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.

A Higher Foci Density of Interstitial Fibrosis and Tubular Atrophy Predicts Progressive CKD after a Radical Nephrectomy for Tumor.

BACKGROUND: Chronic tubulointerstitial injury on kidney biopsy is usually quantified by the percentage of cortex with interstitial fibrosis/tubular atrophy (IF/TA). Whether other patterns of IF/TA or inflammation in the tubulointerstitium have prognostic importance beyond percentage IF/TA is unclear. METHODS: We obtained, stained, and digitally scanned full cortical thickness wedge sections of renal parenchyma from patients who underwent a radical nephrectomy for a tumor over 2000-2015, and morphometrically analyzed the tubulointerstitium of the cortex for percentage IF/TA, IF/TA density (foci per mm2 cortex), percentage subcapsular IF/TA, striped IF/TA, percentage inflammation (both within and outside IF/TA regions), and percentage subcapsular inflammation. Patients were followed with visits every 6-12 months. Progressive CKD was defined as dialysis, kidney transplantation, or 40% decline from the postnephrectomy eGFR. Cox models assessed the risk of CKD or noncancer mortality with morphometric measures of tubulointerstitial injury after adjustment for the percentage IF/TA and clinical characteristics. RESULTS: Among 936 patients (mean age, 64 years; postnephrectomy baseline eGFR, 48 ml/min per 1.73m2), 117 progressive CKD events and 183 noncancer deaths occurred over a median 6.4 years. Higher IF/TA density predicted both progressive CKD and noncancer mortality after adjustment for percentage IF/TA and predicted progressive CKD after further adjustment for clinical characteristics. Independent of percentage IF/TA, age, and sex, higher IF/TA density correlated with lower eGFR, smaller nonsclerosed glomeruli, more global glomerulosclerosis, and smaller total cortical volume. CONCLUSIONS: Higher density of IF/TA foci (a more scattered pattern with more and smaller foci) predicts higher risk of progressive CKD after radical nephrectomy compared with the same percentage of IF/TA but with fewer and larger foci.

Overexpressing STAMP2 attenuates diabetic renal injuries via upregulating autophagy in diabetic rats.

Diabetic nephropathy (DN) is one of the most serious and major renal complications of diabetes. Previously, Six-transmembrane Protein of Prostate 2 (STAMP2) was reported to contribute to nutritional stress. The purpose of this study is to investigate whether overexpression of STAMP2 attenuates diabetic renal injuries in DN rats. We induced the DN rat model by high-fat diet and low-dose streptozotocin and evaluated the metabolite and urine albumin/creatinine. Recombinant adeno-associated virus vectors were injected for overexpression of STAMP2. Pathophysiologic and ultrastructure features of DN by histochemical stain and transmission electron microscope, autophagy-related proteins and signaling pathway by western blotting were assessed. We found the expression of STAMP2 was decreased and autophagy was blunted in DN rat kidneys. Overexpressing STAMP2 significantly ameliorated metabolic disturbance, insulin resistance, and specifically restoring diabetic renal injury. Furthermore, overexpressing STAMP2 improved the autophagy deficiency in DN rats, as revealed by changes in the expressions of Beclin1, p62, and LC3. Furthermore, STAMP2 overexpressing promoted autophagy by inhibiting the mTOR and activating the AMPK/SIRT1 signaling pathway. Our results suggested that STAMP2 overexpression attenuated renal injuries via upregulating autophagy in DN rats. STAMP2 overexpressing promoted autophagy may been involved with inhibition of the mTOR/ULK1 and activation of the AMPK/SIRT1 signaling pathway.

Integrative transcriptomic profiling of a mouse model of hypertension-accelerated diabetic kidney disease.

