kidney volume (htTKV) and age to identify patients at highest risk for disease progression. However, this classification applies only to patients with typical diffuse cystic disease (class 1). Because htTKV poorly predicts eGFR decline for the 5%-10% of patients with atypical morphology (class 2), imaging-based risk modeling remains unresolved. METHODS: Of 558 adults with ADPKD in the HALT-A study, we identified 25 patients of class 2A with prominent exophytic cysts (class 2Ae) and 43 patients of class 1 with prominent exophytic cysts; we recalculated their htTKVs to exclude exophytic cysts. Using original and recalculated htTKVs in association with imaging classification in logistic and mixed linear models, we compared predictions for developing CKD stage 3 and for eGFR trajectory. RESULTS: Using recalculated htTKVs increased specificity for developing CKD stage 3 in all participants from 82.6% to 84.2% after adjustment for baseline age, eGFR, BMI, sex, and race. The predicted proportion of class 2Ae patients developing CKD stage 3 using a cutoff of 0.5 for predicting case status was better calibrated to the observed value of 13.0% with recalculated htTKVs (45.5%) versus original htTKVs (63.6%). Using recalculated htTKVs reduced the mean paired difference between predicted and observed eGFR from 17.6 (using original htTKVs) to 4.0 ml/min per 1.73 m2 for class 2Ae, and from -1.7 (using original htTKVs) to 0.1 ml/min per 1.73 m2 for class 1. CONCLUSIONS: Use of a recalculated htTKV measure that excludes prominent exophytic cysts facilitates inclusion of class 2 patients and reclassification of class 1 patients in the Mayo classification model.

Oxidative Stress and Mitochondrial Abnormalities Contribute to Decreased Endothelial Nitric Oxide Synthase Expression and Renal Disease Progression in Early Experimental Polycystic Kidney Disease.

Vascular abnormalities are the most important non-cystic complications in Polycystic Kidney Disease (PKD) and contribute to renal disease progression. Endothelial dysfunction and oxidative stress are evident in patients with ADPKD, preserved renal function, and controlled hypertension. The underlying biological mechanisms remain unknown. We hypothesized that in early ADPKD, the reactive oxygen species (ROS)-producing nicotinamide adenine dinucleotide phosphate hydrogen (NAD(P)H)-oxidase complex-4 (NOX4), a major source of ROS in renal tubular epithelial cells (TECs) and endothelial cells (ECs), induces EC mitochondrial abnormalities, contributing to endothelial dysfunction, vascular abnormalities, and renal disease progression. Renal oxidative stress, mitochondrial morphology (electron microscopy), and NOX4 expression were assessed in 4- and 12-week-old PCK and Sprague-Dawley (wild-type, WT) control rats (n = 8 males and 8 females each). Endothelial function was assessed by renal expression of endothelial nitric oxide synthase (eNOS). Peritubular capillaries were counted in hematoxylin-eosin (H&E)-stained slides and correlated with the cystic index. The enlarged cystic kidneys of PCK rats exhibited significant accumulation of 8-hydroxyguanosine (8-OHdG) as early as 4 weeks of age, which became more pronounced at 12 weeks. Mitochondria of TECs lining cysts and ECs exhibited loss of cristae but remained preserved in non-cystic TECs. Renal expression of NOX4 was upregulated in TECs and ECs of PCK rats at 4 weeks of age and further increased at 12 weeks. Contrarily, eNOS immunoreactivity was lower in PCK vs. WT rats at 4 weeks and further decreased at 12 weeks. The peritubular capillary index was lower in PCK vs. WT rats at 12 weeks and correlated inversely with the cystic index. Early PKD is associated with NOX4-induced oxidative stress and mitochondrial abnormalities predominantly in ECs and TECs lining cysts. Endothelial dysfunction precedes capillary loss, and the latter correlates with worsening of renal disease. These observations position NOX4 and EC mitochondria as potential therapeutic targets in PKD.

CKD Due to a Novel Mitochondrial DNA Mutation: A Case Report.

