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In the field of sustainable hydrogen production, hematite (α-Fe2O3) is a promising material owing to its optical band gap and water oxidation/reduction energies. In this context, the main objective of this work is devoted to the synthesis of α-Fe2O3 by electrochemical anodization of iron sheet using two anodization voltages (20 and 30 V), followed by annealing at 450°C (1.5 h) in air. Indeed, annealing is a necessary but not sufficient condition for the hematite prepared electrochemically. Furthermore, we discuss the effect of anodizing voltage and analyze its influence on the properties of the films namely structural, morphological, optical, electrochemical and photoelectrochemical (PEC) properties. The annealed Fe2O3 sample, anodized at 30 V, demonstrates a net photocurrent density of 0.81 mA/cm2 at 0.744 VRHE in KOH (1 M). The obtained results indicate that the α-hematite thin films synthesized by anodization technique under optimized conditions is a promising photoanode in PEC cells. [ABSTRACT FROM AUTHOR]

The Pechmann reaction over a series of copper-containing mesoporous silicas with different Cu contents has been investigated. These materials (Cu-HMS-n, where n is the Si/Cu ratio in the precursor gel, n = 15, 25, and 50) have been prepared at room temperature using a route based on hydrogen bonding and self-assembly between neutral primary amine micelles (S°) and neutral inorganic precursors (I°). The obtained materials were analyzed by Chemical analysis, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and nitrogen adsorption to determine the structural order and textural properties. The results show that the copper-containing HMS catalysts exhibit superior catalytic performance in the pechmann condensation reaction (liquid phase condensation of resorcinol and ethyl acetoacetate), the 7-hydroxy-4-methylcoumarin was the only product observed in this reaction. The reaction parameters were optimized to obtain high selectivity of the desired product at maximum resorcinol conversion at a temperature of 150°C. The enhanced catalytic performance of copper-HMS catalysts may be attributed to the homogeneous dispersion and uniformity of the active copper species and to the larger manganese surface areas attained on the HMS support with large pore diameters and surface areas. [ABSTRACT FROM AUTHOR]

Photocatalysis of food dye (tartrazine) was studied using TiO2/ZnAl-LDH material derived from layered-double hydroxide structure, which were synthesized by co-precipitation method. The catalyst was characterized using Fourier-transform infra-red (FTIR) spectroscopy, X-ray diffraction (XRD) spectroscopy, Brunauer–Emmett–Teller (BET) analysis, and scanning electron microscope (SEM), combined with energy dispersive X-ray spectroscopy. The photodegradation experiments were carried out by varying different parameters such as initial dye concentration, pH, irradiation time, and dose photocatalyst. Tartrazine mineralization was also studied by calculating the degradation of chemical oxygen demand. The reusability of TiO2/ZnAl-LDH catalysts was studied and its photocatalytic efficiency was found to be unchanged, even after six cycles of use. The degradation of food dye is achieved at a natural pH 6 and catalyst content of 1 g/L after 120 min under sun with a maximum percentage of degradation 82%. The results of kinetic studies correspond to the Langmuir–Hinshelwood model. [ABSTRACT FROM AUTHOR]

New polymeric membranes were prepared using polymers: polyvinylidene fluoride (PVDF), cellulose triacetate (CTA), and plastified with tris-ethylhexyl phosphate (TEHP). Di (2-ethylhexyl) phosphoric acid (D2EHPA) was selected as carrier. The synthesized membrane was characterized by FTIR, ATG-DSC, and SEM. Mechanical strengths, contact angles as well as physical characteristics were also determined. An attempt was made to separate the metal ions contained in an industrial waste, namely: iron, manganese and zinc. [ABSTRACT FROM AUTHOR]

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Recalcitrant dyes present in effluents constitute a major environmental concern due to their hazardous properties that may cause deleterious effects on aquatic organisms. Tartrazine is a widely-used dye, and it is known to be resistant to biological and chemical degradation processes and by its carcinogenic and mutagenic nature. This study presents the use of TiO 2 (P25) nanoparticles immobilized into a poly(vinylidenefluoride–trifluoroethylene) (P(VDF–TrFE)) membrane to assess the photocatalytic degradation of this dye in a solar photoreactor. The nanocomposite morphological properties were analyzed, confirming an interconnected porous microstructure and the homogeneous distribution of the TiO 2 nanoparticles within the membrane pores. It is shown that the nanocomposite with 8 wt% TiO 2 exhibits a remarkable sunlight photocatalytic activity over five hours, with 78% of the pollutant being degraded. It was also demonstrated that the degradation follows pseudo-first-order kinetics model at low initial tartrazine concentration. Finally, the effective reusability of the produced nanocomposite was also assessed. [ABSTRACT FROM AUTHOR]

