Using Genetics to Inform Interventions Related to Sodium and Potassium in Hypertension.

BACKGROUND: Hypertension is a key risk factor for major adverse cardiovascular events but remains difficult to treat in many individuals. Dietary interventions are an effective approach to lower blood pressure (BP) but are not equally effective across all individuals. BP is heritable, and genetics may be a useful tool to overcome treatment response heterogeneity. We investigated whether the genetics of BP could be used to identify individuals with hypertension who may receive a particular benefit from lowering sodium intake and boosting potassium levels. METHODS: In this observational genetic study, we leveraged cross-sectional data from up to 296 475 genotyped individuals drawn from the UK Biobank cohort for whom BP and urinary electrolytes (sodium and potassium), biomarkers of sodium and potassium intake, were measured. Biologically directed genetic scores for BP were constructed specifically among pathways related to sodium and potassium biology (pharmagenic enrichment scores), as well as unannotated genome-wide scores (conventional polygenic scores). We then tested whether there was a gene-by-environment interaction between urinary electrolytes and these genetic scores on BP. RESULTS: Genetic risk and urinary electrolytes both independently correlated with BP. However, urinary sodium was associated with a larger BP increase among individuals with higher genetic risk in sodium- and potassium-related pathways than in those with comparatively lower genetic risk. For example, each SD in urinary sodium was associated with a 1.47-mm Hg increase in systolic BP for those in the top 10% of the distribution of genetic risk in sodium and potassium transport pathways versus a 0.97-mm Hg systolic BP increase in the lowest 10% (P=1.95×10-3). This interaction with urinary sodium remained when considering estimated glomerular filtration rate and indexing sodium to urinary creatinine. There was no strong evidence of an interaction between urinary sodium and a standard genome-wide polygenic score of BP. CONCLUSIONS: The data suggest that genetic risk in sodium and potassium pathways could be used in a precision medicine model to direct interventions more specifically in the management of hypertension. Intervention studies are warranted.

Isocitrate dehydrogenase 1 upregulation in urinary extracellular vesicles from proximal tubules of type 2 diabetic rats.

Diabetic nephropathy (DN) is a major cause of chronic kidney disease. Microalbuminuria is currently the most common non-invasive biomarker for the early diagnosis of DN. However, renal structural damage may have advanced when albuminuria is detected. In this study, we sought biomarkers for early DN diagnosis through proteomic analysis of urinary extracellular vesicles (uEVs) from type 2 diabetic model rats and normal controls. Isocitrate dehydrogenase 1 (IDH1) was significantly increased in uEVs from diabetic model rats at the early stage despite minimal differences in albuminuria between the groups. Calorie restriction significantly suppressed the increase in IDH1 in uEVs and 24-hour urinary albumin excretion, suggesting that the increase in IDH1 in uEVs was associated with the progression of DN. Additionally, we investigated the origin of IDH1-containing uEVs based on their surface sugar chains. Lectin affinity enrichment and immunohistochemical staining showed that IDH1-containing uEVs were derived from proximal tubules. These findings suggest that the increase in IDH1 in uEVs reflects pathophysiological alterations in the proximal tubules and that IDH1 in uEVs may serve as a potential biomarker of DN in the proximal tubules.

Association of Albuminuria With Chronic Kidney Disease Progression in Persons With Chronic Kidney Disease and Normoalbuminuria : A Cohort Study.

BACKGROUND: Albuminuria is a major risk factor for chronic kidney disease (CKD) progression, especially when categorized as moderate (30 to 300 mg/g) or severe (>300 mg/g). However, there are limited data on the prognostic value of albuminuria within the normoalbuminuric range (<30 mg/g) in persons with CKD. OBJECTIVE: To estimate the increase in the cumulative incidence of CKD progression with greater baseline levels of albuminuria among persons with CKD who had normoalbuminuria (<30 mg/g). DESIGN: Multicenter prospective cohort study. SETTING: 7 U.S. clinical centers. PARTICIPANTS: 1629 participants meeting criteria from the CRIC (Chronic Renal Insufficiency Cohort) study with CKD (estimated glomerular filtration rate [eGFR], 20 to 70 mL/min/1.73 m2) and urine albumin-creatinine ratio (UACR) less than 30 mg/g. MEASUREMENTS: Baseline spot urine albumin divided by spot urine creatinine to calculate UACR as the exposure variable. The 10-year adjusted cumulative incidences of CKD progression (composite of 50% eGFR decline or kidney failure [dialysis or kidney transplantation]) from confounder adjusted survival curves using the G-formula. RESULTS: Over a median follow-up of 9.8 years, 182 of 1629 participants experienced CKD progression. The 10-year adjusted cumulative incidences of CKD progression were 8.7% (95% CI, 5.9% to 11.6%), 11.5% (CI, 8.8% to 14.3%), and 19.5% (CI, 15.4% to 23.5%) for UACR levels of 0 to less than 5 mg/g, 5 to less than 15 mg/g, and 15 mg/g or more, respectively. Comparing persons with UACR 15 mg/g or more to those with UACR 5 to less than 15 mg/g and 0 to less than 5 mg/g, the absolute risk differences were 7.9% (CI, 3.0% to 12.7%) and 10.7% (CI, 5.8% to 15.6%), respectively. The 10-year adjusted cumulative incidence increased linearly based on baseline UACR levels. LIMITATION: UACR was measured once. CONCLUSION: Persons with CKD and normoalbuminuria (<30 mg/g) had excess risk for CKD progression, which increased in a linear fashion with higher levels of albuminuria. PRIMARY FUNDING SOURCE: None.

