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Misfolded protein aggregates may cause toxic proteinopathy, including autosomal dominant tubulointerstitial kidney disease due to uromodulin mutations (ADTKD-UMOD), a leading hereditary kidney disease. There are no targeted therapies. In our generated mouse model recapitulating human ADTKD-UMOD carrying a leading UMOD mutation, we show that autophagy/mitophagy and mitochondrial biogenesis are impaired, leading to cGAS-STING activation and tubular injury. Moreover, we demonstrate that inducible tubular overexpression of mesencephalic astrocyte-derived neurotrophic factor (MANF), a secreted endoplasmic reticulum protein, after the onset of disease stimulates autophagy/mitophagy, clears mutant UMOD, and promotes mitochondrial biogenesis through p-AMPK enhancement, thus protecting kidney function in our ADTKD mouse model. Conversely, genetic ablation of MANF in the mutant thick ascending limb tubular cells worsens autophagy suppression and kidney fibrosis. Together, we have discovered MANF as a biotherapeutic protein and elucidated previously unknown mechanisms of MANF in the regulation of organelle homeostasis, which may have broad therapeutic applications to treat various proteinopathies.
Assuntos
Doenças Renais Policísticas , Humanos , Camundongos , Animais , Autofagia/genética , Homeostase , Fibrose , Fatores de Crescimento Neural/genéticaRESUMO
Misfolded protein aggregates may cause toxic proteinopathy, including autosomal dominant tubulointerstitial kidney disease due to uromodulin mutations (ADTKD- UMOD ), one of the leading hereditary kidney diseases, and Alzheimerâ™s disease etc. There are no targeted therapies. ADTKD is also a genetic form of renal fibrosis and chronic kidney disease, which affects 500 million people worldwide. For the first time, in our newly generated mouse model recapitulating human ADTKD- UMOD carrying a leading UMOD deletion mutation, we show that autophagy/mitophagy and mitochondrial biogenesis are severely impaired, leading to cGAS- STING activation and tubular injury. Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a novel endoplasmic reticulum stress-regulated secreted protein. We provide the first study that inducible tubular overexpression of MANF after the onset of disease stimulates autophagy/mitophagy and clearance of the misfolded UMOD, and promotes mitochondrial biogenesis through p-AMPK enhancement, resulting in protection of kidney function. Conversely, genetic ablation of endogenous MANF upregulated in the mutant mouse and human tubular cells worsens autophagy suppression and kidney fibrosis. Together, we discover MANF as a novel biotherapeutic protein and elucidate previously unknown mechanisms of MANF in regulating organelle homeostasis to treat ADTKD, which may have broad therapeutic application to treat various proteinopathies.
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Ribosomal protein S6 (rpS6) phosphorylation mediates the hypertrophic growth of kidney proximal tubule cells. However, the role of rpS6 phosphorylation in podocyte hypertrophy and podocyte loss during the pathogenesis of focal segmental glomerulosclerosis (FSGS) remains undefined. Here, we examined rpS6 phosphorylation levels in kidney biopsy specimens from patients with FSGS and in podocytes from mouse kidneys with Adriamycin-induced FSGS. Using genetic and pharmacologic approaches in the mouse model of FSGS, we investigated the role of rpS6 phosphorylation in podocyte hypertrophy and loss during development and progression of FSGS. Phosphorylated rpS6 was found to be markedly increased in the podocytes of patients with FSGS and Adriamycin-induced FSGS mice. Genetic deletion of the Tuberous sclerosis 1 gene in kidney glomerular podocytes activated mammalian target of rapamycin complex 1 signaling to rpS6 phosphorylation, resulting in podocyte hypertrophy and pathologic features similar to those of patients with FSGS including podocyte loss, leading to segmental glomerulosclerosis. Since protein phosphatase 1 is known to negatively regulate rpS6 phosphorylation, treatment with an inhibitor increased phospho-rpS6 levels, promoted podocyte hypertrophy and exacerbated formation of FSGS lesions. Importantly, blocking rpS6 phosphorylation (either by generating congenic rpS6 knock-in mice expressing non-phosphorylatable rpS6 or by inhibiting ribosomal protein S6 kinase 1-mediated rpS6 phosphorylation with an inhibitor) significantly blunted podocyte hypertrophy, inhibited podocyte loss, and attenuated formation of FSGS lesions. Thus, our study provides genetic and pharmacologic evidence indicating that specifically targeting rpS6 phosphorylation can attenuate the development of FSGS lesions by inhibiting podocyte hypertrophy and associated podocyte depletion.
