Contralateral renal change in a unilateral ureteral obstruction rat model using intravoxel incoherent motion diffusion-weighted imaging.

OBJECTIVES: Most functional magnetic resonance research has primarily examined alterations in the affected kidney, often neglecting the contralateral kidney. Our study aims to investigate whether imaging parameters accurately depict changes in both the renal cortex and medulla in a unilateral ureteral obstruction rat model, thereby showcasing the utility of intravoxel incoherent motion (IVIM) in evaluating contralateral renal changes. METHODS: Six rats underwent MR scans and were subsequently sacrificed for baseline histological examination. Following the induction of left ureteral obstruction, 48 rats were scanned, and the histopathological examinations were conducted on days 3, 7, 10, 14, 21, 28, 35, and 42. The apparent diffusion coefficient (ADC), pure molecular diffusion (D), pseudodiffusion (D*), and perfusion fraction (f) values were measured using IVIM. RESULTS: On the 10th day of obstruction, both cortical and medullary ADC values differed significantly between the UUO10 group and the sham group (p < 0.01). The cortical D values showed statistically significant differences between UUO3 group and sham group (p < 0.01) but not among UUO groups at other time point. Additionally, the cortical and medullary f values were statistically significant between the UUO21 group and the sham group (p < 0.01). Especially, the cortical f values exhibited significant differences between the UUO21 group and the UUO groups with shorter obstruction time (at time point of 3, 7, 10, 14 day) (p < 0.01). CONCLUSIONS: Significant hemodynamic alterations were observed in the contralateral kidney following renal obstruction. IVIM accurately captures changes in the unobstructed kidney. Particularly, the cortical f value exhibits the highest potential for assessing contralateral renal modifications.

Variation coefficient of stone density and renal cortical thickness: the parameters evaluating non-contrast computed tomography imaging for predict extracorporeal shock wave lithotripsy success.

The stone density (SD) is not the same in all parts of the stone due to the heterogeneous nature of the stone and the shock wave (SW) passes through tissues of many different densities until it reaches the stone. These factors affect the success of Extracorporeal Shock Wave Lithotripsy (ESWL). We aimed to evaluate the effect of the Variation Coefficient of Stone Density (VCSD) and Renal Cortical Tickness (RCT) on the success of ESWL. Between 2020 and 2023, 510 patients who underwent ESWL were divided into 2 groups treatment success (n:304) and treatment failure (n:206). Non-Contrast Computed Tomography (NCCT) imaging values of hydronephrosis degree of the kidney, stone location, stone volume (SV), stone-skin distance (SSD), SD, Standard deviation of Stone Density (SDSD), VCSD, RCT, Soft-Tissue Thickness (STT), Muscle Thickness (MT) were analyzed. VCSD value was obtained by dividing SDSD by SD. Along the SW, tissues were divided into three components: kidney (renal cortex), muscle and other soft tissues. RCT, MT and SSD were measured at three different angles (0°, 45°, and 90°) and these 3 lengths were averaged. In univariate analysis, Body Mass Index (BMI), SV, SD, VCSD, SSD, RCT and STT were demonstrated to affect ESWL success. In multivariate analysis, low BMI, SV, SD, RCT and large VCSD were significant independent predictors of ESWL success. Among these parameters, VCSD had the highest prediction accuracy, followed by SD, SV, RCT and BMI, respectively. This study demonstrated that VCSD value and RCT are predictive parameters in determining the treatment of patients with urinary calculi and selecting suitable ESWL candidates.

Effects of furosemide, acetazolamide and amiloride on renal cortical and medullary tissue oxygenation in non-anaesthetised healthy sheep.