Urinary CD4 + T Cells Predict Renal Relapse in ANCA-Associated Vasculitis.

SIGNIFICANCE STATEMENT: Early identification of patients at risk of renal flares in ANCA vasculitis is crucial. However, current clinical parameters have limitations in predicting renal relapse accurately. This study investigated the use of urinary CD4 + T lymphocytes as a predictive biomarker for renal flares in ANCA vasculitis. This study, including urine samples from 102 patients, found that the presence of urinary CD4 + T cells was a robust predictor of renal relapse within a 6-month time frame, with a sensitivity of 60% and a specificity of 97.8%. The diagnostic accuracy of urinary CD4 + T cells exceeded that of ANCA titers, proteinuria, and hematuria. Monitoring urinary CD4 + T lymphocytes could help assess the risk of future renal relapse, enabling early preventive measures and tailored treatment strategies. BACKGROUND: In ANCA-associated vasculitis, there is a lack of biomarkers for predicting renal relapse. Urinary T cells have been shown to differentiate active GN from remission in ANCA-associated vasculitis, but their predictive value for renal flares remains unknown. METHODS: The PRE-FLARED study was a prospective multicenter biomarker study including 102 individuals with ANCA-associated vasculitis in remission aimed to predict renal relapse by quantifying urinary CD4 + T-cell subsets using flow cytometry at baseline and monitoring clinical outcomes over a 6-month follow-up. RESULTS: Among the participants, ten experienced renal relapses, two had non-renal flares, and 90 remained in stable remission. The median baseline urinary CD4 + T-cell count was significantly higher in patients who relapsed compared with those in remission. Receiver operating characteristic curve analysis of urinary CD4 + T-cell counts showed an area under the curve value of 0.88 for predicting renal flares, outperforming ANCA titers, hematuria, and proteinuria. Using a cutoff of 490 CD4 + T cells per 100 ml urine, the sensitivity and specificity in identifying patients with future renal flares were 60% and 97.8%, respectively. In a post hoc analysis, combining urinary CD4 + T-cell counts with proteinase-3 ANCA levels suggested improved predictive performance in the PR3 + subgroup. In addition, the number of urinary CD4 + T cells showed a limited correlation with a decline in GFR and an increase in proteinuria over the follow-up period. CONCLUSIONS: This study concluded that urinary CD4 + T-cell counts could identify patients with ANCA-associated vasculitis at a substantial risk of renal relapse within 6 months. Combining these counts with ANCA levels further improved the prediction of relapse. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Urinary T Lymphocytes Predict Renal Flares in Patients With Inactive ANCA-associated Glomerulonephritis (PRE-FLARED), NCT04428398 .

Thromboxane biosynthesis and future events in diabetes: the ASCEND trial.

