Loss of Roundabout Guidance Receptor 2 (Robo2) in Podocytes Protects Adult Mice from Glomerular Injury by Maintaining Podocyte Foot Process Structure

Metabolic syndrome (MetS) is associated with nutrient surplus and kidney hyperfiltration, accelerating chronic renal failure. The potential involvement of podocyte damage in early MetS remains unclear. Mitochondrial dysfunction is an important determinant of renal damage, but whether it contributes to MetS-related podocyte injury remains unknown. Domestic pigs were studied after 16 wk of diet-induced MetS, MetS treated with the mitochondria-targeted peptide elamipretide (ELAM; 0.1 mg·kg−1·day−1 sc) for the last month of diet, and lean controls ( n = 6 pigs/group). Glomerular filtration rate (GFR) and renal blood flow (RBF) were measured using multidetector computed tomography, and podocyte and mitochondrial injury were measured by light and electron microscopy. Urinary levels of podocyte-derived extracellular vesicles (pEVs; nephrin positive/podocalyxin positive) were characterized by flow cytometry. Body weight, blood pressure, RBF, and GFR were elevated in MetS. Glomerular size and glomerular injury score were also elevated in MetS and decreased after ELAM treatment. Evidence of podocyte injury, impaired podocyte mitochondria, and foot process width were all increased in MetS but restored with ELAM. The urinary concentration of pEVs was elevated in MetS pigs and directly correlated with renal dysfunction, glomerular injury, and fibrosis and inversely correlated with glomerular nephrin expression. Additionally, pEV numbers were elevated in the urine of obese compared with lean human patients. Early MetS induces podocyte injury and mitochondrial damage, which can be blunted by mitoprotection. Urinary pEVs reflecting podocyte injury might represent early markers of MetS-related kidney disease and a novel therapeutic target.

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Glomerulopathy does not increase renal susceptibility to acute ischemic injury

AJP Renal Physiology ◽

10.1152/ajprenal.1984.246.3.f272 ◽

1984 ◽

Vol 246 (3) ◽

pp. F272-F281 ◽

Cited By ~ 1

Author(s):

R. A. Zager ◽

L. A. Baltes ◽

H. M. Sharma ◽

W. G. Couser

Keyword(s):

Renal Failure ◽

Acute Renal Failure ◽

Nephrotic Syndrome ◽

Renal Injury ◽

Artery Occlusion ◽

Minimal Change ◽

Glomerular Injury ◽

Bilateral Renal Artery ◽

Foot Process ◽

Ischemic Renal Injury

To determine whether preexistent glomerular injury and the nephrotic syndrome increase renal susceptibility to ischemic renal injury, normal rats and rats with either experimental minimal-change disease (Adriamycin nephropathy) (AN) or membranous nephropathy (passive Heymann nephritis) (PHN) underwent renal functional and histologic studies under either basal conditions or 18 h after bilateral renal artery occlusion (over 30 min). Prior to renal ischemia AN and PHN rats had minimally depressed glomerular filtration rate (GFR), normal (AN) or increased (PHN) renal blood flow (RBF), heavy proteinuria, hypoalbuminemia, decreased urine sodium excretion, extensive glomerular foot process fusion, and intratubular hyalin cast formation. Losses of GFR in response to ischemia were comparable among the three groups of rats (controls, 0.29; AN, 0.28; PHN, 0.25 ml X min-1 X 100 g body wt-1) despite prevailing differences in postischemic hemodynamics. Neither light nor transmission electron microscopy showed any differences in the degree of ischemic renal injury. These results suggest that 1) glomerulopathy and the nephrotic syndrome do not significantly increase renal susceptibility to ischemic renal injury; 2) the syndrome of acute renal failure that occurs in patients with minimal-change glomerulopathy is not due to a marked susceptibility of these kidneys to clinically occult ischemic events; and 3) foot process fusion is probably not a pathophysiologically significant lesion in ischemic acute renal failure, as previously suggested.

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The potential roles of NAD(P)H:quinone oxidoreductase 1 in the development of diabetic nephropathy and actin polymerization

Scientific Reports ◽

10.1038/s41598-020-74493-z ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Sung-Je Moon ◽

Jin Young Jeong ◽

Jae-Hoon Kim ◽

Dong-Hee Choi ◽

Hyunsu Choi ◽

...