In human kidney disease, mitochondrial ultrastructural damage has long been recognized. Although the extent to which such mitochondrial changes contribute to human kidney disease is uncertain, experimental studies clearly demonstrate that mitochondrial damage can instigate pathogenetic processes that drive ongoing kidney disease. Clinical credence for this experimentally based hypothesis is provided by the development of kidney disease in patients with primary mitochondrial disorders. In this regard, substantial interest surrounds the occurrence of kidney disease in primary mitochondrial cytopathies, a heterogeneous group of conditions in which mutations in mitochondrial DNA (mtDNA) or nuclear DNA impair the functionality of components of the mitochondrial respiratory chain. We describe a novel mtDNA mutation in a patient who developed chronic kidney disease. The patient exhibited mitochondrial abnormalities in both muscle and kidney, chronic tubulointerstitial changes, and recurrent episodes of rhabdomyolysis. We outline mechanisms that may underlie the occurrence of chronic kidney disease in the setting of this novel mtDNA mutation. We also underscore the need to consider in relevant kidney diseases the presence of an underlying mitochondrial cytopathy because the latter more commonly exists than is generally recognized.

Three-dimensional NMR microscopy of zebrafish specimens.

While zebrafish embryos in the first five days after fertilization are clear and amenable to optical analysis, older juveniles and adults are not, due to pigmentation development and tissue growth. Thus other imaging methods are needed to image adult specimens. NMR is a versatile tool for studies of biological systems and has been successfully used for in vivo zebrafish microscopy. In this work we use NMR microscopy (MRM) for assessment of zebrafish specimens, which includes imaging of formalin fixed (FF), formalin fixed and paraffin embedded (FFPE), fresh (unfixed), and FF gadolinium doped specimens. To delineate the size and shape of various organs we concentrated on 3D MRM. We have shown that at 7 T a 3D NMR image can be obtained with isotropic resolution of 50 µm/pxl within 10 min and 25 µm/pxl within 4 h. Also, we have analyzed sources of contrast and have found that in FF specimens the best contrast is obtained by T1 weighting (3D FLASH, 3D FISP), whereas in FFPE specimens T2 weighting (3D RARE) is the best. We highlight an approach to perform segmentation of the organs in order to study morphological changes associated with mutations. The broader implication of this work is development of NMR methodology for high contrast and high resolution serial imaging and automated analysis of morphology of various zebrafish mutants.

Can we further enrich autosomal dominant polycystic kidney disease clinical trials for rapidly progressive patients? Application of the PROPKD score in the TEMPO trial.

Background: The PROPKD score has been proposed to stratify the risk of progression to end-stage renal disease in autosomal dominant polycystic kidney disease (ADPKD) subjects. We aimed to assess its prognostic value in a genotyped subgroup of subjects from the Tolvaptan Phase 3 Efficacy and Safety Study in Autosomal Dominant Polycystic Kidney Disease (TEMPO3/4) trial. Methods: In the post hoc analysis, PKD1 and PKD2 were screened in 770 subjects and the PROPKD score was calculated in mutation-positive subjects (male: 1 point; hypertension <35 years: 2 points; first urologic event <35 years: 2 points; nontruncating PKD1 mutation: 2 points; truncating PKD1 mutation: 4 points). Subjects were classified into low-risk (LR; 0-3 points), intermediate-risk (IR; 4-6 points) and high-risk (HR; 7-9 points) groups. Results: The PROPKD score was calculated in 749 subjects (LR = 132, IR = 344 and HR = 273); age was inversely related to risk (LR = 43.6 years, IR = 39.5 years, HR = 36.2 years; P < 0.001). Subjects from the HR group had significantly higher height-adjusted total kidney volume (TKV) and rates of TKV growth. While baseline renal function was similar across all risk groups, the rate of estimated glomerular filtration rate (eGFR) decline significantly increased from LR to HR in the placebo group. Tolvaptan treatment effectiveness to reduce TKV growth was similar in all three risk categories. While tolvaptan significantly slowed eGFR decline in the IR (tolvaptan = -2.34 versus placebo = -3.33 mL/min/1.73 m2/year; P = 0.008) and HR groups (tolvaptan = -2.74 versus placebo = -3.94 mL/min/1.73 m2/year; P = 0.002), there was no difference in the LR group (tolvaptan = -2.35 versus placebo = -2.50 mL/min/1.73 m2/year; P = 0.72). Excluding the LR subjects from the analysis improved the apparent treatment effect of tolvaptan on eGFR decline. Conclusion: This study confirms the prognostic value of the PROPKD score and suggests that it could reduce costs and enhance endpoint sensitivity by enriching future study populations for rapidly progressing ADPKD subjects.

The regulatory 1α subunit of protein kinase A modulates renal cystogenesis.