BACKGROUND AND AIMS: Thromboxane (TX) A2, released by activated platelets, plays an important role in atherothrombosis. Urinary 11-dehydro-TXB2 (U-TXM), a stable metabolite reflecting the whole-body TXA2 biosynthesis, is reduced by ∼70% by daily low-dose aspirin. The U-TXM represents a non-invasive biomarker of in vivo platelet activation and is enhanced in patients with diabetes. This study assessed whether U-TXM is associated with the risk of future serious vascular events or revascularizations (SVE-R), major bleeding, or cancer in patients with diabetes. METHODS: The U-TXM was measured pre-randomization to aspirin or placebo in 5948 people with type 1 or 2 diabetes and no cardiovascular disease, in the ASCEND trial. Associations between log U-TXM and SVE-R (n = 618), major bleed (n = 206), and cancer (n = 700) during 6.6 years of follow-up were investigated by Cox regression; comparisons of these associations with the effects of randomization to aspirin were made. RESULTS: Higher U-TXM was associated with older age, female sex, current smoking, type 2 diabetes, higher body size, urinary albumin/creatinine ratio of ≥3 mg/mmol, and higher estimated glomerular filtration rate. After adjustment for these, U-TXM was marginally statistically significantly associated with SVE-R and major bleed but not cancer [hazard ratios per 1 SD higher log U-TXM (95% confidence interval): 1.09 (1.00-1.18), 1.16 (1.01-1.34), and 1.06 (0.98-1.14)]. The hazard ratio was similar to that implied by the clinical effects of randomization to aspirin for SVE-R but not for major bleed. CONCLUSIONS: The U-TXM was log-linearly independently associated with SVE-R in diabetes. This is consistent with the involvement of platelet TXA2 in diabetic atherothrombosis.

Ultrasensitive CCL2 Detection in Urine for Diabetic Nephropathy Diagnosis Using a WS2-Based Plasmonic Biosensor.

The accurate diagnosis of diabetic nephropathy relies on achieving ultrasensitive biosensing for biomarker detection. However, existing biosensors face challenges such as poor sensitivity, complexity, time-consuming procedures, and high assay costs. To address these limitations, we report a WS2-based plasmonic biosensor for the ultrasensitive detection of biomarker candidates in clinical human urine samples associated with diabetic nephropathy. Leveraging plasmonic-based electrochemical impedance microscopy (P-EIM) imaging, we observed a remarkable charge sensitivity in monolayer WS2 single crystals. Our biosensor exhibits an exceptionally low detection limit (0.201 ag/mL) and remarkable selectivity in detecting CC chemokine ligand 2 (CCL2) protein biomarkers, outperforming conventional techniques such as ELISA. This work represents a breakthrough in traditional protein sensors, providing a direction and materials foundation for developing ultrasensitive sensors tailored to clinical applications for biomarker sensing.

Proteomics of urinary small extracellular vesicles in early diagnosis of kidney diseases in children-expectations and limitations.

The primary function of the kidneys is to maintain systemic homeostasis (disruption of renal structure and function results in multilevel impairment of body function). Kidney diseases are characterized by a chronic, progressive course and may result in the development of chronic kidney disease (CKD). Evaluation of the composition of the proteome of urinary small extracellular vesicles (sEVs) as a so-called liquid biopsy is a promising new research direction. Knowing the composition of sEV could allow localization of cellular changes in specific sections of the nephron or the interstitial tissue before fixed changes, detectable only at an advanced stage of the disease, occur. Research is currently underway on the role of sEVs in the diagnosis and monitoring of many disease entities. Reports in the literature on the subject include: diabetic nephropathy, focal glomerulosclerosis in the course of glomerulopathies, renal fibrosis of various etiologies. Studies on pediatric patients are still few, involving piloting if small groups of patients without validation studies. Here, we review the literature addressing the use of sEV for diagnosis of the most common urinary disorders in children. We evaluate the clinical utility and define limitations of markers present in sEV as potential liquid biopsy.

Application of Proteomics and Machine Learning Methods to Study the Pathogenesis of Diabetic Nephropathy and Screen Urinary Biomarkers.

Diabetic nephropathy (DN) has become the main cause of end-stage renal disease worldwide, causing significant health problems. Early diagnosis of the disease is quite inadequate. To screen urine biomarkers of DN and explore its potential mechanism, this study collected urine from 87 patients with type 2 diabetes mellitus (which will be classified into normal albuminuria, microalbuminuria, and macroalbuminuria groups) and 38 healthy subjects. Twelve individuals from each group were then randomly selected as the screening cohort for proteomics analysis and the rest as the validation cohort. The results showed that humoral immune response, complement activation, complement and coagulation cascades, renin-angiotensin system, and cell adhesion molecules were closely related to the progression of DN. Five overlapping proteins (KLK1, CSPG4, PLAU, SERPINA3, and ALB) were identified as potential biomarkers by machine learning methods. Among them, KLK1 and CSPG4 were positively correlated with the urinary albumin to creatinine ratio (UACR), and SERPINA3 was negatively correlated with the UACR, which were validated by enzyme-linked immunosorbent assay (ELISA). This study provides new insights into disease mechanisms and biomarkers for early diagnosis of DN.