It has been proposed that diuretics can improve renal tissue oxygenation through inhibition of tubular sodium reabsorption and reduced metabolic demand. However, the impact of clinically used diuretic drugs on the renal cortical and medullary microcirculation is unclear. Therefore, we examined the effects of three commonly used diuretics, at clinically relevant doses, on renal cortical and medullary perfusion and oxygenation in non-anaesthetised healthy sheep. Merino ewes received acetazolamide (250 mg; n = 9), furosemide (20 mg; n = 10) or amiloride (10 mg; n = 7) intravenously. Systemic and renal haemodynamics, renal cortical and medullary tissue perfusion and P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ , and renal function were then monitored for up to 8 h post-treatment. The peak diuretic response occurred 2 h (99.4 ± 14.8 mL/h) after acetazolamide, at which stage cortical and medullary tissue perfusion and P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ were not significantly different from their baseline levels. The peak diuretic response to furosemide occurred at 1 h (196.5 ± 12.3 mL/h) post-treatment but there were no significant changes in cortical and medullary tissue oxygenation during this period. However, cortical tissue P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ fell from 40.1 ± 3.8 mmHg at baseline to 17.2 ± 4.4 mmHg at 3 h and to 20.5 ± 5.3 mmHg at 6 h after furosemide administration. Amiloride did not produce a diuretic response and was not associated with significant changes in cortical or medullary tissue oxygenation. In conclusion, clinically relevant doses of diuretic agents did not improve regional renal tissue oxygenation in healthy animals during the 8 h experimentation period. On the contrary, rebound renal cortical hypoxia may develop after dissipation of furosemide-induced diuresis.

Metabolic Fluxes in the Renal Cortex Are Dysregulated In Vivo in Response to High-Fat Diet.

Diabetes and obesity are risk factors for kidney disease. Whereas renal glucose production increases in diabetes, recent data suggest that gluconeogenic and oxidative capacity decline in kidney disease. Thus, metabolic dysregulation caused by diet-induced insulin resistance may sensitize the kidney for a loss in function. Here, we examined how diet-induced insulin resistance disrupts mitochondrial metabolic fluxes in the renal cortex in vivo. C57BL/6J mice were rendered insulin resistant through high-fat (HF) feeding; anaplerotic, cataplerotic, and oxidative metabolic fluxes in the cortex were quantified through 13C-isotope tracing during a hyperinsulinemic-euglycemic clamp. As expected, HF-fed mice exhibited increased body weight, gluconeogenesis, and systemic insulin resistance compared with chow-fed mice. Relative to the citric acid cycle, HF feeding increased metabolic flux through pyruvate carboxylation (anaplerosis) and phosphoenolpyruvate carboxykinase (cataplerosis) and decreased flux through the pyruvate dehydrogenase complex in the cortex. Furthermore, the relative flux from nonpyruvate sources of acetyl-CoA profoundly increased in the cortex of HF-fed mice, correlating with a marker of oxidative stress. The data demonstrate that HF feeding spares pyruvate from dehydrogenation at the expense of increasing cataplerosis, which may underpin renal gluconeogenesis during insulin resistance; the results also support the hypothesis that dysregulated oxidative metabolism in the kidney contributes to metabolic disease.

Patterns of cortical oxygenation may predict the response to stenting in subjects with renal artery stenosis: A radiomics-based model.

BACKGROUND: Percutaneous-transluminal renal angioplasty (PTRA) and stenting aim to halt the progression of kidney disease in patients with renal artery stenosis (RAS), but its outcome is often suboptimal. We hypothesized that a model incorporating markers of renal function and oxygenation extracted using radiomics analysis of blood oxygenation-level dependent (BOLD)-MRI images may predict renal response to PTRA in swine RAS. MATERIALS AND METHODS: Twenty domestic pigs with RAS were scanned with CT and BOLD MRI before and 4 weeks after PTRA. Stenotic (STK) and contralateral (CLK) kidney volume, blood flow (RBF), and glomerular filtration rate (GFR) were determined, and BOLD-MRI R2 * maps were generated before and after administration of furosemide, a tubular reabsorption inhibitor. Radiomics features were extracted from pre-PTRA BOLD maps and Robust features were determined by Intraclass correlation coefficients (ICC). Prognostic models were developed to predict post-PTRA renal function based on the baseline functional and BOLD-radiomics features, using Lasso-regression for training, and testing with resampling. RESULTS: Twenty-six radiomics features passed the robustness test. STK oxygenation distribution pattern did not respond to furosemide, whereas in the CLK radiomics features sensitive to oxygenation heterogeneity declined. Radiomics-based model predictions of post-PTRA GFR (r = 0.58, p = 0.007) and RBF (r = 0.68; p = 0.001) correlated with actual measurements with sensitivity and specificity of 92% and 67%, respectively. Models were unsuccessful in predicting post-PTRA systemic measures of renal function. CONCLUSIONS: Several radiomics features are sensitive to cortical oxygenation patterns and permit estimation of post-PTRA renal function, thereby distinguishing subjects likely to respond to PTRA and stenting.

An Objective Computer-Assisted Measurement of Sonographic Renal Cortical Echogenicity: The Splenorenal Index.