The current understanding of molecular mechanisms driving diabetic kidney disease (DKD) is limited, partly due to the complex structure of the kidney. To identify genes and signalling pathways involved in the progression of DKD, we compared kidney cortical versus glomerular transcriptome profiles in uninephrectomized (UNx) db/db mouse models of early-stage (UNx only) and advanced [UNxplus adeno-associated virus-mediated renin-1 overexpression (UNx-Renin)] DKD using RNAseq. Compared to normoglycemic db/m mice, db/db UNx and db/db UNx-Renin mice showed marked changes in their kidney cortical and glomerular gene expression profiles. UNx-Renin mice displayed more marked perturbations in gene components associated with the activation of the immune system and enhanced extracellular matrix remodelling, supporting histological hallmarks of progressive DKD in this model. Single-nucleus RNAseq enabled the linking of transcriptome profiles to specific kidney cell types. In conclusion, integration of RNAseq at the cortical, glomerular and single-nucleus level provides an enhanced resolution of molecular signalling pathways associated with disease progression in preclinical models of DKD, and may thus be advantageous for identifying novel therapeutic targets in DKD.

Early critical cortical infarction by anti-angiotensin II type 1 receptor antibody: A case report and literature review.

RATIONALE: Anti-angiotensin II type 1 receptor antibodies (AT1R-Abs) have been demonstrated to increase the risk of antibody-mediated rejection. We report a case of AT1R-Ab mediated rejection which caused early critical cortical infarction. PATIENT CONCERNS: A 52-year-old man with end-stage kidney disease underwent preemptive kidney transplantation (KT) from his wife. He had no immunologic risk except ABO incompatibility. Proper desensitization treatment were applied prior to KT. On postoperative day 1, he showed stable clinical course with adequate urine output, but there was no decrease in serum creatinine level and imaging studies showed hypoperfusion in the transplanted kidney. DIAGNOSES: Allograft biopsy revealed total cortical infarction with severe necrotizing vasculitis, but the medullary area was preserved. Serum AT1R-Ab concentration was elevated from 10.9 U/mL before KT to 19.1 U/mL on 7 days after KT. INTERVENTIONS: He was treated with plasmapheresis, intravenous immunoglobulin, rituximab, high-dose methylprednisolone, and bortezomib. OUTCOMES: The treatment showed a partial response, and he was discharged with 7.3 mg/dL creatinine level. At 4 months, his creatinine plateaued at 5.5 mg/dL and AT1R-Ab decreased to 3.6 U/mL. LESSONS: This case highlights the risk of early active antibody-mediated rejection by preformed AT1R-Ab, suggesting its ability to exhibit atypical histopathologic findings, such as total cortical infarction.

Shiga Toxin (Stx)-Binding Glycosphingolipids of Primary Human Renal Cortical Epithelial Cells (pHRCEpiCs) and Stx-Mediated Cytotoxicity.

Human kidney epithelial cells are supposed to be directly involved in the pathogenesis of the hemolytic-uremic syndrome (HUS) caused by Shiga toxin (Stx)-producing enterohemorrhagic Escherichia coli (EHEC). The characterization of the major and minor Stx-binding glycosphingolipids (GSLs) globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer), respectively, of primary human renal cortical epithelial cells (pHRCEpiCs) revealed GSLs with Cer (d18:1, C16:0), Cer (d18:1, C22:0), and Cer (d18:1, C24:1/C24:0) as the dominant lipoforms. Using detergent-resistant membranes (DRMs) and non-DRMs, Gb3Cer and Gb4Cer prevailed in the DRM fractions, suggesting their association with microdomains in the liquid-ordered membrane phase. A preference of Gb3Cer and Gb4Cer endowed with C24:0 fatty acid accompanied by minor monounsaturated C24:1-harboring counterparts was observed in DRMs, whereas the C24:1 fatty acid increased in relation to the saturated equivalents in non-DRMs. A shift of the dominant phospholipid phosphatidylcholine with saturated fatty acids in the DRM to unsaturated species in the non-DRM fractions correlated with the GSL distribution. Cytotoxicity assays gave a moderate susceptibility of pHRCEpiCs to the Stx1a and Stx2a subtypes when compared to highly sensitive Vero-B4 cells. The results indicate that presence of Stx-binding GSLs per se and preferred occurrence in microdomains do not necessarily lead to a high cellular susceptibility towards Stx.