Comprehensive Urinary Proteome Profiling Analysis Identifies Diagnosis and Relapse Surveillance Biomarkers for Bladder Cancer.

Bladder cancer (BCa) is the predominant malignancy of the urinary system. Herein, a comprehensive urine proteomic feature was initially established for the noninvasive diagnosis and recurrence monitoring of bladder cancer. 279 cases (63 primary BCa, 87 nontumor controls (NT), 73 relapsed BCa (BCR), and 56 nonrelapsed BCa (BCNR)) were collected to screen urinary protein biomarkers. 4761 and 3668 proteins were qualified and quantified by DDA and sequential window acquisition of all theoretical mass spectra (SWATH-MS) analysis in two discovery sets, respectively. Upregulated proteins were validated by multiple reaction monitoring (MRM) in two independent combined sets. Using the multi-support vector machine-recursive feature elimination (mSVM-RFE) algorithm, a model comprising 13 proteins exhibited good performance between BCa and NT with an AUC of 0.821 (95% CI: 0.675-0.967), 90.9% sensitivity (95% CI: 72.7-100%), and 73.3% specificity (95% CI: 53.3-93.3%) in the diagnosis test set. Meanwhile, an 11-marker classifier significantly distinguished BCR from BCNR with 75.0% sensitivity (95% CI: 50.0-100%), 81.8% specificity (95% CI: 54.5-100%), and an AUC of 0.784 (95% CI: 0.609-0.959) in the test cohort for relapse surveillance. Notably, six proteins (SPR, AK1, CD2AP, ADGRF1, GMPS, and C8A) of 24 markers were newly reported. This paper reveals novel urinary protein biomarkers for BCa and offers new theoretical insights into the pathogenesis of bladder cancer (data identifier PXD044896).

Comparative Analysis of Protein Quantification by the SomaScan Assay versus Orthogonal Methods in Urine from People with Diabetic Kidney Disease.

To our knowledge, calibration curves or other validations for thousands of SomaScan aptamers are not publicly available. Moreover, the abundance of urine proteins obtained from these assays is not routinely validated with orthogonal methods (OMs). We report an in-depth comparison of SomaScan readout for 23 proteins in urine samples from patients with diabetic kidney disease (n = 118) vs OMs, including liquid chromatography-targeted mass spectrometry (LC-MS), ELISA, and nephelometry. Pearson correlation between urine abundance of the 23 proteins from SomaScan 3.2 vs OMs ranged from -0.58 to 0.86, with a median (interquartile ratio, [IQR]) of 0.49 (0.18, 0.53). In multivariable linear regression, the SomaScan readout for 6 of the 23 examined proteins (26%) was most strongly associated with the OM-derived abundance of the same (target) protein. For 3 of 23 (13%), the SomaScan and OM-derived abundance of each protein were significantly associated, but the SomaScan readout was more strongly associated with OM-derived abundance of one or more "off-target" proteins. For the remaining 14 proteins (61%), the SomaScan readouts were not significantly associated with the OM-derived abundance of the targeted proteins. In 6 of the latest group, the SomaScan readout was not associated with urine abundance of any of the 23 quantified proteins. To sum, over half of the SomaScan results could not be confirmed by independent orthogonal methods.

Comprehensive Prostate Fluid-Based Spectral Libraries for Enhanced Protein Detection in Urine.

Biofluids contain molecules in circulation and from nearby organs that can be indicative of disease states. Characterizing the proteome of biofluids with DIA-MS is an emerging area of interest for biomarker discovery; yet, there is limited consensus on DIA-MS data analysis approaches for analyzing large numbers of biofluids. To evaluate various DIA-MS workflows, we collected urine from a clinically heterogeneous cohort of prostate cancer patients and acquired data in DDA and DIA scan modes. We then searched the DIA data against urine spectral libraries generated using common library generation approaches or a library-free method. We show that DIA-MS doubles the sample throughput compared to standard DDA-MS with minimal losses to peptide detection. We further demonstrate that using a sample-specific spectral library generated from individual urines maximizes peptide detection compared to a library-free approach, a pan-human library, or libraries generated from pooled, fractionated urines. Adding urine subproteomes, such as the urinary extracellular vesicular proteome, to the urine spectral library further improves the detection of prostate proteins in unfractionated urine. Altogether, we present an optimized DIA-MS workflow and provide several high-quality, comprehensive prostate cancer urine spectral libraries that can streamline future biomarker discovery studies of prostate cancer using DIA-MS.

