Patient Engagement and Patient Experience Data in Regulatory Review and Health Technology Assessment: A Global Landscape Review.

BACKGROUND: Working with patients through meaningful patient engagement (PE) and incorporating patient experience data (PXD) is increasingly important in medicines and medical device development. However, PE in the planning, organization, generation, and interpretation of PXD within regulatory and health technology assessment (HTA) decision-making processes remains challenging. We conducted a global review of the PE and PXD landscape to identify evolving resources by geography to support and highlight the potential of integration of PE and PXD in regulatory assessment and HTA. METHODS: A review of literature/public information was conducted (August 2021-January 2023), led by a multistakeholder group comprising those with lived or professional experience of PE and PXD, to identify relevant regulatory and HTA initiatives and resources reviewed and categorized by geography and focus area. RESULTS: Overall, 53 relevant initiatives/resources were identified (global, 14; North America, 11; Europe, 11; Asia, nine; UK, six; Latin America, one; Africa, one). Most focused either on PE (49%) or PXD (28%); few (11%) mentioned both PE and PXD (as largely separate activities) or demonstrated an integration of PE and PXD (11%). CONCLUSIONS: Our analysis demonstrates increasing interest in PE, PXD, and guidance on their use individually in decision-making. However, more work is needed to offer guidance on maximizing the value of patient input into decisions by combining both PE and PXD into regulatory and HTA processes; the necessity of integrating PE in the design and interpretation of PXD programs should be highlighted. A co-created framework to achieve this integration is part of a future project.

Fulfillment and Validity of the Kidney Health Evaluation Measure for People with Diabetes.

Objective: To evaluate the fulfillment and validity of the kidney health evaluation for people with diabetes (KED) Healthcare Effectiveness Data Information Set (HEDIS) measure. Patients and Methods: Optum Labs Data Warehouse (OLDW) was used to identify the nationally distributed US population aged 18 years and older, with diabetes, between January 1, 2017, and December 31, 2017. The OLDW includes deidentified medical, pharmacy, laboratory, and electronic health record (EHR) data. The KED fulfillment was defined in 2017 as both estimated glomerular filtration rate (eGFR) and urinary albumin-creatinine ratio testing within the measurement year. The KED validity was assessed using bivariate analyses of KED fulfillment with diabetes care measures in 2017 and chronic kidney disease (CKD) diagnosis and evidence-based kidney protective interventions in 2018. Results: Among eligible 5,635,619 Medicare fee-for-service beneficiaries, 736,875 Medicare advantage (MA) beneficiaries, and 660,987 commercial patients, KED fulfillment was 32.2%, 38.7%, and 37.7%, respectively. Albuminuria testing limited KED fulfillment with urinary albumin-creatinine ratio testing (<40%) and eGFR testing (>90%). The KED fulfillment was positively associated with receipt of diabetes care in 2017, CKD diagnosis in 2018, and evidence-based kidney protective interventions in 2018. The KED fulfillment trended lower for Black race, Medicare-Medicaid dual eligibility status, low neighborhood income, and low education status. Conclusion: Less than 40% of adults with diabetes received guideline-recommended testing for CKD in 2017. Routine KED was associated with diabetes care and evidence-based CKD interventions. Increasing guideline-recommended testing for CKD among people with diabetes should lead to timely and equitable CKD detection and treatment.

Serum Magnesium Levels and Cardiovascular Outcomes in Systolic Blood Pressure Intervention Trial Participants.