The failure of the polycystins (PCs) to function in primary cilia is thought to be responsible for autosomal dominant polycystic kidney disease (ADPKD). Primary cilia integrate multiple cellular signaling pathways, including calcium, cAMP, Wnt, and Hedgehog, which control cell proliferation and differentiation. It has been proposed that mutated PCs result in reduced intracellular calcium, which in turn upregulates cAMP, protein kinase A (PKA) signaling, and subsequently other proliferative signaling pathways. However, the role of PKA in ADPKD has not been directly ascertained in vivo, although the expression of the main regulatory subunit of PKA in cilia and other compartments (PKA-RIα, encoded by PRKAR1A) is increased in a mouse model orthologous to ADPKD. Therefore, we generated a kidney-specific knockout of Prkar1a to examine the consequences of constitutive upregulation of PKA on wild-type and Pkd1 hypomorphic (Pkd1RC) backgrounds. Kidney-specific loss of Prkar1a induced renal cystic disease and markedly aggravated cystogenesis in the Pkd1RC models. In both settings, it was accompanied by upregulation of Src, Ras, MAPK/ERK, mTOR, CREB, STAT3, Pax2 and Wnt signaling. On the other hand, Gli3 repressor activity was enhanced, possibly contributing to hydronephrosis and impaired glomerulogenesis in some animals. To assess the relevance of these observations in humans we looked for and found evidence for kidney and liver cystic phenotypes in the Carney complex, a tumoral syndrome caused by mutations in PRKAR1A These observations expand our understanding of the pathogenesis of ADPKD and demonstrate the importance of PRKAR1A highlighting PKA as a therapeutic target in ADPKD.

Prognostic enrichment design in clinical trials for autosomal dominant polycystic kidney disease: the HALT-PKD clinical trial.

BACKGROUND: Patients with mild autosomal dominant polycystic kidney disease (ADPKD) are less likely to be informative in randomized clinical trials (RCTs). We previously developed an imaging classification of ADPKD (typical diffuse cyst distribution Class 1A-E and atypical cyst distribution Class 2) for prognostic enrichment design in RCTs. We investigated whether using this classification would have increased the power to detect a beneficial treatment effect of rigorous blood pressure (BP) control on HALT-PKD participants with early disease (Study A). METHODS: Post hoc analysis of the early disease HALT-PKD study, an RCT that studied the effect of rigorous versus standard BP control on rates of total kidney volume (TKV) increase and estimated glomerular filtration rate (eGFR) decline in ADPKD patients with eGFR >60 mL/min/1.73 m2. RESULTS: Five hundred and fifty-one patients were classified by two observers (98.2% agreement) into Class 1A (6.2%), 1B (20.3%), 1C (34.1%), 1D (22.1%), 1E (11.8%) and 2 (5.4%). The TKV increase and eGFR decline became steeper from Class 1A through 1E. Rigorous BP control had been shown to be associated with slower TKV increase, without a significant overall effect on the rate of eGFR decline (faster in the first 4 months and marginally slower thereafter). Merging Classes 1A and 2 (lowest severity), 1B and 1C (intermediate severity) and 1D and 1E (highest severity) detected stronger beneficial effects on TKV increase and eGFR decline in Class 1D and E with a smaller number of patients. CONCLUSIONS: Strategies for prognostic enrichment, such as image classification, should be used in the design of RCTs for ADPKD to increase their power and reduce their cost.

Modulation of Polycystic Kidney Disease Severity by Phosphodiesterase 1 and 3 Subfamilies.

Aberrant intracellular calcium levels and increased cAMP signaling contribute to the development of polycystic kidney disease (PKD). cAMP can be hydrolyzed by various phosphodiesterases (PDEs). To examine the role of cAMP hydrolysis and the most relevant PDEs in the pathogenesis of PKD, we examined cyst development in Pde1- or Pde3-knockout mice on the Pkd2(-/WS25) background (WS25 is an unstable Pkd2 allele). These PDEs were selected because of their importance in cross-talk between calcium and cyclic nucleotide signaling (PDE1), control of cell proliferation and cystic fibrosis transmembrane conductance regulator (CFTR) -driven fluid secretion (PDE3), and response to vasopressin V2 receptor activation (both). In Pkd2(-/WS25) mice, knockout of Pde1a, Pde1c, or Pde3a but not of Pde1b or Pde3b aggravated the development of PKD and was associated with higher levels of protein kinase A-phosphorylated (Ser133) cAMP-responsive binding protein (P-CREB), activating transcription factor-1, and CREB-induced CRE modulator proteins in kidney nuclear preparations. Immunostaining also revealed higher expression of P-CREB in Pkd2(-/) (WS25);Pde1a(-/-), Pkd2(-) (/WS25);Pde1c(-/-), and Pkd2(-/) (WS25);Pde3a(-/-) kidneys. The cystogenic effect of desmopressin administration was markedly enhanced in Pkd2(-/WS25);Pde3a(-/-) mice, despite PDE3 accounting for only a small fraction of renal cAMP PDE activity. These observations show that calcium- and calmodulin-dependent PDEs (PDE1A and PDE1C) and PDE3A modulate the development of PKD, possibly through the regulation of compartmentalized cAMP pools that control cell proliferation and CFTR-driven fluid secretion. Treatments capable of increasing the expression or activity of these PDEs may, therefore, retard the development of PKD.