Non-adherence to cardiometabolic medication as assessed by LC-MS/MS in urine and its association with kidney and cardiovascular outcomes in type 2 diabetes mellitus.

AIMS/HYPOTHESIS: Non-adherence to medication is a frequent barrier in the treatment of patients with type 2 diabetes mellitus, potentially limiting the effectiveness of evidence-based treatments. Previous studies have mostly relied on indirect adherence measures to analyse outcomes based on adherence. The aim of this study was to use LC-MS/MS in urine-a non-invasive, direct and objective measure-to assess non-adherence to cardiometabolic drugs and analyse its association with kidney and cardiovascular outcomes. METHODS: This cohort study includes 1125 participants from the PROVALID study, which follows patients with type 2 diabetes mellitus at the primary care level. Baseline urine samples were tested for 79 cardiometabolic drugs and metabolites thereof via LC-MS/MS. An individual was classified as totally adherent if markers for all drugs were detected, partially non-adherent when at least one marker for one drug was detected, and totally non-adherent if no markers for any drugs were detected. Non-adherence was then analysed in the context of cardiovascular (composite of myocardial infarction, stroke and cardiovascular death) and kidney (composite of sustained 40% decline in eGFR, sustained progression of albuminuria, kidney replacement therapy and death from kidney failure) outcomes. RESULTS: Of the participants, 56.3% were totally adherent, 42.0% were partially non-adherent, and 1.7% were totally non-adherent to screened cardiometabolic drugs. Adherence was highest to antiplatelet and glucose-lowering agents and lowest to lipid-lowering agents. Over a median (IQR) follow-up time of 5.10 (4.12-6.12) years, worse cardiovascular outcomes were observed with non-adherence to antiplatelet drugs (HR 10.13 [95% CI 3.06, 33.56]) and worse kidney outcomes were observed with non-adherence to antihypertensive drugs (HR 1.98 [95% CI 1.37, 2.86]). CONCLUSIONS/INTERPRETATION: This analysis shows that non-adherence to cardiometabolic drug regimens is common in type 2 diabetes mellitus and negatively affects kidney and cardiovascular outcomes.

Competitive fitness of asymptomatic bacteriuria E. coli strain 83972 against uropathogens in human urine.

Urinary tract infection (UTI) is one of the most common bacterial infections worldwide. The main causative agent of UTI is uropathogenic Escherichia coli (UPEC). There is an immediate need for novel prophylactic and treatment strategies against UTI because of the increasing incidence of antimicrobial resistance among uropathogens. ABU 83972, an asymptomatic bacteriuria-causing E. coli strain, prevents UTI by suppressing the colonization of UPEC. However, the nature of competition and growth repression of UPEC by ABU 83972 is unclear and is the subject of our investigation. Here, we characterized the growth kinetics of ABU 83972 and uropathogens in human urine and laboratory media. Next, we performed a series of competitive co-culture experiments where ABU 83972 and uropathogens were inoculated at a 1:1 ratio in human urine and in various media, and their relative abundance was determined. In human urine, ABU 83972 outcompeted UPEC and additional uropathogens, reaching up to 90% of the total population after 24 hours of incubation. In contrast, UPEC outcompeted ABU 83972 in LB and M9 minimal media and exhibited superior colonization than ABU 83972 in the mouse urinary bladder. Since engineered living materials (ELMs) can be used to retain an organism of interest in a particular location, we developed ABU 83972-containing ELMs that effectively outcompeted UPEC in human urine. In summary, our work establishes that ABU 83972 outcompetes UPEC in a milieu- and cell-density-dependent manner, highlighting the importance of the metabolites and nutrients found in the human urine as determinants of the competitive fitness of ABU 83972.

Incident Proteinuria by HIV Serostatus Among Men With Pre--Diabetes Mellitus: The Multicenter AIDS Cohort Study.