Keyword(s):

Diabetic Nephropathy ◽

Actin Polymerization ◽

Major Complication ◽

Glomerular Injury ◽

Normal Range ◽

Actin Reorganization ◽

Nitrogen Levels ◽

Urine Albumin ◽

Foot Process

Abstract Diabetic nephropathy (DN) is a major complication of diabetes mellitus. NAD(P)H:quinone oxidoreductase 1 (NQO1) is an antioxidant enzyme that has been involved in the progression of several kidney injuries. However, the roles of NQO1 in DN are still unclear. We investigated the effects of NQO1 deficiency in streptozotocin (STZ)-induced DN mice. NQO1 was upregulated in the glomerulus and podocytes under hyperglycemic conditions. NQO1 knockout (NKO) mice showed more severe changes in blood glucose and body weight than WT mice after STZ treatment. Furthermore, STZ-mediated pathological parameters including glomerular injury, blood urea nitrogen levels, and foot process width were more severe in NKO mice than WT mice. Importantly, urine albumin-to-creatinine ratio (ACR) was higher in healthy, non-treated NKO mice than WT mice. ACR response to STZ or LPS was dramatically increased in the urine of NKO mice compared to vehicle controls, while it maintained a normal range following treatment of WT mice. More importantly, we found that NQO1 can stimulate actin polymerization in an in vitro biochemical assay without directly the accumulation on F-actin. In summary, NQO1 has an important role against the development of DN pathogenesis and is a novel contributor in actin reorganization via stimulating actin polymerization.

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Glomerular permselectivity: barrier function based on discrimination of molecular size and charge

AJP Renal Physiology ◽

10.1152/ajprenal.1978.234.6.f455 ◽

1978 ◽

Vol 234 (6) ◽

pp. F455-F460 ◽

Cited By ~ 21

Author(s):

B. M. Brenner ◽

T. H. Hostetter ◽

H. D. Humes

Keyword(s):

Molecular Size ◽

Glomerular Injury ◽

Cell Dysfunction ◽

Morphological Studies ◽

Molecular Charge ◽

Foot Process ◽

Charged Macromolecules ◽

A Charge ◽

Urinary Space ◽

Positively Charged

The formation of glomerular ultrafiltrate is dependent on the interplay of glomerular pressures and flows as well as the intrinsic permselectivity properties of the glomerular capillary wall. These intrinsic permeability properties serve to exclude macromolecules from the urinary space, based on size as well as net molecular charge discrimination. Neutral dextrans with molecular radii less than 20 A cross the glomerular wall without measurable restriction, whereas dextrans with radii greater than 42 A are almost completely barred. For any given size, negatively charged macromolecules are restricted to a greater extent than neutral molecules. Additionally, positively charged molecules are enhanced in their ability to cross the glomerular wall compared to similarly sized neutral polymers. The concept of a charge barrier, due to fixed negative charges within the glomerular wall, is also supported by morphological studies. Glomerular injury, leading to proteinuria, has been associated with loss of the charge-selective properties of these capillaries. Loss of glomerular fixed negative charges may also result in the foot process fusion and mesangial cell dysfunction often observed in proteinuric states.

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P0143OPPOSING EFFECTS OF P38 MAPK IN PODOCYTES ON ALDOSTERONE-INDUCED GLOMERULAR INJURY IN POD-GC-A CKO MICE

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfaa142.p0143 ◽

2020 ◽

Vol 35 (Supplement_3) ◽

Author(s):

SAYAKA SUGIOKA ◽

Yukiko Kato ◽

Akira Ishii ◽

Keita Mori ◽

Keisuke Osaki ◽

...

Keyword(s):