Rationale and Objective: Serum magnesium levels have been inversely yet inconsistently associated with cardiovascular (CV) outcomes. In this study, we examined the association of serum magnesium levels with CV outcomes in the Systolic Blood Pressure Intervention Trial (SPRINT) participants. Study Design: Case-control post hoc analysis of SPRINT. Setting & Participants: A total of 2,040 SPRINT participants with available serum samples at baseline level were included in this study. Case participants (n = 510) who experienced a CV event during the SPRINT observation period (median follow-up of 3.2 years) and control participants (n = 1,530) without CV events were sampled in a 1:3 ratio for measurements of serum magnesium level at baseline and 2-year follow-up. Predictors: Baseline serum magnesium levels and 2-year percentage change in serum magnesium levels (ΔSMg). Outcome: SPRINT primary composite CV outcome. Analytical Approach: Multivariable conditional logistic regression analysis, accounting for matching factors, was used to evaluate the association of baseline and ΔSMg with CV outcomes. Individual matching of cases and controls was based on the SPRINT treatment arm allocation (standard vs intensive) and prevalence of chronic kidney disease (CKD). Results: The median serum magnesium level at baseline was similar among the case and control groups. In a fully adjusted model, each standard deviation (SD) (0.18 mg/dL) higher of the baseline serum magnesium level was independently associated with a lower risk for composite CV outcomes in all study participants (adjusted odds ratio 95% CI, 0.79 [0.70-0.89]). This association was similar when serum magnesium levels were analyzed in quartiles but dissipated in the standard (vs intensive) arm of SPRINT (0.88 [0.76-1.02] vs 0.65 [0.53-0.79], respectively; Pinteraction = 0.06). The presence or absence of CKD at baseline did not modify this association. ΔSMg was not independently associated with CV outcomes occurring after 2 years. Limitations: ΔSMg was small in magnitude, limiting effect size. Conclusions: Higher baseline serum magnesium levels were independently associated with reduced risk for CV outcomes in all study participants, but ΔSMg was not associated with CV outcomes.

A National Registry for People With All Stages of Kidney Disease: The National Kidney Foundation (NKF) Patient Network.

RATIONALE & OBJECTIVE: The National Kidney Foundation (NKF) launched the first national US kidney disease patient registry, the NKF Patient Network, that is open to patients throughout the continuum of chronic kidney disease (CKD). The Network provides individualized education and will facilitate patient-centered research, clinical care, and health policy decisions. Here, we present the overall design and the results of a feasibility study that was conducted July through December 2020. STUDY DESIGN: Longitudinal observational cohort study of patient-entered data with or without electronic health care record (EHR) linkage in collaboration with health systems. SETTING & PARTICIPANTS: People with CKD, age≥18 years, are invited through their provider, NKF communications, or national outreach campaign. People self-enroll and share their data through a secure portal that offers individualized education and support. The first health system partner is Geisinger. EXPOSURE: Any cause and stage of CKD, including dialysis and kidney transplant recipients. OUTCOME: Feasibility of the EHR data transfer, participants' characteristics, and their perspectives on usability and content. ANALYTICAL APPROACH: Data were collected and analyzed through the registry portal powered by the Pulse Infoframe healthie 2.0 platform. RESULTS: During the feasibility study, 80 participants completed their profile, and 42 completed a satisfaction survey. Mean age was 57.5 years, 51% were women, 83% were White, and 89% were non-Hispanic or Latino. Of the participants, 60% were not aware of their level of estimated glomerular filtration rate and 91% of their urinary albumin-creatinine ratio. LIMITATIONS: Challenges for the Network are lack of awareness of kidney disease for many with CKD, difficulty in recruiting vulnerable populations or those with low digital readiness, and loss to follow-up, all leading to selection bias. CONCLUSIONS: The Network is positioned to become a national and international platform for real-world data that can inform the development of patient-centered research, care, and treatments.

Framework From a Multidisciplinary Approach for Transitioning Variants of Unknown Significance From Clinical Genetic Testing in Kidney Disease to a Definitive Classification.