The present study explored the possibility of using immobilized ZnO to photodegradation C.I. Acid Yellow 23 (tartrazine) Through this immobilization method, the separation process of fine zinc oxide (ZnO) photocatalyst at the end of the treatment can be avoided.The photocatalytic degradation of both dyes was investigated under sunlight irradiation.The results showed that the percentage of dye removal increased with decreasing initial dye concentration and increasing irradiation time. For the dye solution studied, higher removal efficiency was obtained under acidic solution medium.The findings have shown the potential of this application in dye removal from aqueous solution. [ABSTRACT FROM AUTHOR]

In this study, we report a facile hydrothermal synthesis of strontium-doped SnS nanoflowers that were used as a catalyst for the degradation of antibiotic molecules in water. The prepared sample was characterized using X-ray diffraction (XRD), scanning electron microscopy (SEM), and ultraviolet-visible absorption spectroscopy (UV-Vis). The photocatalytic ability of the strontium-doped SnS nanoflowers was evaluated by studying the degradation of metronidazole in an aqueous solution under photocatalytic conditions. The degradation study was conducted for a reaction period of 300 min at neutral pH, and it was found that the degradation of metronidazole reached 91%, indicating the excellent photocatalytic performance of the catalyst. The influence of experimental parameters such as catalyst dosage, initial metronidazole concentration, initial reaction pH, and light source nature was optimized with respect to metronidazole degradation over time. The reusability of the strontium-doped SnS nanoflowers catalyst was investigated, and its photocatalytic efficiency remained unchanged even after four cycles of use.

The efficacy of the B cell-targeting drug rituximab (RTX) in childhood idiopathic nephrotic syndrome (INS) suggests that B cells may be implicated in disease pathogenesis. However, B cell characterization in children with INS remains limited. Here, using single-cell RNA sequencing, we demonstrate that a B cell transcriptional program poised for effector functions represents the major immune perturbation in blood samples from children with active INS. This transcriptional profile was associated with an extrafollicular B cell response marked by the expansion of atypical B cells (atBCs), marginal zone-like B cells, and antibody-secreting cells (ASCs). Flow cytometry of blood from 13 children with active INS and 24 healthy donors confirmed the presence of an extrafollicular B cell response denoted by the expansion of proliferating RTX-sensitive extrafollicular (CXCR5 - ) CD21 low T-bet + CD11c + atBCs and short-lived T-bet + ASCs in INS. Together, our study provides evidence for an extrafollicular origin for humoral immunity in active INS., (© 2023. The Author(s).)

Over the last decades, the growing contamination of wastewater, mainly caused by industrial processes, improper sewage, natural calamities, and a variety of anthropogenic activities, has caused an increase in water-borne diseases. Notably, industrial applications require careful consideration as they pose significant threats to human health and ecosystem biodiversity due to the production of persistent and complex contaminants. The present work reports on the development, characterization, and application of a poly (vinylidene fluoride-hexafluoropropylene) (PVDF-HFP) porous membrane for the remediation of a wide range of contaminants from wastewater withdrawn from industrial applications. The PVDF-HFP membrane showed a micrometric porous structure with thermal, chemical, and mechanical stability and a hydrophobic nature, leading to high permeability. The prepared membranes exhibited simultaneous activity on the removal of organic matter (total suspended and dissolved solids, TSS, and TDS, respectively), the mitigation of salinity in 50%, and the effective removal of some inorganic anions and heavy metals, achieving efficiencies around 60% for nickel, cadmium, and lead. The membrane proved to be a suitable approach for wastewater treatment, as it showed potential for the simultaneous remediation of a wide range of contaminants. Thus, the as-prepared PVDF-HFP membrane and the designed membrane reactor represent an efficient, straightforward, and low-cost alternative as a pretreatment step for continuous treatment processes for simultaneous organic and inorganic contaminants' remediation in real industrial effluent sources.