Ultrasound Findings and Laboratory Predictors of Early Mortality in Patients With Severe Yellow Fever.

OBJECTIVE. Yellow fever is a hemorrhagic disease caused by an arbovirus endemic in South America; outbreaks have occurred in recent years. The purpose of this study was to describe abdominal ultrasound findings in patients with severe yellow fever and correlate them with clinical and laboratory data. MATERIALS AND METHODS. A retrospective cohort study was performed between January and April 2018. The subjects were patients admitted to an ICU with polymerase chain reaction-confirmed yellow fever. Bedside sonography was performed within 48 hours of admission. Images were independently analyzed by two board-certified radiologists. Laboratory test samples were collected within 12 hours of image acquisition. Multivariable logistic regression analysis was performed to identify 30-day mortality predictors; p < .05 was considered statistically significant. RESULTS. Forty-six patients (40 [87%] men, six [13%] women; mean age, 47.5 ± 15.2 years) were evaluated with bedside sonography. Laboratory tests showed high serum levels of aspartate aminotransferase (5319 U/L), total bilirubin (6.2 mg/dL), and creati-nine (4.3 mg/dL). Twenty-six (56.5%) patients died within 30 days of admission (median time to death, 5 days [interquartile range, 2-9 days]). The most frequent ultrasound findings were gallbladder wall thickening (80.4%), increased renal cortex echogenicity (71.7%), increased liver parenchyma echogenicity (65.2%), perirenal fluid (52.2%), and ascites (30.4%). Increased renal echogenicity was associated with 30-day mortality (84.6% versus 55.0%; p = .046) and was an independent predictor of this outcome after multivariate analysis (odds ratio, 10.89; p = .048). CONCLUSION. Reproducible abdominal ultrasound findings in patients with severe yellow fever may be associated with severity of disease and prognosis among patients treated in the ICU.

DHEA-pretreatment attenuates oxidative stress in kidney-cortex and liver of diabetic rabbits and delays development of the disease.

In view of reported discrepancies concerning antioxidant activity of dehydroepiandrosterone (DHEA), a widely used dietary supplement, the current investigation was undertaken to evaluate the antioxidant properties of DHEA in both kidney-cortex and liver of alloxan (ALX)-induced diabetic rabbits, as this diabetogenic compound exhibits the ROS-dependent action. ALX was injected to animals following 7 days of DHEA administration. Four groups of rabbits were used in the experiments: control, DHEA-treated control, diabetic and DHEA-treated diabetic. Our results show for the first time, that in kidney-cortex DHEA resulted in normalization of hydroxyl free radicals (HFR) levels and restoration of catalase (CAT) and glutathione peroxidase (GPx) activities to near the control values, while in liver DHEA prevented the malondialdehyde (MDA) accumulation and normalized glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH) activities. Moreover, in both kidney-cortex and liver DHEA supplementation prevented GSSG elevation accompanied by a decrease in GSH/GSSG ratio. Although DHEA attenuated oxidative stress in both kidney-cortex and liver of ALX-induced diabetic rabbits and significantly delayed the onset of diabetes in time, it did not protect against the final development of diabetes. In conclusion, the current investigation underscores the complexity of the antioxidant action of DHEA. The data are of clinical interest since DHEA supplementation could prevent the deleterious effects of ROS and delay, or even prevent the onset of many diseases. However, in view of the reported pro-oxidant effects of high DHEA doses, the potential use of this agent as a supplement needs a careful evaluation.

Changes in NAD and Lipid Metabolism Drive Acidosis-Induced Acute Kidney Injury.