Introduction: Monogenic causes in over 300 kidney-associated genes account for approximately 12% of end stage kidney disease (ESKD) cases. Advances in sequencing and large customized panels enable the noninvasive diagnosis of monogenic kidney disease at relatively low cost, thereby allowing for more precise management for patients and their families. A major challenge is interpreting rare variants, many of which are classified as variants of unknown significance (VUS). We present a framework in which we thoroughly evaluated and provided evidence of pathogenicity for HNF1B-p.Arg303His, a VUS returned from clinical diagnostic testing for a kidney transplant candidate. Methods: A blueprint was designed by a multidisciplinary team of clinicians, molecular biologists, and diagnostic geneticists. The blueprint included using a health system-based cohort with genetic and clinical information to perform deep phenotyping of VUS heterozygotes to identify the candidate VUS and rule out other VUS, examination of existing genetic databases, as well as functional testing. Results: Our approach demonstrated evidence for pathogenicity for HNF1B-p.Arg303His by showing similar burden of kidney manifestations in this variant to known HNF1B pathogenic variants, and greater burden compared to noncarriers. Conclusion: Determination of a molecular diagnosis for the example family allows for proper surveillance and management of HNF1B-related manifestations such as kidney disease, diabetes, and hypomagnesemia with important implications for safe living-related kidney donation. The candidate gene-variant pair also allows for clinical biomarker testing for aberrations of linked pathways. This working model may be applicable to other diseases of genetic etiology.

Physiologic Regulation of Systemic Klotho Levels by Renal CaSR Signaling in Response to CaSR Ligands and pH o.

BACKGROUND: The kidney is the source of sKlotho and kidney-specific loss of Klotho leads to a phenotype resembling the premature multiorgan failure phenotype in Klotho-hypomorphic mice ( kl/kl mice). Klotho and the Ca-sensing receptor (CaSR) are highly expressed in the distal convoluted tubule (DCT). The physiologic mechanisms that regulate sKlotho levels are unknown. METHODS: We measured sKlotho in WT and tubule-specific CaSR -/- (TS-CaSR -/- ) mice treated with calcimimetics, alkali, or acid, and Klotho shed from minced mouse kidneys, and from HEK-293 cells expressing the CaSR and Klotho, in response to calcimimetics, calcilytics, alkalotic and acidic pH, and ADAM protease inhibitors. The CaSR, Klotho, and ADAM10 were imaged in mouse kidneys and cell expression systems using confocal microscopy. RESULTS: The CaSR, Klotho, and ADAM10 colocalize on the basolateral membrane of the DCT. Calcimimetics and HCO 3 increase serum sKlotho levels in WT but not in CaSR -/- mice, and acidic pH suppresses sKlotho levels in WT mice. In minced kidneys and cultured cells, CaSR activation with high Ca, calcimimetics, or alkali increase shed Klotho levels via ADAM10, as demonstrated using the ADAM10 inhibitor GI254023X and siRNA. In cultured cells, the CaSR, Klotho, and ADAM10 form cell surface aggregates that disperse after CaSR activation. CONCLUSIONS: We identify a novel physiologic mechanism for regulation of sKlotho levels by the renal CaSR-ADAM10-Klotho pathway. We show that CaSR activators, including alkali, increase renal CaSR-stimulated Klotho shedding and predict that this mechanism is relevant to the effects of acidosis and alkali therapy on CKD progression.

Chronic Kidney Disease Testing Among At-Risk Adults in the U.S. Remains Low: Real-World Evidence From a National Laboratory Database.

OBJECTIVE: An estimated 37 million Americans have chronic kidney disease (CKD). Nearly 90% do not know about their condition because of low awareness about the importance of CKD testing and diagnosis among practitioners and people at risk for CKD. This study uses data from a national clinical laboratory to identify guideline-recommended CKD testing rates across the U.S. RESEARCH DESIGN AND METHODS: Patients with Laboratory Corporation of America Holdings (Labcorp) testing between 2013 and 2019 were defined as at risk for CKD if they had any testing ordered with diagnosis codes for diabetes and/or hypertension. Guideline-concordant CKD assessment was defined by estimated glomerular filtration rate (eGFR) and urine albumin-to-creatinine ratio (uACR) testing within the study year. RESULTS: We identified 28,295,982 at-risk patients (mean age 60.6 ± 14.8 years; 53.6% women): 16.2% had diabetes, 63.8% had hypertension, and 20.1% had both comorbidities. Of these, 80.3% did not receive guideline-concordant assessment during the study period. Furthermore, only 21.0% had uACR testing versus 89.6% with eGFR. CKD assessment occurred at least once in 28.7% of patients with diabetes, 10.5% of patients with hypertension, and 41.4% of patients with both conditions. In a state-by-state comparison, annual testing rates ranged from 5 to 30%. The nationwide rate increased modestly each year between 2013 and 2018 (from 10.7% to 15.2%). CONCLUSIONS: Despite guideline recommendations, testing for CKD with uACR and eGFR in U.S. adults with diabetes and hypertension is low in routine clinical care. These data highlight the need for strategies to improve routine CKD assessment nationwide.