Rho GTPases are regulators of the actin cytoskeleton and their activity is modulated by GTPase-activating proteins (GAPs) and guanine nucleotide exchanging factors (GEFs). Glomerular podocytes have numerous actin-based projections called foot processes and their alteration is characteristic of proteinuric kidney diseases. We reported previously that Rac1 hyperactivation in podocytes causes proteinuria and glomerulosclerosis in mice. However, which GAP and GEF modulate Rac1 activity in podocytes remains unknown. Here, using a proximity-based ligation assay, we identified CdGAP (ARHGAP31) and β-PIX (ARHGEF7) as the major regulatory proteins interacting with Rac1 in human podocytes. CdGAP interacted with β-PIX through its basic region, and upon EGF stimulation, they both translocated to the plasma membrane in podocytes. CdGAP-depleted podocytes had altered cell motility and increased basal Rac1 and Cdc42 activities. When stimulated with EGF, CdGAP-depleted podocytes showed impaired β-PIX membrane-translocation and tyrosine phosphorylation, and reduced activities of Src kinase, focal adhesion kinase, and paxillin. Systemic and podocyte-specific CdGAP-knockout mice developed mild but significant proteinuria, which was exacerbated by Adriamycin. Collectively, these findings show that CdGAP contributes to maintain podocyte function and protect them from injury., (© 2022. The Author(s).)

The presence of contaminants of emerging concern (CEC), such as pharmaceuticals, in water sources is one of the main concerns nowadays due to their hazardous properties causing severe effects on human health and ecosystem biodiversity. Niflumic acid (NFA) is a widely used anti-inflammatory drug, and it is known for its non-biodegradability and resistance to chemical and biological degradation processes. In this work, a 10 wt.% TiO 2 /PVDF-TrFE nanocomposite membrane (NCM) was prepared by the solvent casting technique, fully characterized, and implemented on an up-scaled photocatalytic membrane reactor (PMR). The photocatalytic activity of the NCM was evaluated on NFA degradation under different experimental conditions, including NFA concentration, pH of the media, irradiation time and intensity. The NCM demonstrated a remarkable photocatalytic efficiency on NFA degradation, as efficiency of 91% was achieved after 6 h under solar irradiation at neutral pH. The NCM proved effective in long-term use, with maximum efficiency losses of 7%. An artificial neural network (ANN) model was designed to model NFA's photocatalytic degradation behavior, demonstrating a good agreement between experimental and predicted data, with an R 2 of 0.98. The relative significance of each experimental condition was evaluated, and the irradiation time proved to be the most significant parameter affecting the NFA degradation efficiency. The designed ANN model provides a reliable framework l for modeling the photocatalytic activity of TiO 2 /PVDF-TrFE and related NCM.

Two significant limitations of using TiO 2 nanoparticles for water treatment applications are reduced photocatalytic activity under visible radiation and difficulty recovering the particles after use. In this study, round-shaped Ag@TiO 2 nanocomposites with a ≈21 nm diameter and a bandgap energy of 2.8 eV were synthesised by a deposition-precipitation method. These nanocomposites were immobilised into a porous poly (vinylidene fluoride-hexafluoropropylene) (PVDF-HFP) matrix and well-distributed within the pores. The photocatalytic activity of Ag@TiO 2 /PVDF-HFP against metronidazole (MNZ) under solar radiation was evaluated. Further, an adaptive neuro-fuzzy inference system (ANFIS) was applied to predict the effect of four independent variables, including initial pollutant concentration, pH, light irradiation intensity, and reaction time, on the photocatalytic performance of the composite membrane on MNZ degradation. The 10% Ag@TiO 2 /PVDF-HFP composite membrane showed a maximum removal efficiency of 100% after 5 h under solar radiation. After three use cycles, this efficiency remained practically constant, demonstrating the membranes' reusability and suitability for water remediation applications.

In this work, the synthesis of pure and (Ce, Ag) co-doped ZnO was successfully accomplished using a solvothermal process. The synthesized samples were characterized by ultraviolet-visible spectroscopy, X-ray diffraction, and scanning electron microscopy. The photocatalytic ability of the samples is estimated through degradation of tartrazine in aqueous solution under photocatalytic conditions. The degradation study carried out for a reaction period of 90 min at and a free pH = 6.0 found that dye degradation is 44.82% for pure ZnO and 98.91% for (Ce, Ag) co-doped ZnO samples, indicating its excellent photocatalytic ability. Tartrazine mineralization was also studied by calculating the degradation of chemical oxygen demand. The effect of operating parameters such as catalyst dose, initial concentration of tartrazine, initial reaction pH, and nature of light source has been optimized for tartrazine degradation as a function of time. The reusability of ZnO and (Ce, Ag) co-doped ZnO catalysts was studied and its photocatalytic efficiency was found to be unchanged, even after six cycles of use. The mechanism of photocatalytic activity was also proposed.