ABSTRACT: Renal cortical echogenicity represents a marker of renal function. However, evaluation of the renal echotexture is subjective and thus disposed to error and interrater variability. Computer-aided image analysis may be used to objectively assess renal cortical echogenicity by comparing the echogenicity of the left kidney to that of the spleen; the resultant ratio is referred to as the splenorenal index (SRI). We performed a retrospective review of all adult patients who received a renal ultrasound over a 45-day period at our institution. Demographic data and kidney function laboratory values were documented for each patient. Regions of interest (ROIs) were selected in the left renal cortex and spleen using ImageJ software. The SRI was calculated as a ratio of the mean pixel brightness of the left kidney cortex ROI to the mean pixel brightness of the spleen ROI. The SRI was then correlated with serum creatinine, blood urea nitrogen, and estimated glomerular filtration rate. We found that among the 94 patients included in the study, the SRI had a significant positive correlation with serum creatinine ( r = 0.43, P < 0.001) and serum blood urea nitrogen ( r = 0.45, P < 0.001) and negative correlation with estimated glomerular filtration rate ( r = -0.47, P < 0.001). Our data indicate that SRI may serve as a valuable tool for sonographic evaluation of renal parenchymal disease.

A murine monoclonal antibody against H5N1 avian influenza virus cross-reacts with human kidney cortex cells.

To investigate the biological characteristics of monoclonal antibodies (mAbs) against avian influenza virus (AIV) and the possible mechanism of AIV-related kidney injury. BALB/c mice were immunized with inactivated H5N1 AIV to prepare monoclonal antibody H5-32, and its subtype, titer and cross-reactivity with other influenza viruses were identified. The reactivity of monoclonal antibody with normal human tissue was analyzed by immunohistochemistry. Immunofluorescence and confocal laser scanning technique were used to detect the binding sites between mAb and human renal cortical cells, and Western blotting was used to detect the size of binding fragments. Immunohistochemical analysis confirmed that monoclonal antibody H5-32 cross-reacted with normal human kidney tissue. In human kidney, mAb H5-32 was localized in the cytoplasm of human renal tubular epithelial cells, and its binding fragment size was about 43 kDa. H5N1 AIV appears to bind to human renal tubular epithelial cells, which may be one of the mechanisms of kidney injury caused by AIV infection.

3D autofluorescence imaging of hydronephrosis and renal anatomical structure using cryo-micro-optical sectioning tomography.

Rationale: Mesoscopic visualization of the main anatomical structures of the whole kidney in vivo plays an important role in the pathological diagnosis and exploration of the etiology of hydronephrosis. However, traditional imaging methods cannot achieve whole-kidney imaging with micron resolution under conditions representing in vivo perfusion. Methods: We used in vivo cryofixation (IVCF) to fix acute obstructive hydronephrosis (unilateral ureteral obstruction, UUO), chronic spontaneous hydronephrosis (db/db mice), and their control mouse kidneys for cryo-micro-optical sectioning tomography (cryo-MOST) autofluorescence imaging. We quantitatively assessed the kidney-wide pathological changes in the main anatomical structures, including hydronephrosis, renal subregions, arteries, veins, glomeruli, renal tubules, and peritubular functional capillaries. Results: By comparison with microcomputed tomography imaging, we confirmed that IVCF can maintain the status of the kidney in vivo. Cryo-MOST autofluorescence imaging can display the main renal anatomical structures with a cellular resolution without contrast agents. The hydronephrosis volume reached 26.11 ± 6.00 mm3 and 13.01 ± 3.74 mm3 in 3 days after UUO and in 15-week-old db/db mouse kidneys, respectively. The volume of the cortex and inner stripe of the outer medulla (ISOM) increased while that of the inner medulla (IM) decreased in UUO mouse kidneys. Db/db mice also showed an increase in the volume of the cortex and ISOM volume but no atrophy in the IM. The diameter of the proximal convoluted tubule and proximal straight tubule increased in both UUO and db/db mouse kidneys, indicating that proximal tubules were damaged. However, some renal tubules showed abnormal central bulge highlighting in the UUO mice, but the morphology of renal tubules was normal in the db/db mice, suggesting differences in the pathology and severity of hydronephrosis between the two models. UUO mouse kidneys also showed vascular damage, including segmental artery and vein atrophy and arcuate vein dilation, and the density of peritubular functional capillaries in the cortex and IM was reduced by 37.2% and 49.5%, respectively, suggesting renal hypoxia. In contrast, db/db mouse kidneys showed a normal vascular morphology and peritubular functional capillary density. Finally, we found that the db/db mice displayed vesicoureteral reflux and bladder overactivity, which may be the cause of hydronephrosis formation. Conclusions: We observed and compared main renal structural changes in hydronephrosis under conditions representing in vivo perfusion in UUO, db/db, and control mice through cryo-MOST autofluorescence imaging. The results indicate that cryo-MOST with IVCF can serve as a simple and powerful tool to quantitatively evaluate the in vivo pathological changes in three dimensions, especially the distribution of body fluids in the whole kidney. This method is potentially applicable to the three-dimensional visualization of other tissues, organs, and even the whole body, which may provide new insights into pathological changes in diseases.