P38 Mapk ◽

Renal Injury ◽

High Salt ◽

Glomerular Injury ◽

Podocyte Damage ◽

Foot Process ◽

Genetic Deletion ◽

P38 Mapk Inhibitor ◽

Mapk Inhibitor

Abstract Background and Aims Previously, we demonstrated that uninephrectomized aldosterone-infused, high salt-fed podocyte-specific guanylyl cyclase-A (natriuretic peptide receptor 1) conditional KO (pod-GC-A cKO) mice exhibited glomerular injury and that pharmacological inhibition of p38 MAPK ameliorates podocyte damage. However, the effects of genetic deletion of p38 MAPK in podocytes of pod-GC-A cKO mice have been unclarified. Method We generated p38 MAPK(fl/fl);Nephrin-Cre (pod-p38 MAPK cKO) mice and p38 MAPK(fl/fl);GC-A(fl/fl);Nephrin-Cre (pod-p38MAPK/GC-A DKO) mice. For induction of glomerular injury, we treated them with aldosterone and high salt at 2 months of age for 3 weeks without nephrectomy (B-ALDO). In vitro, we examined the effect of p38 MAPK inhibitor in cultured human podocytes transfected with GC-A siRNA. Results B-ALDO-treated pod-p38 MAPK/GC-A DKO mice resulted in significant elevation of serum Cr (0.29 ± 0.04 mg/dl), massive albuminuria (42,660 ± 20,200 μg/mgCr) and severe foot process effacement in addition to intracapillary fibrin thrombi which indicated endothelial damage. Vehicle-treated DKO mice, B-ALDO-treated pod-GC-A cKO mice, and B-ALDO-treated pod-p38 MAPK cKO showed normal serum Cr levels (0.14 ± 0.01, 0.18 ± 0.02, 0.20 ± 0.01 mg/dl, respectively), mild increase of albuminuria (223 ± 6.5, 1,496 ± 592, 649 ± 303 μg/mgCr, respectively) and only segmental foot process effacement. Blood pressure was not elevated in either mutant mice compared with that of B-ALDO control mice. Furthermore, glomerular mRNA expressions of MCP-1, PAI-1, and FN were upregulated and that of VEGF-A was downregulated in DKO mice. In vitro, suppression of GC-A mRNA by siRNA in combination with p38 MAPK inhibitor downregulated VEGF mRNA in human cultured podocytes. Our previous works showed that pharmacological inhibition of p38 MAPK in the whole body ameliorated podocyte damage, whereas our current result showed that genetic deletion of p38 MAPK in podocytes aggravated renal injury. In order to explain the discrepancy in these results, we added an analysis of podocyte specific GC-A fl/fl p38 fl/+ cKO mice. Pod GC-A fl/fl p38 fl/+ cKO mice exhibited considerably milder renal damage than pod GC-A fl/fl p38 fl/fl double cKO mice. Conclusion Genetic complete p38 MAPK deletion in GC-A-nul podocytes exacerbated aldosterone-induced glomerular endothelial cell injury as well as podocytes, and resulted in renal dysfunction, probably through VEGF downregulation, whereas partial p38 MAPK inhibition in podocytes ameliorated aldosterone-induced glomerular injury in pod-GC-A cKO mice. These results suggest a certain level of p38 MAPK in podocytes is necessary to protect endothelial and epithelial cells from aldosterone-induced renal injury.

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Deletion of inositol-requiring enzyme-1α in podocytes disrupts glomerular capillary integrity and autophagy

Molecular Biology of the Cell ◽

10.1091/mbc.e16-12-0828 ◽

2017 ◽

Vol 28 (12) ◽

pp. 1636-1651 ◽

Cited By ~ 12

Author(s):

Daniel Robert Kaufman ◽

Joan Papillon ◽

Louise Larose ◽

Takao Iwawaki ◽

Andrey V. Cybulsky

Keyword(s):

Functional Role ◽

Plasma Membranes ◽

Normal Development ◽

Dominant Negative ◽

Male Mice ◽

Glomerular Injury ◽

Cross Sectional ◽

Foot Process ◽

Essential Function

Inositol-requiring enzyme-1α (IRE1α) is an endoplasmic reticulum (ER)–transmembrane endoribonuclease kinase that plays an essential function in extraembryonic tissues during normal development and is activated during ER stress. To address the functional role of IRE1α in glomerular podocytes, we produced podocyte-specific IRE1α-deletion mice. In male mice, deletion of IRE1α in podocytes resulted in albuminuria beginning at 5 mo of age and worsening with time. Electron microscopy revealed focal podocyte foot-process effacement in 9-mo-old male IRE1α-deletion mice, as well as microvillous transformation of podocyte plasma membranes. Compared with control, glomerular cross-sectional and capillary lumenal areas were greater in deletion mice, and there was relative podocyte depletion. Levels of microtubule-associated protein 1A/1B-light chain 3 (LC3)-II expression and c-Jun N-terminal kinase-1 phosphorylation were decreased in IRE1α-deletion glomeruli, in keeping with reduced autophagy. Deletion of IRE1α exacerbated glomerular injury in anti–glomerular basement membrane nephritis. In cell culture, IRE1α dominant-negative mutants reduced the physiological (basal) accumulation of LC3B-II and the size of autophagic vacuoles but did not affect ER-associated degradation. Thus IRE1α is essential for maintaining podocyte and glomerular integrity as mice age and in glomerulonephritis. The mechanism is related, at least in part, to the maintenance of autophagy in podocytes.

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AMP-Kinase mediates regulation of glomerular volume and podocyte survival

10.1101/2021.05.21.445180 ◽

2021 ◽

Author(s):

Khadija Banu ◽

Qisheng Lin ◽

John M Basgen ◽

Marina Planoutene ◽

Chengguo Wei ◽

...