BACKGROUND: The kidney plays an important role in maintaining normal blood pH. Metabolic acidosis (MA) upregulates the pathway that mitochondria in the proximal tubule (PT) use to produce ammonia and bicarbonate from glutamine, and is associated with AKI. However, the extent to which MA causes AKI, and thus whether treating MA would be beneficial, is unclear. METHODS: Gavage with ammonium chloride induced acute MA. Multiphoton imaging of mitochondria (NADH/membrane potential) and transport function (dextran/albumin uptake), oxygen consumption rate (OCR) measurements in isolated tubules, histologic analysis, and electron microscopy in fixed tissue, and urinary biomarkers (KIM-1/clara cell 16) assessed tubular cell structure and function in mouse kidney cortex. RESULTS: MA induces an acute change in NAD redox state (toward oxidation) in PT mitochondria, without changing the mitochondrial energization state. This change is associated with a switch toward complex I activity and decreased maximal OCR, and a major alteration in normal lipid metabolism, resulting in marked lipid accumulation in PTs and the formation of large multilamellar bodies. These changes, in turn, lead to acute tubular damage and a severe defect in solute uptake. Increasing blood pH with intravenous bicarbonate substantially improves tubular function, whereas preinjection with the NAD precursor nicotinamide (NAM) is highly protective. CONCLUSIONS: MA induces AKI via changes in PT NAD and lipid metabolism, which can be reversed or prevented by treatment strategies that are viable in humans. These findings might also help to explain why MA accelerates decline in function in CKD.

Does the renal parenchymal thickness affect the efficacy of the retrograde intrarenal surgery? A prospective cohort study.

Retrograde intrarenal surgery (RIRS) is one of the minimally invasive main treatment modalities in renal stone disease. There are many factors which affect stone-free rate (SFR). Our study was based on the hypothesis that higher renal parenchymal thickness (RPT) which may include higher average number of nephrons provides better diuresis. We investigated the efficacy of RPT on success of RIRS. This study is a single-centered prospective surgical cohort study. A total of 383 patients were analyzed. Regularly followed 304 patients with unilateral kidney stone at single pole or renal pelvis and who underwent single-session RIRS were included in the final analysis, and the patients' preoperative and postoperative 1st and 3rd months' data were evaluated. RPT was measured on the non-contrast computed tomography (CT) images. ROC analysis was performed to estimate the cutoff value of RPT for SFR. Univariate and multivariate logistic regression analyses were used to model the relationship between RPT and SFR after RIRS. ROC analysis revealed the best cutoff value of the RPT for predicting residual stone as 19 mm for both the 1st and 3rd month visits with Youden indexes of 0.397 and 0.406, respectively. To the best of our knowledge, this is the first study which evaluated the effect of RPT on the efficacy of RIRS. RPT measurement is a cost-effective method that can be easily performed on routinely applied non-contrast CT and may have predictive value for the surgical success in patients with nephrolithiasis.

Kidney Single-cell Transcriptomes Predict Spatial Corticomedullary Gene Expression and Tissue Osmolality Gradients.

BACKGROUND: Single-cell transcriptomes from dissociated tissues provide insights into cell types and their gene expression and may harbor additional information on spatial position and the local microenvironment. The kidney's cells are embedded into a gradient of increasing tissue osmolality from the cortex to the medulla, which may alter their transcriptomes and provide cues for spatial reconstruction. METHODS: Single-cell or single-nuclei mRNA sequencing of dissociated mouse kidneys and of dissected cortex, outer, and inner medulla, to represent the corticomedullary axis, was performed. Computational approaches predicted the spatial ordering of cells along the corticomedullary axis and quantitated expression levels of osmo-responsive genes. In situ hybridization validated computational predictions of spatial gene-expression patterns. The strategy was used to compare single-cell transcriptomes from wild-type mice to those of mice with a collecting duct-specific knockout of the transcription factor grainyhead-like 2 (Grhl2CD-/-), which display reduced renal medullary osmolality. RESULTS: Single-cell transcriptomics from dissociated kidneys provided sufficient information to approximately reconstruct the spatial position of kidney tubule cells and to predict corticomedullary gene expression. Spatial gene expression in the kidney changes gradually and osmo-responsive genes follow the physiologic corticomedullary gradient of tissue osmolality. Single-nuclei transcriptomes from Grhl2CD-/- mice indicated a flattened expression gradient of osmo-responsive genes compared with control mice, consistent with their physiologic phenotype. CONCLUSIONS: Single-cell transcriptomics from dissociated kidneys facilitated the prediction of spatial gene expression along the corticomedullary axis and quantitation of osmotically regulated genes, allowing the prediction of a physiologic phenotype.