Computational Modeling of Substrate-Dependent Mitochondrial Respiration and Bioenergetics in the Heart and Kidney Cortex and Outer Medulla.

Integrated computational modeling provides a mechanistic and quantitative framework to characterize alterations in mitochondrial respiration and bioenergetics in response to different metabolic substrates in-silico. These alterations play critical roles in the pathogenesis of diseases affecting metabolically active organs such as heart and kidney. Therefore, the present study aimed to develop and validate thermodynamically constrained integrated computational models of mitochondrial respiration and bioenergetics in the heart and kidney cortex and outer medulla (OM). The models incorporated the kinetics of major biochemical reactions and transport processes as well as regulatory mechanisms in the mitochondria of these tissues. Intrinsic model parameters such as Michaelis-Menten constants were fixed at previously estimated values, while extrinsic model parameters such as maximal reaction and transport velocities were estimated separately for each tissue. This was achieved by fitting the model solutions to our recently published respirometry data measured in isolated rat heart and kidney cortex and OM mitochondria utilizing various NADH- and FADH2-linked metabolic substrates. The models were validated by predicting additional respirometry and bioenergetics data, which were not used for estimating the extrinsic model parameters. The models were able to predict tissue-specific and substrate-dependent mitochondrial emergent metabolic system properties such as redox states, enzyme and transporter fluxes, metabolite concentrations, membrane potential, and respiratory control index under diverse physiological and pathological conditions. The models were also able to quantitatively characterize differential regulations of NADH- and FADH2-linked metabolic pathways, which contribute differently toward regulations of oxidative phosphorylation and ATP synthesis in the heart and kidney cortex and OM mitochondria.

Effects of ROS pathway inhibitors and NADH and FADH2 linked substrates on mitochondrial bioenergetics and ROS emission in the heart and kidney cortex and outer medulla.

Mitochondria are major sources of reactive oxygen species (ROS), which play important roles in both physiological and pathological processes. However, the specific contributions of different ROS production and scavenging components in the mitochondria of metabolically active tissues such as heart and kidney cortex and outer medulla (OM) are not well understood. Therefore, the goal of this study was to determine contributions of different ROS production and scavenging components and provide detailed comparisons of mitochondrial respiration, bioenergetics, ROS emission between the heart and kidney cortex and OM using tissues obtained from the same Sprague-Dawley rat under identical conditions and perturbations. Specifically, data were obtained using both NADH-linked substrate pyruvate + malate and FADH2-linked substrate succinate followed by additions of inhibitors of different components of the electron transport chain (ETC) and oxidative phosphorylation (OxPhos) and other ROS production and scavenging systems. Currently, there is limited data available for the mitochondria of kidney cortex and OM, the two major energy-consuming tissues in the body only next to the heart, and scarce quantitative information on the interplay between mitochondrial ROS production and scavenging systems in the three tissues. The findings from this study demonstrate significant differences in mitochondrial respiratory and bioenergetic functions and ROS emission among the three tissues. The results quantify the rates of ROS production from different complexes of the ETC, identify the complexes responsible for variations in mitochondrial membrane depolarization and regulations of ROS production, and quantify the contributions of ROS scavenging enzymes towards overall mitochondrial ROS emission. These findings advance our fundamental knowledge of tissue-specific and substrate-dependent mitochondrial respiratory and bioenergetic functions and ROS emission. This is important given the critical role that excess ROS production, oxidative stress, and mitochondrial dysfunction in the heart and kidney cortex and OM play in the pathogenesis of cardiovascular and renal diseases, including salt-sensitive hypertension.