Predicted Mutation Strength of Nontruncating PKD1 Mutations Aids Genotype-Phenotype Correlations in Autosomal Dominant Polycystic Kidney Disease.

Autosomal dominant polycystic kidney disease (ADPKD) often results in ESRD but with a highly variable course. Mutations to PKD1 or PKD2 cause ADPKD; both loci have high levels of allelic heterogeneity. We evaluated genotype-phenotype correlations in 1119 patients (945 families) from the HALT Progression of PKD Study and the Consortium of Radiologic Imaging Study of PKD Study. The population was defined as: 77.7% PKD1, 14.7% PKD2, and 7.6% with no mutation detected (NMD). Phenotypic end points were sex, eGFR, height-adjusted total kidney volume (htTKV), and liver cyst volume. Analysis of the eGFR and htTKV measures showed that the PKD1 group had more severe disease than the PKD2 group, whereas the NMD group had a PKD2-like phenotype. In both the PKD1 and PKD2 populations, men had more severe renal disease, but women had larger liver cyst volumes. Compared with nontruncating PKD1 mutations, truncating PKD1 mutations associated with lower eGFR, but the mutation groups were not differentiated by htTKV. PKD1 nontruncating mutations were evaluated for conservation and chemical change and subdivided into strong (mutation strength group 2 [MSG2]) and weak (MSG3) mutation groups. Analysis of eGFR and htTKV measures showed that patients with MSG3 but not MSG2 mutations had significantly milder disease than patients with truncating cases (MSG1), an association especially evident in extreme decile populations. Overall, we have quantified the contribution of genic and PKD1 allelic effects and sex to the ADPKD phenotype. Intrafamilial correlation analysis showed that other factors shared by families influence htTKV, with these additional genetic/environmental factors significantly affecting the ADPKD phenotype.

Utilizing magnetization transfer imaging to investigate tissue remodeling in a murine model of autosomal dominant polycystic kidney disease.

PURPOSE: Noninvasive imaging techniques that quantify renal tissue composition are needed to more accurately ascertain prognosis and monitor disease progression in polycystic kidney disease (PKD). Given the success of magnetization transfer (MT) imaging to characterize various tissue remodeling pathologies, it was tested on a murine model of autosomal dominant PKD. METHODS: C57Bl/6 Pkd1 R3277C mice at 9, 12, and 15 months were imaged with a 16.4T MR imaging system. Images were acquired without and with RF saturation in order to calculate MT ratio (MTR) maps. Following imaging, the mice were euthanized and kidney sections were analyzed for cystic and fibrotic indices, which were compared with statistical parameters of the MTR maps. RESULTS: The MTR-derived mean, median, 25th percentile, skewness, and kurtosis were all closely related to indices of renal pathology, including kidney weight/body weight, cystic index, and percent of remaining parenchyma. The correlation between MTR and histology-derived cystic and fibrotic changes was R(2) = 0.84 and R(2) = 0.70, respectively. CONCLUSION: MT imaging provides a new, noninvasive means of measuring tissue remodeling PKD changes and may be better suited for characterizing renal impairment compared with conventional MR techniques.

Automatic total kidney volume measurement on follow-up magnetic resonance images to facilitate monitoring of autosomal dominant polycystic kidney disease progression.