This study focuses on the synthesis of various nanocomposites with heterojunction structures, MgAl-LDH (LDH = layered double hydroxides) hybrid with semiconductor such as MoO 3 and CuO. These solids were synthesized by co-precipitation method at constant pH and have been characterized extensively using atomic absorption spectroscopy (AAS), X-ray diffraction (XRD), Fourier transform infrared (FTIR) and transmission electron microscopy-energy dispersive X-ray (TEM-EDX) methods. The catalytic activity of nanocomposites was tested in the photocatalytic degradation under solar irradiation of emerging pollutants as the pharmaceutical metronidazole (MNZ). The experimental parameters, including initial MNZ concentration, the nature of oxide incorporate in the photocatalyst, catalyst loading were explored. All the synthesized samples showed high photocatalytic performances; the highest photocatalysis efficiency was achieved with the photocatalyst dose 1.5 g/L and initial MNZ concentration of 10 mg/L at neutral pH. The photocatalytic experimental results were fitted very well to the Langmuir-Hinshelwood model. From the obtained results the calcined LDH/semiconductors could be efficient for the photocatalytic process under solar irradiation of pharmaceuticals and may contribute in environmental remediation.

Background: Previous studies showed that Cdc42, a member of the prototypical Rho family of small GTPases and a regulator of the actin cytoskeleton, is critical for the normal development and health of podocytes. However, upstream regulatory mechanisms for Cdc42 activity in podocytes are largely unknown., Methods: We used a proximity-based ligation assay, BioID, to identify guanine nucleotide exchange factors that activate Cdc42 in immortalized human podocytes. We generated podocyte-specific ARHGEF7 (commonly known as β -PIX) knockout mice by crossing β -PIX floxed mice with Podocin-Cre mice. Using shRNA, we established cultured mouse podocytes with β -PIX knockdown and their controls., Results: We identified β -PIX as a predominant guanine nucleotide exchange factor that interacts with Cdc42 in human podocytes. Podocyte-specific β -PIX knockout mice developed progressive proteinuria and kidney failure with global or segmental glomerulosclerosis in adulthood. Glomerular podocyte density gradually decreased in podocyte-specific β -PIX knockout mice, indicating podocyte loss. Compared with controls, glomeruli from podocyte-specific β -PIX knockout mice and cultured mouse podocytes with β -PIX knockdown exhibited significant reduction in Cdc42 activity. Loss of β -PIX promoted podocyte apoptosis, which was mediated by the reduced activity of the prosurvival transcriptional regulator Yes-associated protein., Conclusions: These findings indicate that β -PIX is required for the maintenance of podocyte architecture and glomerular function via Cdc42 and its downstream Yes-associated protein activities. This appears to be the first evidence that a Rho-guanine nucleotide exchange factor plays a critical role in podocytes., (Copyright © 2020 by the American Society of Nephrology.)

The compounds found in industrial wastewater typically show high toxicity, and in this way, they have become a primary environmental concern. Several techniques have been applied in industrial effluent remediation. In spite of the efforts, these techniques are yet to be ineffective to treat oily wastewater before it can be discharged safely to the environment. Membrane technology is an attractive approach to treat oily wastewater. This is dedicated to the immobilisation of TiO 2 nanoparticles on poly(vinylidene fluoride-trifluoro ethylene) (PVDF-TrFE) porous matrix by solvent casting. Membranes with interconnected pores with an average diameter of 60 µm and a contact angle of 97°, decorated with TiO 2 nanoparticles, are obtained. The degradation of oily wastewater demonstrated the high photocatalytic efficiency of the nanocomposite membranes: Under sunlight irradiation for seven hours, colourless water was obtained.

The Houbara bustard Chlamydotisundulata (Jacquin, 1784) is an emblematic and endangered bird of steppes and desert spaces of North Africa. This species belonging to Otidiformes is recognized as vulnerable by the International Union for Nature Conservation. The critical situation of this species and the revision of its classification on the tree of birds encouraged the authors to start accumulating chromosome data. For that, we propose the GTG- and RBG-banded karyotypes of the Houbara bustard prepared from primary fibroblast cell cultures. The first eight autosomal pairs and sex chromosomes have been described and compared to those of the domestic fowl Gallusdomesticus (Linnaeus, 1758). The diploid number has been estimated as 78 chromosomes with 8 macrochromosomes pairs and 30 microchromosomes pairs, attesting of the stability of chromosome number in avian karyotypes. The description of the karyotype of the Houbara is of crucial importance for the management of the reproduction of this species in captivity. It can be used as a reference in the detection of chromosomal abnormalities, which would be responsible of the early embryonic mortalities.