2D Shear wave elastography for evaluation of renal cortex and medulla: stiffness values in healthy children.

PURPOSE: To evaluate renal cortical and medullary stiffness using 2D Shear-wave elastography (SWE) in healthy children. METHODS: In this IRB approved prospective study, we measured the stiffness of cortex and medulla of children (4 months-17 years) at the upper pole, mid pole, and lower pole in bilateral kidneys. RESULTS: The median (IQR) values of renal cortex in <1 year age group was 8.7 (5.7-11.7) kPa for right and 8.7 (4.2-14.1) kPa for the left side. For 1-5 years age group, it was 7.3 (5.3-10) kPa for the right and 8.9 (6-12.3) kPa for the left side. For >5 years, it was 7.4 (5.3-11.2) kPa for the right and 9.6 (6.2-12.7) kPa for the left side. The median (IQR) values of renal medulla in <1 year age group was 7.1 (5.1-12.5) kPa for right and 6.8 (4-10.6) kPa for the left side. For 1-5 years age group, it was 7.2 (4.9-9.7) kPa for the right and 6.9 (5.6-9.9) kPa for the left side. For >5 years, it was 6.8 (5.1-9.6) kPa for the right and 7 (5-10.2) kPa for the left side. The differences in the elasticity values amongst these groups were statistically insignificant (p>0.05). There was a significant correlation between SWE values of cortex and medulla of right kidney (ρ=0.64) and of left kidney (ρ=0.61), respectively. CONCLUSION: SWE values of renal cortical and medullary stiffness in healthy children do not correlate with age. There is a significant correlation between SWE values of cortex and medulla of the kidneys in healthy children.

Dapagliflozin Treatment Augments Bioactive Phosphatidylethanolamine Concentrations in Kidney Cortex Membrane Fractions of Hypertensive Diabetic db/db Mice and Alters the Density of Lipid Rafts in Mouse Proximal Tubule Cells.

In addition to inhibiting renal glucose reabsorption and allowing for glucose excretion, the sodium/glucose cotransporter 2 (SGLT2) inhibitor dapagliflozin may be efficacious in treating various comorbidities associated with type 2 diabetes mellitus (T2DM). The molecular mechanisms by which dapagliflozin exerts its beneficial effects are largely unknown. We hypothesized dapagliflozin treatment in the diabetic kidney alters plasma membrane lipid composition, suppresses extracellular vesicle (EV) release from kidney cells, and disrupts lipid rafts in proximal tubule cells. In order to test this hypothesis, we treated diabetic db/db mice with dapagliflozin (N = 8) or vehicle (N = 8) and performed mass spectrometry-based lipidomics to investigate changes in the concentrations of membrane lipids in the kidney cortex. In addition, we isolated urinary EVs (uEVs) from urine samples collected during the active phase and the inactive phase of the mice and then probed for changes in membrane proteins enriched in the EVs. Multiple triacylglycerols (TAGs) were enriched in the kidney cortex membrane fractions of vehicle-treated diabetic db/db mice, while the levels of multiple phosphatidylethanolamines were significantly higher in similar mice treated with dapagliflozin. EV concentration and size were lesser in the urine samples collected during the inactive phase of dapagliflozin-treated diabetic mice. In cultured mouse proximal tubule cells treated with dapagliflozin, the lipid raft protein caveolin-1 shifted from less dense fractions to more dense sucrose density gradient fractions. Taken together, these results suggest dapagliflozin may regulate lipid-mediated signal transduction in the diabetic kidney.

The nuclear factor of activated T cells 5 (NFAT5) contributes to the renal corticomedullary differences in gene expression.

The corticomedullary osmotic gradient between renal cortex and medulla induces a specific spatial gene expression pattern. The factors that controls these differences are not fully addressed. Adaptation to hypertonic environment is mediated by the actions of the nuclear factor of activated T-cells 5 (NFAT5). NFAT5 induces the expression of genes that lead to intracellular accumulation of organic osmolytes. However, a systematical analysis of the NFAT5-dependent gene expression in the kidneys was missing. We used primary cultivated inner medullary collecting duct (IMCD) cells from control and NFAT5 deficient mice as well as renal cortex and inner medulla from principal cell specific NFAT5 deficient mice for gene expression profiling. In primary NFAT5 deficient IMCD cells, hyperosmolality induced changes in gene expression were abolished. The majority of the hyperosmolality induced transcripts in primary IMCD culture were determined to have the greatest expression in the inner medulla. Loss of NFAT5 altered the expression of more than 3000 genes in the renal cortex and more than 5000 genes in the inner medulla. Gene enrichment analysis indicated that loss of NFAT5 is associated with renal inflammation and increased expression of kidney injury marker genes, like lipocalin-2 or kidney injury molecule-1. In conclusion we show that NFAT5 is a master regulator of gene expression in the kidney collecting duct and in vivo loss of NFAT function induces a kidney injury like phenotype.