International Medical Graduates in Nephrology: A Guide for Trainees and Programs.

A significant portion of the nephrology workforce has traditionally consisted of non-United States (US) citizen international medical graduates and international research trainees. Although international medical graduates are offered opportunities for training and professional growth that are beyond those available in their countries of origin, they typically encounter barriers to transition from training to practice and early-stage career development. In this article, we describe the exchange visitor and temporary worker visas granted to foreign trainees in the United States, focusing on the transition from training to nephrology practice and/or research. While we provide general recommendations on how to navigate this tedious and unpredictable process for both programs and trainees, consultation with immigration attorneys is indispensable for a successful outcome. Trainees are therefore encouraged to seek continuous support from their programs/sponsors and assistance from immigration representatives at their training institutions. We provide a positive message to emphasize that there are many pathways to arrive at the desired post-training destination.

Interstitial microRNA miR-214 attenuates inflammation and polycystic kidney disease progression.

Renal cysts are the defining feature of autosomal dominant polycystic kidney disease (ADPKD); however, the substantial interstitial inflammation is an often-overlooked aspect of this disorder. Recent studies suggest that immune cells in the cyst microenvironment affect ADPKD progression. Here we report that microRNAs (miRNAs) are new molecular signals in this crosstalk. We found that miR-214 and its host long noncoding RNA Dnm3os are upregulated in orthologous ADPKD mouse models and cystic kidneys from humans with ADPKD. In situ hybridization revealed that interstitial cells in the cyst microenvironment are the primary source of miR-214. While genetic deletion of miR-214 does not affect kidney development or homeostasis, surprisingly, its inhibition in Pkd2- and Pkd1-mutant mice aggravates cyst growth. Mechanistically, the proinflammatory TLR4/IFN-γ/STAT1 pathways transactivate the miR-214 host gene. miR-214, in turn as a negative feedback loop, directly inhibits Tlr4. Accordingly, miR-214 deletion is associated with increased Tlr4 expression and enhanced pericystic macrophage accumulation. Thus, miR-214 upregulation is a compensatory protective response in the cyst microenvironment that restrains inflammation and cyst growth.

Renal tubular cell spliced X-box binding protein 1 (Xbp1s) has a unique role in sepsis-induced acute kidney injury and inflammation.

Sepsis is a systemic inflammatory state in response to infection, and concomitant acute kidney injury (AKI) increases mortality significantly. Endoplasmic reticulum stress is activated in many cell types upon microbial infection and modulates inflammation. The role of endoplasmic reticulum signaling in the kidney during septic AKI is unknown. Here we tested the role of the spliced X-box binding protein 1 (Xbp1s), a key component of the endoplasmic reticulum stress-activated pathways, in the renal response to sepsis in the lipopolysaccharide (LPS) model. Xbp1s was increased in the kidneys of mice treated with LPS but not in other models of AKI, or several chronic kidney disease models. The functional significance of Xbp1s induction was examined by genetic manipulation in renal tubules. Renal tubule-specific overexpression of Xbp1s caused severe tubule dilation and vacuolation with expression of the injury markers Kim1 and Ngal, the pro-inflammatory molecules interleukin-6 (Il6) and Toll-like receptor 4 (Tlr4), decreased kidney function and 50% mortality in five days. Renal tubule-specific genetic ablation of Xbp1 had no phenotype at baseline. However, after LPS, Xbp1 knockdown mice displayed lower renal NGAL, pro-apoptotic factor CHOP, serum creatinine levels, and a tendency towards lower Tlr4 compared to LPS-treated mice with intact Xbp1s. LPS treatment in Xbp1s-overexpressing mice caused a mild increase in NGAL and CHOP compared to LPS-treated mice without genetic Xbp1s overexpression. Thus, increased Xbp1s signaling in renal tubules is unique to sepsis-induced AKI and contributes to renal inflammation and injury. Inhibition of this pathway may be a potential portal to alleviate injury.