Although sequence variants in CD2-associated protein (CD2AP) have been identified in patients with focal segmental glomerulosclerosis (FSGS), definitive proof of causality in human disease is meager. By whole-exome sequencing, we identified a homozygous frame-shift mutation in CD2AP (p.S198fs) in three siblings born of consanguineous parents who developed childhood-onset FSGS and end stage renal disease. When the same frameshift mutation was introduced in mice by gene editing, the mice developed FSGS and kidney failure. These results provide conclusive evidence that homozygous mutation of CD2AP causes FSGS in humans., (Copyright © 2018 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Nephrotic syndrome is a kidney disease featured by heavy proteinuria. It is caused by injury to the specialized epithelial cells called "podocytes" within the filtration unit of the kidney, glomerulus. Previous studies showed that hyperactivation of the RhoGTPase, Rac1, in podocytes causes podocyte injury and glomerulosclerosis (accumulation of extracellular matrix in the glomerulus). However, the mechanism by which Rac1 is activated during podocyte injury is unknown. Trio is a guanine nucleotide exchange factor (GEF) known to activate Rac1. By RNA-sequencing, we found that Trio mRNA is abundantly expressed in cultured human podocytes. Trio mRNA was also significantly upregulated in humans with minimal change disease and focal segmental glomerulosclerosis, two representative causes of nephrotic syndrome. Reduced expression of Trio in cultured human podocytes decreased basal Rac1 activity, cell size, attachment to laminin, and motility. Furthermore, while the pro-fibrotic cytokine, transforming growth factor β1 increased Rac1 activity in control cells, it decreases Rac1 activity in cells with reduced Trio expression. This was likely due to simultaneous activation of the Rac1-GTPase activation protein, CdGAP. Thus, Trio is important in the basal functions of podocytes and may also contribute to glomerular pathology, such as sclerosis, via Rac1 activation., Competing Interests: The authors declare no conflict of interest.

Nephrin is a key structural component of the podocyte slit diaphragm, and proper expression of nephrin on the cell surface is critical to ensure integrity of the blood filtration barrier. Maintenance of nephrin within this unique cell junction has been proposed to require dynamic phosphorylation events and endocytic recycling, although the molecular mechanisms that control this interplay are poorly understood. Here, we investigated the possibility that the phosphotyrosine adaptor protein ShcA regulates nephrin turnover. Western blotting and immunostaining analysis confirmed that ShcA is expressed in podocytes. In immunoprecipitation and pulldown assays, ShcA, via its SH2 domain, was associated with several phosphorylated tyrosine residues on nephrin. Overexpression of ShcA promoted nephrin tyrosine phosphorylation and reduced nephrin signaling and cell surface expression in vitro In a rat model of reversible podocyte injury and proteinuria, phosphorylated nephrin temporally colocalized with endocytic structures coincident with upregulation of ShcA expression. In vivo biotinylation assays confirmed that nephrin expression decreased at the cell surface and correspondingly increased in the cytosol during the injury time course. Finally, immunostaining in kidney biopsy specimens demonstrated overexpression of ShcA in several human proteinuric kidney diseases compared with normal conditions. Our results suggest that increases in ShcA perturb nephrin phosphosignaling dynamics, leading to aberrant nephrin turnover and slit diaphragm disassembly., (Copyright © 2018 by the American Society of Nephrology.)