The Ischemia and Reperfusion Injury Involves the Toll-Like Receptor-4 Participation Mainly in the Kidney Cortex.

BACKGROUND/AIMS: The renal inflammatory response and kidney regeneration in ischemia-reperfusion injury (IRI) are associated with Toll-like receptor 4 (TLR4). Here we study the role of TLR4 during IRI in the renal cortex and medulla separately, using wild-type (TLR4-WT) and Knockout (TLR4-KO) TLR4 mice. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 48 hours of reperfusion in C57BL/6 mice. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and proteostasis in the renal cortex and medulla by qRT-PCR and Western blot. In addition, we studied kidney morphology by H/E and PAS. RESULTS: Renal ischemia (30min) and reperfusion (48hrs) induced the mRNA and protein of TLR4 in the renal cortex. In addition, Serum Creatinine (SCr), blood urea nitrogen (BUN), Neutrophil gelatinase-associated lipocalin (NGAL), and acute tubular necrosis (ATN) were increased in TLR4-WT by IRI. Interestingly, the SCr and BUN had normal levels in TLR-KO during IRI. However, ATN and high levels of NGAL were present in the kidneys of TLR4-KO mice. The pro-inflammatory (IL-6 and TNF-α) and anti-inflammatory (Foxp3 and IL-10) markers increased by IRI only in the cortex of TLR4-WT but not in TLR4-KO mice. Furthermore, the M1 (CD38 and Frp2) and M2 (Arg-I, Erg-2, and c-Myc) macrophage markers increased by IRI only in the cortex of TLR4-WT. The TLR4-KO blunted the IRI-upregulation of M1 but not the M2 macrophage polarization. Vimentin increased in the renal cortex and medulla of TLR4-WT animals but not in the cortex of TLR4-KO mice. In addition, iNOS and clusterin were increased by IRI only in the cortex of TLR4-WT, and the absence of TLR4 inhibited only clusterin upregulation. Finally, Hsp27 and Hsp70 protein levels increased by IRI in the cortex and medulla of TLR4-WT and TRL4-KO lost the IRI-upregulation of Hsp70. In summary, TLR4 participates in renal ischemia and reperfusion through pro-inflammatory and anti-inflammatory responses inducing impaired kidney function (SCr and BUN). However, the IRI-upregulation of M2 macrophage markers (cortex), iNOS (cortex), IL-6 (medulla), vimentin (medulla), and Hsp27 (cortex and medulla) were independent of TLR4. CONCLUSION: The TLR4 inactivation during IRI prevented the loss of renal function due to the inactivation of inflammation response, avoiding M1 and preserving the M2 macrophage polarization in the renal cortex.

1400W Prevents Renal Injury in the Renal Cortex But Not in the Medulla in a Murine Model of Ischemia and Reperfusion Injury.

BACKGROUND/AIMS: Acute kidney injury (AKI) carries high morbidity and mortality, and the inducible nitric oxide synthase (iNOS) is a potential molecular target to prevent kidney dysfunction. In previous work, we reported that the pharmacological inhibitions of iNOS before ischemia/reperfusion (I/R) attenuate the I/R-induced AKI in mice. Here, we study the iNOS inhibitor 1400W [N-(3-(Aminomethyl)benzyl] acetamide, which has been described to be much more specific to iNOS inhibition than other compounds. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 24 hours of reperfusion in Balb/c mice. 1400w (10 mg/kg i.p) was applied before I/R injury. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and nephrogenic genes by qRT-PCR in the renal cortex and medulla. The Periodic Acid-Schiff (PAS) was used to study the kidney morphology. RESULTS: Remarkably, we found that 1400W affects the renal cortex and medulla in different ways. Thus, in the renal cortex, 1400W prevented the I/R-upregulation of 1. NGAL, Clusterin, and signs of morphological damage; 2. IL-6 and TNF-α; 3. TGF-ß; 4. M2(Arg1, Erg2, cMyc) and M1(CD38, Fpr2) macrophage polarization makers; and 5. Vimentin and FGF2 levels but not in the renal medulla. CONCLUSION: 1400W conferred protection in the kidney cortex compared to the kidney medulla. The present investigation provides relevant information to understand the opportunity to use 1400W as a therapeutic approach in AKI treatment.