BACKGROUND: Renal imaging examinations provide high-resolution information about the anatomic structure of the kidneys and are used to measure total kidney volume (TKV) in autosomal dominant polycystic kidney disease (ADPKD) patients. TKV has become the gold-standard image biomarker for ADPKD progression at early stages of the disease and is used in clinical trials to characterize treatment efficacy. Automated methods to segment the kidneys and measure TKV are desirable because of the long time requirement for manual approaches such as stereology or planimetry tracings. However, ADPKD kidney segmentation is complicated by a number of factors, including irregular kidney shapes and variable tissue signal at the kidney borders. METHODS: We describe an image processing approach that overcomes these problems by using a baseline segmentation initialization to provide automatic segmentation of follow-up scans obtained years apart. We validated our approach using 20 patients with complete baseline and follow-up T1-weighted magnetic resonance images. Both manual tracing and stereology were used to calculate TKV, with two observers performing manual tracings and one observer performing repeat tracings. Linear correlation and Bland-Altman analysis were performed to compare the different approaches. RESULTS: Our automated approach measured TKV at a level of accuracy (mean difference ± standard error = 0.99 ± 0.79%) on par with both intraobserver (0.77 ± 0.46%) and interobserver variability (1.34 ± 0.70%) of manual tracings. All approaches had excellent agreement and compared favorably with ground-truth manual tracing with interobserver, stereological and automated approaches having 95% confidence intervals ∼ ± 100 mL. CONCLUSIONS: Our method enables fast, cost-effective and reproducible quantification of ADPKD progression that will facilitate and lower the costs of clinical trials in ADPKD and other disorders requiring accurate, longitudinal kidney quantification. In addition, it will hasten the routine use of TKV as a prognostic biomarker in ADPKD.

Effect of genotype on the severity and volume progression of polycystic liver disease in autosomal dominant polycystic kidney disease.

BACKGROUND: The autosomal dominant polycystic kidney disease (APDKD) genotype influences renal phenotype severity but its effect on polycystic liver disease (PLD) is unknown. Here we analyzed the influence of genotype on liver phenotype severity. METHODS: Clinical data were retrieved from electronic records of patients who were mutation screened with the available liver imaging (n = 434). Liver volumes were measured by stereology (axial or coronal images) and adjusted to height (HtLV). RESULTS: Among the patients included, 221 (50.9%) had truncating PKD1 (PKD1-T), 141 (32.5%) nontruncating PKD1 (PKD1-NT) and 72 (16.6%) PKD2 mutations. Compared with PKD1-NT and PKD2, patients with PKD1-T had greater height-adjusted total kidney volumes (799 versus 610 and 549 mL/m; P < 0.001). HtLV was not different (1042, 1095 and 1058 mL/m; P = 0.64) between the three groups, but females had greater HtLVs compared with males (1114 versus 1015 mL/m; P < 0.001). Annualized median liver growth rates were 1.68, 1.5 and 1.24% for PKD1-T, PKD1-NT and PKD2 mutations, respectively (P = 0.49), and remained unaffected by the ADPKD genotype when adjusted for age, gender and baseline HtLV. Females <48 years of age had higher annualized growth rates compared with those who were older (2.65 versus 0.09%; P < 0.001). After age 48 years, 58% of females with severe PLD had regression of HtLV, while HtLV continued to increase in males. CONCLUSIONS: In contrast to the renal phenotype, the ADPKD genotype was not associated with the severity or growth rate of PLD in ADKPD patients. This finding, along with gender influence, indicates that modifiers beyond the disease gene significantly influence the liver phenotype.

Volume regression of native polycystic kidneys after renal transplantation.