Tissue-selective alternate promoters guide NLRP6 expression.

The pryin domain (PYD) domain is involved in protein interactions that lead to assembly of immune-sensing complexes such as inflammasomes. The repertoire of PYD-containing genes expressed by a cell type arms tissues with responses against a range of stimuli. The transcriptional regulation of the PYD gene family however is incompletely understood. Alternative promoter utilization was identified as a mechanism regulating the tissue distribution of human PYD gene family members, including NLRP6 that is translationally silenced outside of intestinal tissue. Results show that alternative transcriptional promoters mediate NLRP6 silencing in mice and humans, despite no upstream genomic synteny. Human NLRP6 contains an internal alternative promoter within exon 2 of the PYD, resulting in a truncated mRNA in nonintestinal tissue. In mice, a proximal promoter was used that expanded the 5' leader sequence restricting nuclear export and abolishing translational efficiency. Nlrp6 was dispensable in disease models targeting the kidney, which expresses noncanonical isoforms. Thus, alternative promoter use is a critical mechanism not just for isoform modulation but for determining expression profile and function of PYD family members.

Histone Deacetylase Activity and the Renin-Angiotensin-Aldosterone System: Key Elements in Cardiorenal Alterations Provoked by Chronic Malnutrition in Male Adult Rats.

BACKGROUND/AIMS: Chronic malnutrition (M) affects >1 billion people worldwide. Epidemiological data point to long-term renal and cardiovascular outcomes (e.g. arterial hypertension, cardiorenal syndromes). The renin-angiotensin-aldosterone system (RAAS) has been implicated in the physiopathology of these disturbances, but M-induced alterations in RAAS-modulated renal Na+ handling and their cardiovascular repercussions are not known. Moreover, altered tissue-specific histone deacetylases (HDAC) results in arterial hypertension and the use of sodium Valproate (Val; a HDAC inhibitor) reduces blood pressure. However, there are no reports regarding the renal and cardiovascular effects of HDAC inhibition in M, or on the signaling pathways involved. The central aim of our study has been to investigate whether alterations in the HDAC/RAAS axis underpin alterations in active Na+ transport in the kidney and heart, and affects blood pressure. METHODS: Male rats aged 28 days were given either a control (C) or a multideficient diet (Regional Basic Diet, RBD), which mimics alimentary habits from developing countries. Subgroups received Losartan (Los), a blocker of type 1 Angiotensin II receptors. When the rats reached 70 days, new subgroups received Val until they were 90 days of age. Homogenates and enriched plasma membrane fractions from renal cortex corticis and cardiomyocytes were obtained by differential centrifugation of the tissues. The activity of renal and cardiac deacetylases was assayed by measuring - after incubation with the membranes - the amount of deacetylated lysines in a substrate containing an acetylated lysine side chain. Protein kinases activities were measured following the incorporation of the γ-phosphoryl group of [γ-32P]ATP into Ser/Thr residues of histone type III-S. The activity of Na+-transporting ATPases (kidney and heart) was quantified by measuring the release of Pi from ATP that was sensitive to ouabain ((Na++K+)ATPase), or sensitive to furosemide (Na+-ATPase). Tail-cuff plethysmography was used to measure systolic blood pressure and heart rate. RESULTS: M provoked HDAC downregulation, which was reversed by Los and Val, either alone or in combination, with selective upregulation of protein kinases C and A (PKC, PKA) in renal cortex corticis, but not in left ventricle cardiomyocytes. The 2 kinases were strongly inhibited by Los and Val in both organs. Malnourished rats developed elevated systolic arterial pressure (SAP) and heart rate (HR) at 70 days of age; Los and Val restored the control SAP, but not HR. Functional and the above biochemical alterations were associated with the deregulation of renal and cardiac Na+-transporting ATPases. (Na++K+)ATPase activities were downregulated in M rats in both organs, and were further inhibited by the pharmacological treatments in the renal cortex corticis (C and M groups) and the left ventricle (only in C rats). No additional effect was found in cardiac (Na++K+)ATPase from M rats. Ouabain-resistant Na+-ATPase was upregulated in renal cortex corticis and downregulated in cardiomyocytes, returning to C values after administration of Los and Val. CONCLUSION: The HDAC/RAAS axis appears to be a key regulator of Na+-transporting ATPases in renal cortex corticis and cardiomyocytes via an appropriate balance of PKC and PKA activities. Modifications within the HDAC/RAAS axis provoked by chronic M - with repercussions in renal and cardiac Na+ transport - underpin alterations in bodily Na+ homeostasis that culminate with the onset of arterial hypertension and potential cardiorenal syndrome.