Hyper-activation of Rac1, a small GTPase, in glomerular podocytes has been implicated in the pathogenesis of familial proteinuric kidney diseases. However, the role of Rac1 in acquired nephrotic syndrome is unknown. To gain direct insights into this, we generated a transgenic mouse model expressing a doxycycline-inducible constitutively active form of Rac1 (CA-Rac1) in podocytes. Regardless of the copy number, proteinuria occurred rapidly within five days, and the histology resembled minimal change disease. The degree and severity of proteinuria were dependent on the transgene copy number. Upon doxycycline withdrawal, proteinuria resolved completely (one copy) or nearly completely (two copy). After one month of doxycycline treatment, two-copy mice developed glomerulosclerosis that resembled focal segmental glomerulosclerosis (FSGS) with urinary shedding of transgene-expressing podocytes. p38 MAPK was activated in podocytes upon CA-Rac1 induction while a p38 inhibitor attenuated proteinuria, podocyte loss, and glomerulosclerosis. Mechanistically, activation of Rac1 in cultured mouse podocytes reduced adhesiveness to laminin and induced redistribution of β1 integrin, and both were partially reversed by the p38 inhibitor. Activation of Rac1 in podocytes was also seen in kidney biopsies from patients with minimal change disease and idiopathic FSGS by immunofluorescence while sera from the same patients activated Rac1 in cultured human podocytes. Thus, activation of Rac1 in podocytes causes a spectrum of disease ranging from minimal change disease to FSGS, due to podocyte detachment from the glomerular basement membrane that is partially dependent on p38 MAPK., (Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Glomerular visceral epithelial cells (podocytes) play a critical role in the maintenance of glomerular permselectivity. Podocyte injury, manifesting as proteinuria, is the cause of many glomerular diseases. We reported previously that calcium-independent phospholipase A2γ (iPLA2γ) is cytoprotective against complement-mediated glomerular epithelial cell injury. Studies in iPLA2γ KO mice have demonstrated an important role for iPLA2γ in mitochondrial lipid turnover, membrane structure, and metabolism. The aim of the present study was to employ iPLA2γ KO mice to better understand the role of iPLA2γ in normal glomerular and podocyte function as well as in glomerular injury. We show that deletion of iPLA2γ did not cause detectable albuminuria; however, it resulted in mitochondrial structural abnormalities and enhanced autophagy in podocytes as well as loss of podocytes in aging KO mice. Moreover, after induction of anti-glomerular basement membrane nephritis in young mice, iPLA2γ KO mice exhibited significantly increased levels of albuminuria, podocyte injury, and loss of podocytes compared with wild type. Thus, iPLA2γ has a protective functional role in the normal glomerulus and in glomerulonephritis. Understanding the role of iPLA2γ in glomerular pathophysiology provides opportunities for the development of novel therapeutic approaches to glomerular injury and proteinuria., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

Nephrotic syndrome (NS) describes a group of kidney disorders in which there is injury to podocyte cells, specialized cells within the kidney's glomerular filtration barrier, allowing proteins to leak into the urine. Three mutations in ARHGDIA, which encodes Rho GDP dissociation inhibitor α (GDIα), have been reported in patients with heritable NS and encode the following amino acid changes: ΔD185, R120X, and G173V. To investigate the impact of these mutations on podocyte function, endogenous GDIα was knocked-down in cultured podocytes by shRNA and then the cells were re-transfected with wild-type or mutant GDIα constructs. Among the 3 prototypical Rho-GTPases, Rac1 was markedly hyperactivated in podocytes with any of the 3 mutant forms of GDIα while the activation of RhoA and Cdc42 was modest and variable. All three mutant GDIα proteins resulted in slow podocyte motility, suggesting that podocytes are sensitive to the relative balance of Rho-GTPase activity. In ΔD185 podocytes, both random and directional movements were impaired and kymograph analysis of the leading edge showed increased protrusion and retraction of leading edge (phase switching). The mutant podocytes also showed impaired actin polymerization, smaller cell size, and increased cellular projections. In the developing kidney, GDIα expression increased as podocytes matured. Conversely, active Rac1 was detected only in immature, but not in mature, podocytes. The results indicate that GDIα has a critical role in suppressing Rac1 activity in mature podocytes, to prevent podocyte injury and nephrotic syndrome.

Rho family GTPases act as molecular switches regulating actin cytoskeleton dynamics. Attenuation of their signaling capacity is provided by GTPase-activating proteins (GAPs), including p190A, that promote the intrinsic GTPase activity of Rho proteins. In the current study we have performed a small-scale ENU mutagenesis screen and identified a novel loss of function allele of the p190A gene Arhgap35, which introduces a Leu1396 to Gln substitution in the GAP domain. This results in decreased GAP activity for the prototypical Rho-family members, RhoA and Rac1, likely due to disrupted ordering of the Rho binding surface. Consequently, Arhgap35-deficient animals exhibit hypoplastic and glomerulocystic kidneys. Investigation into the cystic phenotype shows that p190A is required for appropriate primary cilium formation in renal nephrons. P190A specifically localizes to the base of the cilia to permit axoneme elongation, which requires a functional GAP domain. Pharmacological manipulations further reveal that inhibition of either Rho kinase (ROCK) or F-actin polymerization is able to rescue the ciliogenesis defects observed upon loss of p190A activity. We propose a model in which p190A acts as a modulator of Rho GTPases in a localized area around the cilia to permit the dynamic actin rearrangement required for cilia elongation. Together, our results establish an unexpected link between Rho GTPase regulation, ciliogenesis and glomerulocystic kidney disease.