Low serum magnesium is associated with faster decline in kidney function: the Dallas Heart Study experience.

Hypomagnesemia associates with inflammation and risk of diabetes and hypertension, which may contribute to kidney function decline. We hypothesized that low serum magnesium (SMg) levels independently associate with a significant decline in estimated glomerular filtration rate (eGFR). We analyzed SMg levels in 2056 participants from the Dallas Heart Study, a longitudinal, population-based, multiethnic, cohort study involving residents of Dallas County, Texas, USA. The primary study outcome was the change in eGFR using multivariable linear regression models adjusted for demographics, anthropometric and biochemical parameters, medications, C reactive protein levels, prevalent hypertension and diabetes. During a median follow-up of 7.0 years (25th, 75th percentile: 6.5, 7.6), the median decrease in eGFR was -0.71 (25th, 75th percentile: -2.43, +0.68) mL/min/1.73 m2 per year in the entire cohort. In a fully adjusted model, the lowest SMg quintile (≤1.9 mg/dL or ≤0.8 mM) was associated with a -0.50 mL/min/1.73 m2 per year drop in eGFR (95% CI -0.95 to -0.05; p=0.028) compared with the highest SMg quintile (≥2.3 mg/dL or ≥1.0 mM). Every 0.2 mg/dL (0.08 mM) decrease in SMg was associated with an eGFR decline of -0.23 mL/min/1.73 m2 per year (95% CI -0.38 to -0.08; p=0.003), a decline that was more pronounced in participants with prevalent diabetes compared with patients without diabetes (-0.51 vs -0.18 mL/min/1.73 m2 per year, respectively). In conclusion, low SMg was independently associated with eGFR decline. Further studies are needed to determine whether Mg repletion can ameliorate inflammation, lower blood pressure and serum glucose and ultimately prevent or retard kidney function decline.

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.

Association of serum magnesium with all-cause mortality in patients with and without chronic kidney disease in the Dallas Heart Study.

Background: Low serum magnesium (SMg) has been linked to increased mortality and cardiovascular disease (CVD) in the general population. We examined whether this association is similar in participants with versus without prevalent chronic kidney disease (CKD) in the multiethnic Dallas Heart Study (DHS) cohort. Methods: SMg was analyzed as a continuous variable and divided into tertiles. Study outcomes were all-cause death, cardiovascular (CV) death or event, and CVD surrogate markers, evaluated using multivariable Cox regression models adjusted for demographics, comorbidity, anthropometric and biochemical parameters including albumin, phosphorus and parathyroid hormone, and diuretic use. Median follow-up was 12.3 years (11.9-12.8, 25th percentile-75th percentile). Results: Among 3551 participants, 306 (8.6%) had prevalent CKD. Mean SMg was 2.08 ± 0.19 mg/dL (0.85 ± 0.08 mM, mean ± SD) in the CKD and 2.07 ± 0.18 mg/dL (0.85 ± 0.07 mM) in the non-CKD subgroups. During the follow-up period, 329 all-cause deaths and 306 CV deaths or events occurred. In a fully adjusted model, every 0.2 mg/dL decrease in SMg was associated with ∼20-40% increased hazard for all-cause death in both CKD and non-CKD subgroups. In CKD participants, the lowest SMg tertile was also independently associated with all-cause death (adjusted hazard ratio 2.31; 95% confidence interval 1.23-4.36 versus 1.15; 0.55-2.41; for low versus high tertile, respectively). Conclusions: Low SMg levels (1.4-1.9 mg/dL; 0.58-0.78 mM) were independently associated with all-cause death in patients with prevalent CKD in the DHS cohort. Randomized clinical trials are important to determine whether Mg supplementation affects survival in CKD patients.