Blood pressure and the kidney cortex transcriptome response to high-sodium diet challenge in female nonhuman primates.

Blood pressure (BP) is influenced by genetic variation and sodium intake with sex-specific differences; however, studies to identify renal molecular mechanisms underlying the influence of sodium intake on BP in nonhuman primates (NHP) have focused on males. To address the gap in our understanding of molecular mechanisms regulating BP in female primates, we studied sodium-naïve female baboons (n = 7) fed a high-sodium (HS) diet for 6 wk. We hypothesized that in female baboons variation in renal transcriptional networks correlates with variation in BP response to a high-sodium diet. BP was continuously measured for 64-h periods throughout the study by implantable telemetry devices. Sodium intake, blood samples for clinical chemistries, and ultrasound-guided kidney biopsies were collected before and after the HS diet for RNA-Seq and bioinformatic analyses. We found that on the LS diet but not the HS diet, sodium intake and serum 17 ß-estradiol concentration correlated with BP. Furthermore, kidney transcriptomes differed by diet-unbiased weighted gene coexpression network analysis revealed modules of genes correlated with BP on the HS diet but not the LS diet. Our results showed variation in BP on the HS diet correlated with variation in novel kidney gene networks regulated by ESR1 and MYC; i.e., these regulators have not been associated with BP regulation in male humans or rodents. Validation of the mechanisms underlying regulation of BP-associated gene networks in female NHP will inform better therapies toward greater precision medicine for women.

Effects of Norepinephrine on Renal Cortical and Medullary Blood Flow in Atherosclerotic Rabbits.

OBJECTIVE: The aim of this study was to explore the effect of norepinephrine (NE) on renal cortical and medullary blood flow in atherosclerotic rabbits without renal artery stenosis. METHODS: Atherosclerosis was induced in 21 New Zealand white rabbits by feeding them a cholesterol-rich diet for 16 weeks. Thirteen healthy New Zealand white rabbits were randomly selected as controls. After atherosclerosis induction, standard ultrasonography was performed to confirm that there was no plaque or accelerated flow at the origin of the renal artery. Contrast-enhanced ultrasound (CEUS) was performed at baseline and during intravenous injection of NE. The degree of contrast enhancement of renal cortex and medulla after the injection of contrast agents was quantified by calculating the enhanced intensity. RESULTS: The serum nitric oxide (NO) level in atherosclerotic rabbits was higher than that in healthy rabbits (299.6±152 vs. 136.5±49.5, P<0.001). The infusion of NE induced a significant increase in the systolic blood pressure (112±14 mmHg vs. 84±9 mmHg, P=0.016) and a significant decrease in the enhanced intensity in renal cortex (17.78±2.07 dB vs. 21.19±2.03 dB, P<0.001) and renal medulla (14.87±1.82 dB vs. 17.14±1.89 dB, P<0.001) during CEUS. However, the enhanced intensity in the cortex and medulla of healthy rabbits after NE infusion showed no significant difference from that at baseline. CONCLUSION: NE may reduce renal cortical and medullary blood flow in atherosclerotic rabbits without renal artery stenosis, partly by reducing the serum NO level.

Analysis of the Hypoxic Response in a Mouse Cortical Collecting Duct-Derived Cell Line Suggests That Esrra Is Partially Involved in Hif1α-Mediated Hypoxia-Inducible Gene Expression in mCCDcl1 Cells.