Combinational Use of Antiplatelet Medication Sarpogrelate with Therapeutic Drug Rosuvastatin in Treating High-Cholesterol Diet-Induced Chronic Kidney Disease in ApoE-Deficient Mice.

A number of metabolic disorders, including hyperlipidemia, potentially cause chronic kidney disease (CKD), one of their major chronic complications and comorbidities. Rosuvastatin is one of the widely used antiatherogenic drugs among hyperlipidemic patients. Meanwhile, sarpogrelate is not only a 5-hydroxytryptamine receptor antagonist but also an antiplatelet agent, inhibiting platelet-stimulated blood coagulation and improving peripheral circulation. In this study, a combination of sarpogrelate and/or rosuvastatin was used on CKD mice induced by a high-fat diet for 8 weeks. The mice were tested for pathological changes using histological evaluation. Tremendous alterations were found, including a remarked increase in total cholesterol and low-density lipoprotein cholesterol levels, glomerular endothelial proliferation, and mesangial expansion. Also, tubular damage and extracellular matrix accumulation occurred, namely, a marked increase in the macula densa, scattered and apoptotic loss of the apical brush border with vacuolated basophilic cytoplasm and heavily stained nuclei, and expanded Bowman's space, which were at least partially ameliorated by sarpogrelate and/or rosuvastatin treatment. The analysis of expression profiles at both the RNA and protein levels, using real-time quantitative polymerase chain reaction and Western blot analysis, indicated that LDL-R/CD68/LOX-1-positive monocyte/macrophage-mediated enhanced proinflammatory activation, including the significant upregulation of tumor necrosis factor-α and interleukin-6, was actually attenuated by sarpogrelate and/or rosuvastatin treatment. The findings indicated that sarpogrelate and/or rosuvastatin treatment potentially ameliorates CKD progression in patients with the aforementioned comorbid metabolic disorders.

CaMKIV regulates mitochondrial dynamics during sepsis.

Sepsis and shock states impose mitochondrial stress, and in response, adaptive mechanisms such as fission, fusion and mitophagy are induced to eliminate damaged portions of or entire dysfunctional mitochondria. The mechanisms underlying these events are being elucidated; yet a direct link between loss of mitochondrial membrane potential ΔΨm and the initiation of fission, fusion and mitophagy remains to be well characterized. The direct association between the magnitude of the ΔΨm and the capacity for mitochondria to buffer Ca2+ renders Ca2+ uniquely suited as the signal engaging these mechanisms in circumstances of mitochondrial stress that lower the ΔΨm. Herein, we show that the calcium/calmodulin-dependent protein kinase (CaMK) IV mediates an adaptive slowing in oxidative respiration that minimizes oxidative stress in the kidneys of mice subjected to either cecal ligation and puncture (CLP) sepsis or endotoxemia. CaMKIV shifts the balance towards mitochondrial fission and away from fusion by 1) directly phosphorylating an activating Serine616 on the fission protein DRP1 and 2) reducing the expression of the fusion proteins Mfn1/2 and OPA-1. CaMKIV, through its function as a direct PINK1 kinase and regulator of Parkin expression, also enables mitophagy. These data support that CaMKIV serves as a keystone linking mitochondrial stress with the adaptive mechanisms of mitochondrial fission, fusion and mitophagy that mitigate oxidative stress in the kidneys of mice responding to sepsis.