Effect of urine pH and magnesium on calcium oxalate saturation.

Hypomagnesiuria is a common biochemical finding in patients with calcium oxalate (CaOx) nephrolithiasis. Clinical trials using Mg supplements as therapy against CaOx stones have shown mixed results. We tested the effect of Mg administration in healthy subjects under conditions of controlled urine pH (UpH) on urinary Ca excretion rate (UCaV) and CaOx saturation. This is a 4-phase, double blind, placebo-controlled, metabolic crossover study performed in healthy volunteers. Mg lactate (MgLact2) was used as Mg supplement. High UpH and low UpH were achieved by administration of potassium citrate (K3Citrate) and ammonium chloride (NH4Cl), respectively, with potassium balance maintained by KCl. Eight participants completed 4 phases of study. The interventions successfully modulated 24-h UpH (7.0 ± 0.4 vs. 5.7 ± 0.6 in high vs low pH phases; P<0.001). Administration of MgLact2 increased UMgV [175.8 ± 40.2 vs 93.4 ± 39.7 mg/day (7.2 ± 1.7 vs 3.8 ± 1.6 mmol/day), high vs low Mg phase; P<0.001], and increased pH both at low (5.6 ± 0.5 to 5.8 ± 0.7; P = 0.02) and high UpH (6.9 ± 0.4 to 7.0 ± 0.3; P = 0.01). At a given urine pH, Mg supplementation marginally increased UCaV, but did not alter UOxV or CaOx saturation. Provision of an alkali load significantly lowered UCaV and saturation of CaOx at any level of UMgV. Compared to changes in UMgV, changes in UpH play a more significant role in determining urine CaOx saturation in healthy subjects. Mg supplements are likely to reduce CaOx saturation if they also raise urine pH.

Mutations in PCBD1 cause hypomagnesemia and renal magnesium wasting.

Mutations in PCBD1 are causative for transient neonatal hyperphenylalaninemia and primapterinuria (HPABH4D). Until now, HPABH4D has been regarded as a transient and benign neonatal syndrome without complications in adulthood. In our study of three adult patients with homozygous mutations in the PCBD1 gene, two patients were diagnosed with hypomagnesemia and renal Mg(2+) loss, and two patients developed diabetes with characteristics of maturity onset diabetes of the young (MODY), regardless of serum Mg(2+) levels. Our results suggest that these clinical findings are related to the function of PCBD1 as a dimerization cofactor for the transcription factor HNF1B. Mutations in the HNF1B gene have been shown to cause renal malformations, hypomagnesemia, and MODY. Gene expression studies combined with immunohistochemical analysis in the kidney showed that Pcbd1 is expressed in the distal convoluted tubule (DCT), where Pcbd1 transcript levels are upregulated by a low Mg(2+)-containing diet. Overexpression in a human kidney cell line showed that wild-type PCBD1 binds HNF1B to costimulate the FXYD2 promoter, the activity of which is instrumental in Mg(2+) reabsorption in the DCT. Of seven PCBD1 mutations previously reported in HPABH4D patients, five mutations caused proteolytic instability, leading to reduced FXYD2 promoter activity. Furthermore, cytosolic localization of PCBD1 increased when coexpressed with HNF1B mutants. Overall, our findings establish PCBD1 as a coactivator of the HNF1B-mediated transcription necessary for fine tuning FXYD2 transcription in the DCT and suggest that patients with HPABH4D should be monitored for previously unrecognized late complications, such as hypomagnesemia and MODY diabetes.

Early development of hyperparathyroidism due to loss of PTH transcriptional repression in patients with HNF1ß mutations?