The kidney is strongly dependent on a continuous oxygen supply, and is conversely highly sensitive to hypoxia. Controlled oxygen gradients are essential for renal control of solutes and urine-concentrating mechanisms, which also depend on various hormones including aldosterone. The cortical collecting duct (CCD) is part of the aldosterone-sensitive distal nephron and possesses a key function in fine-tuned distal salt handling. It is well known that aldosterone is consistently decreased upon hypoxia. Furthermore, a recent study reported a hypoxia-dependent down-regulation of sodium currents within CCD cells. We thus investigated the possibility that cells from the cortical collecting duct are responsive to hypoxia, using the mouse cortical collecting duct cell line mCCDcl1 as a model. By analyzing the hypoxia-dependent transcriptome of mCCDcl1 cells, we found a large number of differentially-expressed genes (3086 in total logFC< −1 or >1) following 24 h of hypoxic conditions (0.2% O2). A gene ontology analysis of the differentially-regulated pathways revealed a strong decrease in oxygen-linked processes such as ATP metabolic functions, oxidative phosphorylation, and cellular and aerobic respiration, while pathways associated with hypoxic responses were robustly increased. The most pronounced regulated genes were confirmed by RT-qPCR. The low expression levels of Epas1 under both normoxic and hypoxic conditions suggest that Hif-1α, rather than Hif-2α, mediates the hypoxic response in mCCDcl1 cells. Accordingly, we generated shRNA-mediated Hif-1α knockdown cells and found Hif-1α to be responsible for the hypoxic induction of established hypoxically-induced genes. Interestingly, we could show that following shRNA-mediated knockdown of Esrra, Hif-1α protein levels were unaffected, but the gene expression levels of Egln3 and Serpine1 were significantly reduced, indicating that Esrra might contribute to the hypoxia-mediated expression of these and possibly other genes. Collectively, mCCDcl1 cells display a broad response to hypoxia and represent an adequate cellular model to study additional factors regulating the response to hypoxia.

Inhibition of phosphate transport by NAD+/NADH brush border membrane vesicles.

Nicotinamide is an important regulator of Pi homeostasis after conversion into NAD+/NADH. In this work, we have studied the classical inhibition of Pi transport by these compounds in the brush border membrane vesicles (BBMV) of rat kidney and rat intestine, and we examined the effects in opossum kidney (OK) cells and in phosphate transporter-expressing Xenopus laevis oocytes. In BBMV, NAD+ required preincubation at either room temperature or on ice to inhibit Pi uptake in BBMV. However, no effects were observed in the known Slc34 or Slc20 Pi transporters expressed in Xenopus oocytes, in OK cells, or in isolated rat cortical nephron segments. In BBMV from jejunum or kidney cortex, the inhibition of Pi transport was specific, dose-related, and followed a competitive inhibition pattern, as shown by linear transformation and nonlinear regression analyses. A Ki value of 538 µM NAD+ in kidney BBMV was obtained. Ribosylation inhibitors and ribosylation assays revealed no evidence that this reaction was responsible for inhibiting Pi transport. An analysis of the persistence of NAD+/NADH revealed a half-life of just 2 min during preincubation. Out of several metabolites of NAD degradation, only ADP-ribose was able to inhibit Pi uptake. Pi concentration also increased during 30 min of preincubation, up to 0.67 mM, most likely as a metabolic end product. In conclusion, the classical inhibition of Pi transport by NAD+/NADH in BBMV seems to be caused by the degradation metabolites of these compounds during the preincubation time.

Supplementation of amino acids and organic acids prevents the increase in blood pressure induced by high salt in Dahl salt-sensitive rats.

A high-salt (HS) diet leads to metabolic disorders in Dahl salt-sensitive (SS) rats, and promotes the development of hypertension. According to the changes in the metabolites of SS rats, a set of combined dietary supplements containing amino acids and organic acids (AO) were designed. The purpose of the present study was to evaluate the effect of AO supplementation on the blood pressure of SS rats after the HS diet and clarify the mechanism of AO by metabolomics and biochemical analyses. The results showed that AO supplementation avoided the elevation of blood pressure induced by the HS diet in SS rats, increased the renal antioxidant enzyme activities (catalase, superoxide dismutase, glutathione reductase, and glutathione S-transferase), reduced the H2O2 and MDA levels, and restored the normal antioxidant status of the serum and kidneys. AO also reversed the decrease in the nitric oxide (NO) levels and NO synthase activity induced by the HS feed, which involved the L-arginine/NO pathway. Metabolomics analysis showed that AO administration increased the levels of amino acids such as cysteine, glycine, hypotaurine, and lysine in the renal medulla and the levels of leucine, isoleucine, and serine in the renal cortex. Of note, lysine, hypotaurine and glycine had higher metabolic centrality in the metabolic correlation network of the renal medulla after AO administration. In conclusion, AO intervention could prevent HS diet-induced hypertension in SS rats by restoring the metabolic homeostasis of the kidneys. Hence, AO has the potential to become a functional food additive to improve salt-sensitive hypertension.