BACKGROUND: Pre-diabetes mellitus (DM) is associated with proteinuria, a risk factor for chronic kidney disease. While people with human immunodeficiency virus (HIV; PWH) have a higher risk of proteinuria than people without HIV (PWOH), it is unknown whether incident proteinuria differs by HIV serostatus among prediabetic persons. METHODS: The urine protein-to-creatinine ratio was measured at semiannual visits among men in the Multicenter AIDS Cohort Study since April 2006. Men with pre-DM on or after April 2006 and no prevalent proteinuria or use of antidiabetic medications were included. Pre-DM was defined as a fasting glucose level of 100-125 mg/dL confirmed within a year by a repeated fasting glucose or hemoglobin A1c measurement of 5.7%-6.4%. Incident proteinuria was defined as a urine protein-to-creatinine ratio (UPCR) >200 mg/g, confirmed within a year. We used Poisson regression models to determine whether incident proteinuria in participants with pre-DM differed by HIV serostatus and, among PWH, whether HIV-specific factors were related to incident proteinuria. RESULTS: Between 2006 and 2019, among 1276 men with pre-DM, proteinuria developed in 128 of 613 PWH (21%) and 50 of 663 PWOH (8%) over a median 10-year follow-up. After multivariable adjustment, the incidence of proteinuria in PWH with pre-DM was 3.3 times (95% confidence interval, 2.3-4.8 times) greater than in PWOH (P < .01). Among PWH, current CD4 cell count <50/µL (P < .01) and current use of protease inhibitors (P = .03) were associated with incident proteinuria, while lamivudine and integrase inhibitor use were associated with a lower risk. CONCLUSIONS: Among men with pre-DM, the risk of incident proteinuria was 3 times higher in PWH. Strategies to preserve renal function are needed in this population.

Natural Protein Photon Upconversion Supramolecular Assemblies for Background-Free Biosensing.

The diagnosis of disease biomarkers is crucial for the identification, monitoring, and prognostic assessment of malignant disease. However, biological samples with autofluorescence, complex components, and heterogeneity pose major challenges to reliable biosensing. Here, we report the self-assembly of natural proteins and the triplet-triplet annihilation upconversion (TTA-UC) pair to form upconverted protein clusters (∼8.2 ± 1.1 nm), which were further assembled into photon upconversion supramolecular assemblies (PUSA). This PUSA exhibited unique features, including a small size (∼44.1 ± 4.1 nm), oxygen tolerance, superior biocompatibility, and easy storage via lyophilization, all of which are long sought after for photon upconversion materials. Further, we have revealed that the steric hindrance of the annihilator suppresses the stacking of the annihilator in PUSA, which is vital for maintaining the water dispersibility and enhancing the upconversion performance of PUSA. In conjunction with sarcosine oxidase, this near infrared (NIR)-excitable PUSA nanoprobe could perform background-free biosensing of urinary sarcosine, which is a common biomarker for prostatic carcinoma (PCa). More importantly, this nanoprobe not only allows for qualitative identification of urinary samples from PCa patients by the unaided eye under NIR-light-emitting diode (LED) illumination but also quantifies the concentration of urinary sarcosine. These remarkable findings have propelled photon upconversion materials to a new evolutionary stage and expedited the progress of upconversion biosensing in clinical diagnostics.

Renal calcium and magnesium handling during pregnancy: modeling and analysis.

Pregnancy is associated with elevated demand of most nutrients, with many trace elements and minerals critical for the development of fetus. In particular, calcium (Ca2+) and magnesium (Mg2+) are essential for cellular function, and their deficiency can lead to impaired fetal growth. A key contributor to the homeostasis of these ions is the kidney, which in a pregnant rat undergoes major changes in morphology, hemodynamics, and molecular structure. The goal of this study is to unravel the functional implications of these pregnancy-induced changes in renal handling of Ca2+ and Mg2+, two cations that are essential in a healthy pregnancy. To achieve that goal, we developed computational models of electrolyte and water transport along the nephrons of a rat in mid and late pregnancy. Model simulations reveal a substantial increase in the reabsorption of Mg2+ along the proximal tubules and thick ascending limbs. In contrast, the reabsorption of Ca2+ is increased in the proximal tubules but decreased in the thick ascending limbs, due to the lower transepithelial concentration gradient of Ca2+ along the latter. Despite the enhanced transport capacity, the marked increase in glomerular filtration rate results in elevated urinary excretions of Ca2+ and Mg2+ in pregnancy. Furthermore, we conducted simulations of hypocalcemia and hypomagnesemia. We found that hypocalcemia lowers Ca2+ excretion substantially more than Mg2+ excretion, with this effect being more pronounced in virgin rats than in pregnant ones. Conversely, hypomagnesemia reduces the excretion of Mg2+ and Ca2+ to more similar degrees. These differences can be explained by the greater sensitivity of the calcium-sensing receptor (CaSR) to Ca2+ compared with Mg2+.NEW & NOTEWORTHY A growing fetus' demands of minerals, notably calcium and magnesium, necessitate adaptations in pregnancy. In particular, the kidney undergoes major changes in morphology, hemodynamics, and molecular structure. This computational modeling study provides insights into how these pregnancy-induced renal adaptation impact calcium and magnesium transport along different nephron segments. Model simulations indicate that, despite the enhanced transport capacity, the marked increase in glomerular filtration rate results in elevated urinary excretions of calcium and magnesium in pregnancy.