CONTEXT: Heterozygous mutations or deletions of the transcription factor hepatocyte nuclear factor 1ß (HNF1ß) result in a heterogeneous syndrome characterized by renal cysts and diabetes, together with a variety of other extrarenal and renal manifestations. Interestingly, in several patients with HNF1ß abnormalities, we observed early hyperparathyroidism and PTH levels that we judged inappropriately high compared with the degree of renal function decline. OBJECTIVE: Based on the above clinical observations, we tested the hypothesis of a direct role of HNF1ß in the transcriptional regulation of the human PTH gene in the parathyroid gland. DESIGN, SETTING, AND PATIENTS: Immunostaining of human parathyroid sections, RT-PCR, chromatin immunoprecipitation (ChIP), and luciferase reporter assays in human embryonic kidney cells (HEK293) were performed. We eventually report clinical data from all 11 HNF1ß patients known at our institute, 9 with heterozygous HNF1ß whole-gene deletions and 2 with heterozygous HNF1ß mutations. RESULTS: PTH levels were high in 8 patients. In 2 of these patients, the hyperparathyroidism was clearly appropriate for the level of kidney function, whereas PTH might be discrepant in the others. We demonstrated HNF1ß expression in PTH-positive cells of human parathyroid gland. Chromatin immunoprecipitation analysis showed that HNF1ß directly binds responsive elements within the human PTH promoter. Cotransfection of a PTH promoter- luciferase construct with a wild-type HNF1ß construct resulted in a maximal reduction of 30% of PTH promoter activity. Importantly, HNF1ß mutants lacked this inhibitory property. Serial deletions in the PTH promoter construct revealed that the inhibitory effect of HNF1ß resides between -200 and -70 bp from the transcription initiation site. CONCLUSIONS: Our data demonstrate that HNF1ß is a novel repressor of human PTH gene transcription, which could contribute to the development of hyperparathyroidism in patients with HNF1ß mutations or deletions.

Vitamin D down-regulates TRPC6 expression in podocyte injury and proteinuric glomerular disease.

The transient receptor potential cation channel C6 (TRPC6) is a slit diaphragm protein expressed by podocytes. TRPC6 gain-of-function mutations cause autosomal dominant focal segmental glomerulosclerosis. In acquired proteinuric renal disease, glomerular TRPC6 expression is increased. We previously demonstrated that acquired increased TRPC6 expression is ameliorated by antiproteinuric angiotensin receptor blockers and angiotensin-converting enzyme inhibitors. Vitamin D also has an antiproteinuric effect. We hypothesized that vitamin D reduces proteinuria by affecting TRPC6 expression in podocytes. Adriamycin-induced nephropathy increased TRPC6 mRNA and protein expression and induced proteinuria in rats. Treatment with 1,25-dihydroxyvitamin D3 (1,25-D3) normalized TRPC6 expression and reduced proteinuria. In vitro, podocyte injury induced by adriamycin exposure in cultured podocytes increased TRPC6 expression. Treatment of injured podocytes with 1,25-D3 dose dependently reduced adriamycin-induced TRPC6 expression. Chromatin immunoprecipitation analysis demonstrated that the vitamin D receptor directly binds to the TRPC6 promoter. Moreover, 1,25-D3 reduced TRPC6 promoter activity in a luciferase reporter assay. In 1,25-D3-deficient 25-hydroxy-1α-hydroxylase knockout mice, TRPC6 expression was increased, accompanied by podocyte foot process effacement and proteinuria. 1,25-D3 supplementation normalized TRPC6 expression, podocyte morphology, and proteinuria in these mice. These results demonstrate that vitamin D down-regulates the enhanced TRPC6 expression in in vivo and in vitro podocyte injury, possibly through a direct effect on TRPC6 promoter activity. This TRPC6 down-regulation could contribute to the antiproteinuric effect of vitamin D.

Sensing mechanisms involved in Ca2+ and Mg2+ homeostasis.