BACKGROUND: The natural course of native kidneys after renal transplantation (RT) or dialysis in patients with autosomal dominant polycystic kidney disease (ADPKD) remains poorly understood. METHODS: We measured the total volumes of native kidneys and liver in 78 and 68 ADPKD patients, respectively, who had pre-transplant (within 2 years) and at least one post-transplant computed tomography (CT)/magnetic resonance imaging (MRI); in 40 patients with at least two post-transplant but no pre-transplant CT/MRIs; in 9 patients on chronic hemodialysis with at least one CT/MRI before and after beginning dialysis; and in 5 patients who had no image before and more than one image after dialysis. The last imaging was used in patients with multiple studies. RESULTS: Mean total kidney volume (TKV) ( ± SD) prior to transplantation was 3187 ± 1779 mL in the 78 patients who had imaging before and after transplantation and decreased by 20.2, 28.6, 38.3 and 45.8% after 0.5-1 (mean 0.7), 1-3 (1.8), 3-10 (5.7) and >10 (12.6) years, respectively. In the multivariable analysis, time on dialysis prior to RT and time from baseline to transplantation were negatively associated with reduction in TKV, whereas estimated glomerular filtration rate (eGFR) after transplantation and time from transplantation were positively associated with percent reduction in TKV. In the 40 patients with imaging only after transplantation, TKV decreased by 3.2 ± 16.3% between 7.2 ± 6.0 and 11.2 ± 6.8 years after transplantation (P < 0.001). TKV was 11.2 ± 35.6% higher (P = NS) after a follow-up of 3.4 ± 2.0 years in the 9 patients with imaging before and after initiation of hemodialysis and 3.4 ± 40.2% lower (P = NS) in the 5 patients with imaging between 2.0 ± 2.1 and 3.5 ± 3.6 years after initiation of hemodialysis. In the 68 patients with liver measurements, volume increased by 5.8 ± 17.9% between baseline and follow-up at 3.7 ± 3.8 years after transplantation (P = 0.009). CONCLUSIONS: TKV of native polycystic kidneys decreases substantially after RT. The reduction occurs mainly during the early post-transplantation period and more slowly thereafter.

Identification of Biomarkers for PKD1 Using Urinary Exosomes.

Autosomal dominant polycystic kidney disease (ADPKD) is a common cause of ESRD. Affected individuals inherit a defective copy of either PKD1 or PKD2, which encode polycystin-1 (PC1) or polycystin-2 (PC2), respectively. PC1 and PC2 are secreted on urinary exosome-like vesicles (ELVs) (100-nm diameter vesicles), in which PC1 is present in a cleaved form and may be complexed with PC2. Here, label-free quantitative proteomic studies of urine ELVs in an initial discovery cohort (13 individuals with PKD1 mutations and 18 normal controls) revealed that of 2008 ELV proteins, 9 (0.32%) were expressed at significantly different levels in samples from individuals with PKD1 mutations compared to controls (P<0.03). In samples from individuals with PKD1 mutations, levels of PC1 and PC2 were reduced to 54% (P<0.02) and 53% (P<0.001), respectively. Transmembrane protein 2 (TMEM2), a protein with homology to fibrocystin, was 2.1-fold higher in individuals with PKD1 mutations (P<0.03). The PC1/TMEM2 ratio correlated inversely with height-adjusted total kidney volume in the discovery cohort, and the ratio of PC1/TMEM2 or PC2/TMEM2 could be used to distinguish individuals with PKD1 mutations from controls in a confirmation cohort. In summary, results of this study suggest that a test measuring the urine exosomal PC1/TMEM2 or PC2/TMEM2 ratio may have utility in diagnosis and monitoring of polycystic kidney disease. Future studies will focus on increasing sample size and confirming these studies. The data were deposited in the ProteomeXchange (identifier PXD001075).

Imaging classification of autosomal dominant polycystic kidney disease: a simple model for selecting patients for clinical trials.

The rate of renal disease progression varies widely among patients with autosomal dominant polycystic kidney disease (ADPKD), necessitating optimal patient selection for enrollment into clinical trials. Patients from the Mayo Clinic Translational PKD Center with ADPKD (n=590) with computed tomography/magnetic resonance images and three or more eGFR measurements over ≥6 months were classified radiologically as typical (n=538) or atypical (n=52). Total kidney volume (TKV) was measured using stereology (TKVs) and ellipsoid equation (TKVe). Typical patients were randomly partitioned into development and internal validation sets and subclassified according to height-adjusted TKV (HtTKV) ranges for age (1A-1E, in increasing order). Consortium for Radiologic Imaging Study of PKD (CRISP) participants (n=173) were used for external validation. TKVe correlated strongly with TKVs, without systematic underestimation or overestimation. A longitudinal mixed regression model to predict eGFR decline showed that log2HtTKV and age significantly interacted with time in typical patients, but not in atypical patients. When 1A-1E classifications were used instead of log2HtTKV, eGFR slopes were significantly different among subclasses and, except for 1A, different from those in healthy kidney donors. The equation derived from the development set predicted eGFR in both validation sets. The frequency of ESRD at 10 years increased from subclass 1A (2.4%) to 1E (66.9%) in the Mayo cohort and from 1C (2.2%) to 1E (22.3%) in the younger CRISP cohort. Class and subclass designations were stable. An easily applied classification of ADPKD based on HtTKV and age should optimize patient selection for enrollment into clinical trials and for treatment when one becomes available.