Altered kidney distribution and loss of ACE2 into the urine in acute kidney injury.

There are diverse pathophysiological mechanisms involved in acute kidney injury (AKI). Among them, overactivity of the renin-angiotensin system (RAS) has been described. Angiotensin-converting enzyme 2 (ACE2) is a tissue RAS enzyme expressed in the apical border of proximal tubules. Given the important role of ACE2 in the metabolism of angiotensin II, this study aimed to characterize kidney and urinary ACE2 in a mouse model of AKI. Ischemia-reperfusion injury (IRI) was induced in C57BL/6 mice by clamping of the left renal artery followed by removal of the right kidney. In kidneys harvested 48 h after IRI, immunostaining revealed a striking maldistribution of ACE2 including spillage into the tubular lumen and the presence of ACE2-positive luminal casts in the medulla. In cortical membranes, ACE2 protein and enzymatic activity were both markedly reduced (37 ± 4 vs. 100 ± 6 ACE2/ß-actin, P = 0.0004, and 96 ± 14 vs. 152 ± 6 RFU/µg protein/h, P = 0.006). In urine, full-length membrane-bound ACE2 protein (100 kDa) was markedly increased (1,120 ± 405 vs. 100 ± 46 ACE2/µg creatinine, P = 0.04), and casts stained for ACE2 were recovered in the urine sediment. In conclusion, in AKI caused by IRI, there is a marked loss of ACE2 from the apical tubular border with deposition of ACE2-positive material in the medulla and increased urinary excretion of full-length membrane-bound ACE2 protein. The deficiency of tubular ACE2 in AKI suggests that provision of this enzyme could have therapeutic applications and that its excretion in the urine may also serve as a diagnostic marker of severe proximal tubular injury.NEW & NOTEWORTHY This study provides novel insights into the distribution of kidney ACE2 in a model of AKI by IRI showing a striking detachment of apical ACE2 from proximal tubules and its loss in urine and urine sediment. The observed deficiency of kidney ACE2 protein and enzymatic activity in severe AKI suggests that administration of forms of this enzyme may mitigate AKI and that urinary ACE2 may serve as a potential biomarker for tubular injury.

Sex-disaggregated analysis of acute kidney injury in hospitalized children with sickle cell anemia in Uganda.

A growing body of research is categorizing sex differences in both sickle cell anemia (SCA) and acute kidney injury (AKI); however, most of this work is being conducted in high-resource settings. Here, we evaluated risk factors and clinical parameters associated with AKI and AKI severity, stratified by sex, in a cohort of children hospitalized with SCA and vaso-occlusive pain crisis (VOC). The purpose of this study was to explore sex disparities in a high-risk, vulnerable population. This study was a secondary analysis of data collected from a cohort of Ugandan children between 2 and 18 yr of age prospectively enrolled. A total of 185 children were enrolled in the primary study; 41.6% were female and 58.4% were male, with a median age of 8.9 yr. Incident or worsening AKI (P = 0.026) occurred more frequently in female compared with male children, despite no differences in AKI on admission. Female children also had altered markers of renal function including higher creatinine levels at admission (P = 0.03), higher peak creatinine (P = 0.006), and higher urine neutrophil gelatinase-associated lipocalin (NGAL) at admission (P = 0.003) compared with male children. Female children had elevated total (P = 0.045) and conjugated bilirubin at admission (P = 0.02) compared with male children and higher rates of hematuria at admission (P = 0.004). Here, we report sex differences in AKI in children with SCA and VOC, including increased incidence and worsening of AKI in female pediatric patients, in association with an increase in biological indicators of poor renal function including creatinine, estimated glomerular filtration rate, and NGAL.NEW & NOTEWORTHY In this study, we report an increased risk of developing acute kidney injury (AKI) during hospitalization, worsening AKI, and death among females with sickle cell anemia (SCA) hospitalized with an acute pain crisis compared with males. The sex differences in AKI were not explained by socioeconomic differences, severity of pain, or disease severity among females compared with males. Together, these data suggest that female children with SCA may be at increased risk of AKI.

Effect of the DASH diet on the sodium-chloride cotransporter and aquaporin-2 in urinary extracellular vesicles.