Calcium (Ca(2+)) and magnesium (Mg(2+)) ions are involved in many vital physiological functions. In the human body, Ca(2+) and Mg(2+) homeostatic systems rely on three components: (i) tissues (re)absorbing or storing Ca(2+) and Mg(2+), mainly kidney, intestine, and bone; (ii) hormones that modulate the transport and mobilization of these minerals; and (iii) sensors controlling the transport of Ca(2+) and Mg(2+) in tissues. So far, the Ca(2+)-sensing receptor (CaSR) is the only known sensing mechanism involved in the response to systemic and local fluctuations in Ca(2+) and Mg(2+) concentrations. The tight coupling of the Ca(2+) and Mg(2+) homeostasis is frequently observed in humans and in animal models. Moreover, during the past decade, the intense research on mineral disorders, seen in the course of chronic kidney disease with secondary hyperparathyroidism, has revealed the interplay between Ca(2+) and Mg(2+) homeostasis. Altogether this evidence points to the CaSR as an interesting target to study mutual disturbances in Ca(2+) and Mg(2+) balance. Although many homeostatic processes involving CaSR in the parathyroid glands and kidneys have been elucidated, the current knowledge of the inter-regulation of divalent handling by the CaSR is still limited. Insight into the systemic and local sensing of Ca(2+) and Mg(2+) by the CaSR, as well as the identification of other receptors/sensors for these cations, will unravel the mechanism controlling their coupled regulation, and will lead to new treatment concepts for mineral disorders.

Role of the distal convoluted tubule in renal Mg2+ handling: molecular lessons from inherited hypomagnesemia.

In healthy individuals, Mg(2+) homeostasis is tightly regulated by the concerted action of intestinal absorption, exchange with bone, and renal excretion. The kidney, more precisely the distal convoluted tubule (DCT), is the final determinant of plasma Mg(2+) concentrations. Positional cloning strategies in families with hereditary hypomagnesemia identified defects in several proteins localized in the DCT as causative factors. So far, the identified actors involved in Mg(2+) handling in the DCT include: the transient receptor potential channel melastatin member 6, the pro-epidermal growth factor, the thiazide-sensitive Na(+)-Cl(-) cotransporter, the γ-subunit of the Na(+)/K(+)-ATPase, the hepatocyte nuclear factor 1B, the potassium channels Kv1.1 and Kir4.1, and the cyclin M2. In the years to come, the identification of new magnesiotropic genes and related proteins will further clarify the role of the kidney in Mg(2+) homeostasis, and will potentially lead to new therapeutic approaches for hypomagnesemia.

HNF-1B specifically regulates the transcription of the γa-subunit of the Na+/K+-ATPase.

Hepatocyte nuclear factor-1B (HNF-1B) is a transcription factor involved in embryonic development and tissue-specific gene expression in several organs, including the kidney. Recently heterozygous mutations in the HNF1B gene have been identified in patients with hypomagnesemia due to renal Mg(2+) wasting. Interestingly, ChIP-chip data revealed HNF-1B binding sites in the FXYD2 gene, encoding the γ-subunit of the Na(+)/K(+)-ATPase. The γ-subunit has been described as one of the molecular players in the renal Mg(2+) reabsorption in the distal convoluted tubule (DCT). Of note, the FXYD2 gene can be alternatively transcribed into two main variants, namely γa and γb. In the present study, we demonstrated via two different reporter gene assays that HNF-1B specifically acts as an activator of the γa-subunit, whereas the γb-subunit expression was not affected. Moreover, the HNF-1B mutations H69fsdelAC, H324S325fsdelCA, Y352finsA and K156E, previously identified in patients with hypomagnesemia, prevented transcription activation of γa-subunit via a dominant negative effect on wild type HNF1-B. By immunohistochemistry, it was shown that the γa- and γb-subunits colocalize at the basolateral membrane of the DCT segment of mouse kidney. On the basis of these data, we suggest that abnormalities involving the HNF-1B gene may impair the relative abundance of γa and γb, thus affecting the transcellular Mg(2+) reabsorption in the DCT.