Keyword(s):

Protective Role ◽

Negative Regulator ◽

Amp Kinase ◽

Glomerular Injury ◽

Podocyte Injury ◽

Kinase Activation ◽

Glomerular Volume ◽

Foot Process ◽

Injury Models

We reported that Shroom3 knockdown, via Fyn inhibition, induced albuminuria with foot process effacement (FPE) without glomerulosclerosis (FSGS) or podocytopenia. Interestingly, knockdown mice had reduced podocyte volumes. Human minimal change disease, where podocyte Fyn inactivation was reported, also showed lower glomerular volumes than FSGS. We hypothesized that lower glomerular volume prevented the progression to podocytopenia. To test this hypothesis, we utilized unilateral- and 5/6th nephrectomy models in Shroom3 knockdown mice. Knockdown mice exhibited lower glomerular volume, and less glomerular and podocyte hypertrophy after nephrectomy. FYN-knockdown podocytes had similar reductions in podocyte volume, implying Fyn was downstream of Shroom3. Using SHROOM3- or FYN-knockdown, we confirmed reduced podocyte protein content, along with significantly increased phosphorylated AMP-kinase, a negative regulator of anabolism. AMP-Kinase activation resulted from increased cytoplasmic redistribution of LKB1 in podocytes. Inhibition of AMP-Kinase abolished the reduction in glomerular volume and induced podocytopenia in mice with FPE, suggesting a protective role for AMP-Kinase activation. In agreement with this, treatment of glomerular injury models with AMP-Kinase activators restricted glomerular volume, podocytopenia and progression to FSGS. In summary, we demonstrate the important role of AMP-Kinase in glomerular volume regulation and podocyte survival. Our data suggest that AMP-Kinase activation adaptively regulates glomerular volume to prevent podocytopenia in the context of podocyte injury.

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Binucleate Spermiogenesis in the Mouse

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010009381x ◽

1972 ◽

Vol 30 ◽

pp. 112-113

Author(s):

John J. Wolosewick

Keyword(s):

Electron Microscopy ◽

Phosphate Buffer ◽

Seminiferous Epithelium ◽

Mouse Testis ◽

Germinal Epithelium ◽

Intercellular Bridges ◽

Adult Mice ◽

Cervical Dislocation ◽

Human And Mouse

Classically, the male germinal epithelium is depicted as synchronously developing uninucleate spermatids conjoined by intercellular bridges. Recently, binucleate and multinucleate spermatids from human and mouse testis have been reported. The present paper describes certain developmental events in one type of binucleate spermatid in the seminiferous epithelium of the mouse.Testes of adult mice (ABP Jax) were removed from the animals after cervical dislocation and placed into 2.5% glutaraldehyde/Millonig's phosphate buffer (pH 7.2). Testicular capsules were gently split and separated, exposing the tubules. After 15 minutes the tissue was carefully cut into cubes (approx. 1mm), fixed for an additional 45 minutes and processed for electron microscopy.

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Ineffective Thrombopoiesis

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100158881 ◽

1990 ◽

Vol 48 (3) ◽

pp. 266-267

Author(s):

JM Radley ◽

SL Ellis

Keyword(s):

Microscopic Examination ◽

Phosphate Buffer ◽

Primary Myelofibrosis ◽

Transient Increase ◽

Major Difficulty ◽

Adult Mice ◽

Dense Cytoplasm

In effective thrombopoies is has been inferred to occur in several disease sates from considerations of megakaryocyte mass and platelet kinetics. Microscopic examination has demonstrated increased numbers of megakaryocytes, with a typical forms particularly pronounced, in primary myelofibrosis. It has not been documented if megakaryocyte ever fail to reach maturity in non-pathological situations. A major difficulty of establishing this is that the number of megakaryocytes normally present in the marrow is extremely low. A large transient increase in megakaryocytopoiesis can how ever be induced in mice by an injection of 5-fluorouracil. We have utilised this treatment and report here evidence for in effective thrombopoies is in healthy mice.Adult mice were perfused (2% glutaraldehyde in 0.08M phosphate buffer, pH 7.4) 8 days following an injection of 5-fluorouracil (150mg/kg). Femurs were subsequently decalcified in 10% neutral E.D.T.A. and embedded in Spurrs resin. Transverse sections of marrow revealed many megakaryocytes at various stages of maturity. Occasional megakaryocytes (less than 1%) were found to be under going degeneration prior to achieving full maturation and releasing cytoplasm as platelets. These cells were characterized by a peripheral rim of dense cytoplasm which enveloped a mass of organelles and vacuoles (Fig. 1). Numerous microtubules were foundaround and with in the organelle-rich zone (Fig 2).

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INTRINSIC RENAL CELL DERIVED IL‐12 IS AN EFFECTOR OF RENAL GLOMERULAR INJURY IN CRESCENTIC GLOMERULONEPHRITIS

Nephrology ◽

10.1046/j.1440-1797.2000.abs127.x ◽

2000 ◽

Vol 5 (3) ◽

pp. A98-A98

Author(s):

Timoshanko Jr ◽

Kitching Ar ◽

Holdsworth Sr ◽

Tipping PG.

Keyword(s):

Renal Cell ◽

Crescentic Glomerulonephritis ◽

Glomerular Injury

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