The dietary approach to stop hypertension (DASH) diet combines the antihypertensive effect of a low sodium and high potassium diet. In particular, the potassium component of the diet acts as a switch in the distal convoluted tubule to reduce sodium reabsorption, similar to a diuretic but without the side effects. Previous trials to understand the mechanism of the DASH diet were based on animal models and did not characterize changes in human ion channel protein abundance. More recently, protein cargo of urinary extracellular vesicles (uEVs) has been shown to mirror tissue content and physiological changes within the kidney. We designed an inpatient open label nutritional study transitioning hypertensive volunteers from an American style diet to DASH diet to examine physiological changes in adults with stage 1 hypertension otherwise untreated (Sacks FM, Svetkey LP, Vollmer WM, Appel LJ, Bray GA, Harsha D, Obarzanek E, Conlin PR, Miller ER 3rd, Simons-Morton DG, Karanja N, Lin PH; DASH-Sodium Collaborative Research Group. N Engl J Med 344: 3-10, 2001). Urine samples from this study were used for proteomic characterization of a large range of pure uEVs (small to large) to reveal kidney epithelium changes in response to the DASH diet. These samples were collected from nine volunteers at three time points, and mass spectrometry identified 1,800 proteins from all 27 samples. We demonstrated an increase in total SLC12A3 [sodium-chloride cotransporter (NCC)] abundance and a decrease in aquaporin-2 (AQP2) in uEVs with this mass spectrometry analysis, immunoblotting revealed a significant increase in the proportion of activated (phosphorylated) NCC to total NCC and a decrease in AQP2 from day 5 to day 11. This data demonstrates that the human kidney's response to nutritional interventions may be captured noninvasively by uEV protein abundance changes. Future studies need to confirm these findings in a larger cohort and focus on which factor drove the changes in NCC and AQP2, to which degree NCC and AQP2 contributed to the antihypertensive effect and address if some uEVs function also as a waste pathway for functionally inactive proteins rather than mirroring protein changes.NEW & NOTEWORTHY Numerous studies link DASH diet to lower blood pressure, but its mechanism is unclear. Urinary extracellular vesicles (uEVs) offer noninvasive insights, potentially replacing tissue sampling. Transitioning to DASH diet alters kidney transporters in our stage 1 hypertension cohort: AQP2 decreases, NCC increases in uEVs. This aligns with increased urine volume, reduced sodium reabsorption, and blood pressure decline. Our data highlight uEV protein changes as diet markers, suggesting some uEVs may function as waste pathways. We analyzed larger EVs alongside small EVs, and NCC in immunoblots across its molecular weight range.

KS-WNK1 is required for the renal response to extreme changes in potassium intake.

Kidney-specific with-no-lysine kinase 1 (KS-WNK1) is an isoform of WNK1 kinase that is predominantly found in the distal convoluted tubule of the kidney. The precise physiological function of KS-WNK1 remains unclear. Some studies have suggested that it could play a role in regulating potassium renal excretion by modulating the activity of the Na+-Cl- cotransporter (NCC). However, changes in the potassium diet from normal to high failed to reveal a role for KS-WNK1, but under a normal-potassium diet, the expression of KS-WNK1 is negligible. It is only detectable when mice are exposed to a low-potassium diet. In this study, we investigated the role of KS-WNK1 in regulating potassium excretion under extreme changes in potassium intake. After following a zero-potassium diet (0KD) for 10 days, KS-WNK1-/- mice had lower plasma levels of K+ and Cl- while exhibiting higher urinary excretion of Na+, Cl-, and K+ compared with KS-WNK1+/+ mice. After 10 days of 0KD or normal-potassium diet (NKD), all mice were challenged with a high-potassium diet (HKD). Plasma K+ levels markedly increased after the HKD challenge only in mice previously fed with 0KD, regardless of genotype. KSWNK1+/+ mice adapt better to HKD challenge than KS-WNK1-/- mice after a potassium-retaining state. The difference in the phosphorylated NCC-to-NCC ratio between KS-WNK1+/+ and KS-WNK1-/- mice after 0KD and HKD indicates a role for KS-WNK1 in both NCC phosphorylation and dephosphorylation. These observations show that KS-WNK1 helps the distal convoluted tubule to respond to extreme changes in potassium intake, such as those occurring in wildlife.NEW & NOTEWORTHY The findings of this study demonstrate that kidney-specific with-no-lysine kinase 1 plays a role in regulating urinary electrolyte excretion during extreme changes in potassium intake, such as those occurring in wildlife. .