Nephrotic syndrome is a disease of glomerular permselectivity that can arise as a consequence of heritable or acquired changes to the integrity of the glomerular filtration barrier. We recently reported two siblings with heritable nephrotic syndrome caused by a loss of function mutation in the gene ARHGDIA, which encodes for Rho guanine nucleotide dissociation inhibitor-α (GDIα). GDIs are known to negatively regulate Rho-GTPase signaling. We hypothesized that loss of GDIα sensitizes podocytes to external injury via hyperactivation of Rho-GTPases and p38 MAPK. We examined the response of cultured podocytes with and without knockdown of GDIα to LPS injury by assessing the levels of phospho-p38 as well as the degree of synaptopodin loss. GDIα knockdown podocytes showed more pronounced and sustained p38 phosphorylation in response to LPS compared with control podocytes, and this was blunted significantly by the Rac1 inhibitor. In LPS-treated control podocytes, synaptopodin degradation occurred, and this was dependent on p38, the proteasome, and cathepsin L. In GDIα knockdown podocytes, the same events were triggered, but the levels of synaptopodin after LPS treatment were significantly lower than in control podocytes. These experiments reveal a common pathway by which heritable and environmental risk factors converge to injure podocytes, from Rac1 hyperactivation to p38 phosphorylation and synaptopodin degradation via the ubiquitin-proteasome pathway and cathepsin L., (Copyright © 2015 the American Physiological Society.)

Protein tyrosine phosphatase 1B (PTP1B) is a ubiquitously expressed nonreceptor protein-tyrosine phosphatase that regulates various cellular functions, including migration. Recent studies suggest that an increased migratory phenotype of podocytes may be responsible for proteinuria and foot process effacement. The current study addresses the role of PTP1B in podocyte injury and proteinuria. PTP1B was markedly up-regulated in the glomerulus, notably in podocytes, in three rodent models of podocyte injury. Podocyte-specific ablation of the PTP1B gene ameliorated proteinuria induced by lipopolysaccharide and Adriamycin (doxorubicin). The use of a specific PTP1B inhibitor also protected against lipopolysaccharide-induced proteinuria. In contrast, podocyte-specific PTP1B transgenic male mice developed spontaneous proteinuria and foot process effacement. In cultured mouse podocytes, PTP1B knockdown and/or pretreatment with the PTP1B inhibitor blunted lipopolysaccharide-induced cell migration, activation of Src-family kinases (SFKs), and phosphorylation of focal adhesion kinase at Y397 (pFAK(Y397)), the latter being crucial for cell migration. Lipopolysaccharide-injected mice showed increased glomerular expression of active SFKs and pFAK(Y397), both of which were inhibited by podocyte-specific PTP1B knockout and the PTP1B inhibitor. Moreover, podocyte-specific PTP1B transgenic mice showed increased glomerular expression of active SFKs and pFAK(Y397). In summary, PTP1B up-regulation in podocytes induces a migratory response by activating SFKs and FAK, leading to foot process effacement and proteinuria. Pharmacological inhibition of PTP1B may have therapeutic potential in the treatment of proteinuric diseases., (Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Visceral glomerular epithelial cells (GEC), also known as podocytes, are vital for the structural and functional integrity of the glomerulus. The actin cytoskeleton plays a central role in maintaining GEC morphology. In a rat model of experimental membranous nephropathy (passive Heymann nephritis (PHN)), complement C5b-9-induced proteinuria was associated with the activation of the actin regulator small GTPase, RhoA. The mechanisms of RhoA activation, however, remained unknown. In this study, we explored the role of the epithelial guanine nucleotide exchange factor, GEF-H1, in complement-induced RhoA activation. Using affinity precipitation to monitor GEF activity, we found that GEF-H1 was activated in glomeruli isolated from rats with PHN. Complement C5b-9 also induced parallel activation of GEF-H1 and RhoA in cultured GEC. In GEC in which GEF-H1 was knocked down, both basal and complement-induced RhoA activity was reduced. On the other hand, GEF-H1 knockdown augmented complement-mediated cytolysis, suggesting a role for GEF-H1 and RhoA in protecting GEC from cell death. The MEK1/2 inhibitor, U0126, and mutation of the ERK-dependent phosphorylation site (T678A) prevented complement-induced GEF-H1 activation, indicating a role for the ERK pathway. Further, complement induced GEF-H1 and microtubule accumulation in the perinuclear region. However, both the perinuclear accumulation and the activation of GEF-H1 were independent of microtubules and myosin-mediated contractility, as shown using drugs that interfere with microtubule dynamics and myosin II activity. In summary, we have identified complement-induced ERK-dependent GEF-H1 activation as the upstream mechanism of RhoA stimulation, and this pathway has a protective role against cell death.