Comprehensive Lipidome Profiling of the Kidney in Early-Stage Diabetic Nephropathy.

Metabolic changes associated with diabetes are reported to lead to the onset of early-stage diabetic nephropathy (DN). Furthermore, lipotoxicity is implicated in renal dysfunction. Most studies of DN have focused on a single or limited number of lipids, and the lipidome of the kidney during early-stage DN remains to be elucidated. In the present study, we aimed to comprehensively identify lipid abnormalities during early-stage DN; to this end, we established an early-stage DN rat model by feeding a high-sucrose and high-fat diet combined with administration of low-dose streptozotocin. Using a high-coverage, targeted lipidomic approach, we established the lipid profile, comprising 437 lipid species and 25 lipid classes, of the kidney cortex in normal rats and the DN rat model. Our findings additionally confirmed that the DN rat model had been successfully established. We observed distinct lipidomic signatures in the DN kidney, with characteristic alterations in side chain composition and degree of unsaturation. Glyceride lipids, especially cholesteryl esters, showed a significant increase in the DN kidney cortex. The levels of most phospholipids exhibited a decline, except those of phospholipids with side chain of 36:1. Furthermore, the levels of lyso-phospholipids and sphingolipids, including ceramide and its derivatives, were dramatically elevated in the present DN rat model. Our findings, which provide a comprehensive lipidome of the kidney cortex in rats with DN, are expected to be useful for the identification of pathologically relevant lipid species in DN. Furthermore, the results represent novel insights into the mechanistic basis of DN.

Melatonin Suppresses Renal Cortical Fibrosis by Inhibiting Cytoskeleton Reorganization and Mitochondrial Dysfunction through Regulation of miR-4516.

Renal fibrosis, a major risk factor for kidney failure, can lead to chronic kidney disease (CKD) and is caused by cytoskeleton reorganization and mitochondrial dysfunction. In this study, we investigated the potential of melatonin treatment to reduce renal fibrosis by recovering the cytoskeleton reorganization and mitochondrial dysfunction. We found that miR-4516 expression was downregulated in the renal cortex of CKD mice and P-cresol-treated TH1 cells. Decreased miR-4516 expression stimulated cytoskeleton reorganization and mitochondrial dysfunction, and induced renal fibrosis. Melatonin treatment suppressed fibrosis by inhibiting cytoskeleton reorganization and restoring mitochondrial function via increased miR-4516 expression. More specifically, melatonin treatment increased miR-4516 expression while decreasing ITGA9 expression, thereby inhibiting cytoskeleton reorganization. In addition, increased expression of miR-4516 by melatonin treatment reduced ROS formation and restored mitochondrial function. These findings suggest that melatonin may be a promising treatment for patients with CKD having renal fibrosis. Moreover, regulation of miR-4516 expression may be a novel strategy for the treatment of renal fibrosis.

The correlation analysis between the Oxford classification of Chinese IgA nephropathy children and renal outcome - a retrospective cohort study.

BACKGROUND: The 2016 Oxford Classification's MEST-C scoring system predicts outcomes in adults with IgA nephropathy (IgAN), but it lacks tremendous cohort validation in children with IgAN in China. We sought to verify whether the Oxford classification could be used to predict the renal outcome of children with IgAN. METHODS: In this retrospective cohort study, 1243 Chinese IgAN children who underwent renal biopsy in Jinling Hospital were enregistered from 2000 to 2017. The combined endpoint was defined as either a ≥ 50% reduction in estimated glomerular filtration rate (eGFR) or end-stage renal disease (ESRD). We probed into the relevance betwixt the Oxford classification and renal prognosis. RESULTS: There were 29% of children with mesangial proliferation(M1), 35% with endocapillary proliferation (E1), 37% with segmental sclerosis/adhesion lesion (S1), 23% with moderate tubular atrophy/interstitial fibrosis (T1 25-50% of cortical area involved), 4.3% with severe tubular atrophy/interstitial fibrosis (T2 > 50% of cortical area involved), 44% with crescent in< 25% of glomeruli(C1), and 4.6% with crescent in> 25% of glomeruli (C2). All children were followed for a medial of 7.2 (4.6-11.7) years, 171 children (14%) arrived at the combined endpoint. The multivariate COX regression model revealed that the presence of lesions S (HR2.7,95%CI 1.8 ~ 4.2, P<0.001) and T (HR6.6,95%CI 3.9 ~ 11.3, P<0.001) may be the reason for poorer prognosis in the whole cohort. In contrast, C lesion showed a significant association with the outcome only in children received no immunosuppressive treatment. CONCLUSIONS: This study revealed that S and T lesions were useful as the long-term renal prognostic factors among Chinese IgAN children.