The noncanonical Wnt/planar cell polarity (PCP) pathway controls a variety of cell behaviors such as polarized protrusive cell activity, directional cell movement, and oriented cell division and is crucial for the normal development of many tissues. Mutations in the PCP genes cause malformation in multiple organs. Recently, the PCP pathway was shown to control endocytosis of PCP and non-PCP proteins necessary for cell shape remodeling and formation of specific junctional protein complexes. During formation of the renal glomerulus, the glomerular capillary becomes enveloped by highly specialized epithelial cells, podocytes, that display unique architecture and are connected via specialized cell-cell junctions (slit diaphragms) that restrict passage of protein into the urine; podocyte differentiation requires active remodeling of cytoskeleton and junctional protein complexes. We report here that in cultured human podocytes, activation of the PCP pathway significantly stimulates endocytosis of the core slit diaphragm protein, nephrin, via a clathrin/β-arrestin-dependent endocytic route. In contrast, depletion of the PCP protein Vangl2 leads to an increase of nephrin at the cell surface; loss of Vangl2 functions in Looptail mice results in disturbed glomerular maturation. We propose that the PCP pathway contributes to podocyte development by regulating nephrin turnover during junctional remodeling as the cells differentiate.

Glomerulosclerosis is featured by accumulation of the extracellular matrixes in the glomerulus. We showed previously that activation of the small GTPase RhoA in podocytes induces heavy proteinuria and glomerulosclerosis in the mouse. In the current study, we investigated the mechanism by which RhoA stimulates the production of one of the extracellular matrixes, fibronectin, by podocytes, specifically testing the role of nuclear factor of activated T cells (NFAT). Expression of constitutively active RhoA in cultured podocytes activated the fibronectin promoter, upregulated fibronectin protein, and activated NFAT. Expression of constitutively active NFAT in podocytes also activated the fibronectin promoter and upregulated fibronectin protein. RhoA-induced NFAT activation and fibronectin upregulation were both dependent on the calcium/calmodulin pathway and Rho kinase. NFAT activation was also observed in vivo in the rat and mouse models of podocyte injury and proteinuria, and NFAT inhibition ameliorated fibronectin upregulation in the latter. RhoA activation induced a rise of intracellular calcium ion concentration ([Ca(2+)]i), which was at least in part dependent on the transient receptor potential canonical 6 (TRPC6) cation channel. The results indicate that RhoA activates NFAT by inducing a rise of [Ca(2+)]i in podocytes, which in turn contributes to fibronectin upregulation. This pathway may be responsible for the pathogenesis of certain glomerular diseases such as hypertension-mediated glomerulosclerosis.

Proper organization of the actin cytoskeleton is essential for the normal structure and function of podocytes. RhoA modulates actin dynamics but its role in podocyte biology is controversial. Here, we generated transgenic mice that express a constitutively active form of RhoA in a podocyte-specific and doxycycline-inducible manner. Induction of activated RhoA with doxycycline resulted in significant albuminuria. Furthermore, both the degree of albuminuria and the histologic changes in the glomerulus positively correlated with the level of constitutively active RhoA expression: low levels of expression associated with segmental foot-process effacement without changes observable by light microscopy, whereas higher levels of expression associated with both extensive foot-process effacement and histologic features of focal segmental glomerulosclerosis (FSGS). In addition, induction of activated RhoA markedly upregulated glomerular mRNA expression of fibronectin and collagen IA1, and the degree of upregulation positively correlated with the level of albuminuria. Withdrawal of doxycycline led to a decline in albuminuria toward basal levels in most mice, but heavy albuminuria persisted in some mice. Taken together, these data suggest that activation of RhoA in podocytes leads to albuminuria accompanied by a range of histologic changes characteristic of minimal change disease and FSGS in humans. Although most changes are reversible, severe and prolonged activation of RhoA may cause irreversible glomerulosclerosis.

Glomerular podocytes are critical for the barrier function of the glomerulus in the kidney and their dysfunction causes protein leakage into the urine (proteinuria). Nephrin is a key podocyte protein, which regulates the actin cytoskeleton via tyrosine phosphorylation of its cytoplasmic domain. Here we report that two protein tyrosine phosphatases, PTP1B and PTP-PEST negatively regulate nephrin tyrosine phosphorylation. PTP1B directly binds to and dephosphorylates nephrin, while the action of PTP-PEST is indirect. The two phosphatases are also upregulated in the glomerulus in the rat model of puromycin aminonucleoside nephrosis. Both overexpression and inhibition of PTP1B deranged the actin cytoskeleton in cultured mouse podocytes. Thus, protein tyrosine phosphatases may affect podocyte function via regulating nephrin tyrosine phosphorylation.