The Importance of Kidney Medullary Tissue for the Accurate Diagnosis of BK Virus Allograft Nephropathy.

BACKGROUND AND OBJECTIVES: The published tissue adequacy requirement of kidney medulla for BK virus allograft nephropathy diagnosis lacks systematic verification and competes against potential increased procedural risks from deeper sampling. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: We evaluated whether the presence of kidney medulla improved the diagnostic rate of BK nephropathy in 2244 consecutive biopsy samples from 856 kidney transplants with detailed histologic and virologic results. RESULTS: Medulla was present in 821 samples (37%) and correlated with maximal core length (r=0.35; P<0.001). BK virus allograft nephropathy occurred in 74 (3% overall) but increased to 5% (42 of 821) with medulla compared with 2% (32 of 1423) for cortical samples (P<0.001). Biopsy medulla was associated with infection after comprehensive multivariable adjustment of confounders, including core length, glomerular number, and number of cores (adjusted odds ratio, 1.81; 95% confidence interval, 1.02 to 3.21; P=0.04). In viremic cases (n=275), medulla was associated with BK virus nephropathy diagnosis (39% versus 19% for cortex; P<0.001) and tissue polyomavirus load (Banff polyomavirus score 0.64±0.96 versus 0.33±1.00; P=0.006). Biopsy medulla was associated with BK virus allograft nephropathy using generalized estimating equation (odds ratio, 2.04; 95% confidence interval, 1.05 to 3.96; n=275) and propensity matched score comparison (odds ratio, 2.24; 95% confidence interval, 1.11 to 4.54; P=0.03 for 156 balanced pairs). Morphometric evaluation of Simian virus 40 large T immunohistochemistry found maximal infected tubules within the inner cortex and medullary regions (P<0.001 versus outer cortex). CONCLUSIONS: Active BK virus replication concentrated around the corticomedullary junction can explain the higher detection rates for BK virus allograft nephropathy with deep sampling. The current adequacy requirement specifying targeting medulla can be justified to minimize a missed diagnosis from undersampling.

RNA-seq reveals altered gene expression levels in proximal tubular cell cultures compared to renal cortex but not during early glucotoxicity.

Cell cultures are often used to study physiological processes in health and disease. It is well-known that cells change their gene expression in vitro compared to in vivo, but it is rarely experimentally addressed. High glucose is a known trigger of apoptosis in proximal tubular cells (PTC). Here we used RNA-seq to detect differentially expressed genes in cultures of primary rat PTC, 3 days old, compared to cells retrieved directly from rat outer renal cortex and between PTC exposed to 15 mM glucose and control for 8 h. The expression of 6,174 genes was significantly up- or downregulated in the cultures of PTC compared to the cells in the outer renal cortex. Most altered were mitochondrial and metabolism related genes. Gene expression of proapoptotic proteins were upregulated and gene expression of antiapoptotic proteins were downregulated in PTC. Expression of transporter related genes were generally downregulated. After 8 h, high glucose had not altered the gene expression in PTC. The current study provides evidence that cells alter their gene expression in vitro compared to in vivo and suggests that short-term high glucose exposure can trigger apoptosis in PTC without changing the gene expression levels of apoptotic proteins.