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Glomerulosclerose Segmentar e Focal , Podócitos , Animais , Doxorrubicina , Glomerulosclerose Segmentar e Focal/metabolismo , Humanos , Hipertrofia , Mamíferos/metabolismo , Camundongos , Fosforilação , Podócitos/patologia , Proteínas Serina-Treonina Quinases , Proteína S6 Ribossômica/metabolismoRESUMO
Nod-like receptor protein 3 (NLRP3), as an inflammatory regulator, has been implicated in acute kidney injury (AKI). Failed recovery after AKI can lead to chronic kidney disease (CKD). However, the role of NLRP3 in the AKI-CKD transition is still unknown. A mild or severe AKI mouse model was performed by using ischemia-reperfusion injury (IRI). We evaluated the renal NLRP3 expression in acute and chronic phases of ischemic AKI, respectively. Although serum creatinine (Cr) and blood urea nitrogen (BUN) levels in AKI chronic phase were equivalent to normal baseline, histological analysis and fibrotic markers revealed that severe AKI-induced maladaptive tubular repair with immune cell infiltration and fibrosis. Tubular damage was restored completely in mild AKI rather than in severe AKI. Of note, persistent overexpression of NLRP3 was also found in severe AKI but not in mild AKI. In the severe AKI-induced chronic phase, there was a long-term high level of NLRP3 in serum or urine. Overt NLRP3 was mainly distributed in the abnormal tubules surrounded by inflammatory infiltrates and fibrosis, which indicated the maladaptive repair. Renal Nlrp3 overexpression was correlated with infiltrating macrophages and fibrosis. Renal NLRP3 signaling-associated genes were upregulated after severe AKI by RNA-sequencing. Furthermore, NLRP3 was found increased in renal tubular epitheliums from CKD biopsies. Together, persistent NLRP3 overexpression was associated with chronic pathological changes following AKI, which might be a new biomarker for evaluating the possibility of AKI-CKD transition.
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Tuberous sclerosis complex (TSC) is characterized by hamartomatous lesions in multiple organs, with most patients developing polycystic kidney disease and leading to a decline of renal function. TSC is caused by loss-of-function mutations in either Tsc1 or Tsc2 gene, but currently, there is no effective treatment for aberrant kidney growth in TSC patients. By generating a renal proximal tubule-specific Tsc1 gene-knockout (Tsc1 ptKO) mouse model, we observed that Tsc1 ptKO mice developed aberrantly enlarged kidneys primarily due to hypertrophy and proliferation of proximal tubule cells, along with some cystogenesis, interstitial inflammation, and fibrosis. Mechanistic studies revealed inhibition of AMP-activated protein kinase (AMPK) phosphorylation at Thr-172 and activation of Akt phosphorylation at Ser-473 and Thr-308. We therefore treated Tsc1 ptKO mice with the AMPK activator, metformin, by daily intraperitoneal injection. Our results indicated that metformin increased the AMPK phosphorylation, but decreased the Akt phosphorylation. These signaling modulations resulted in inhibition of proliferation and induction of apoptosis in the renal proximal tubule cells of Tsc1 ptKO mice. Importantly, metformin treatment effectively prevented aberrant kidney enlargement and cyst growth, inhibited inflammatory response, attenuated interstitial fibrosis, and protected renal function. The effects of metformin were further confirmed by in vitro experiments. In conclusion, this study indicates a potential therapeutic effect of metformin on Tsc1 deletion-induced kidney pathology, although currently metformin is primarily prescribed to treat patients with type 2 diabetes.
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UCH-L1 is a de-ubiquitination enzyme comprehensively distributed in neural cells and podocytes, which is involved in several kinds of nervous system and kidney related diseases. Our previous studies have demonstrated the aberrant up-regulation of UCH-L1 in podocytes of renal diseases, but how dose podocytes are injured by up-regulated UCH-L1 is waiting to be elucidated. Here, we observed the cytoskeleton rearrangement in podocytes with over-expression of UCH-L1, accompanied with a down-regulation of synaptopodin and RhoA, which are closely related to cytoskeletal stabilization. However, we did not see any alteration of RhoA ubiquitination level under the stimulation of UCH-L1 in podocytes. Subsequently, mass spectrum was applied in UCH-L1-flag immunoprecipitation and plakoglobin was screened out, which was among the UCH-L1-combined proteins and most likely related to cytoskeleton rearrangement. Our experiment demonstrates UCH-L1 may not injure podocytes cytoskeleton through a direct regulation on RhoA/Synaptopodin, but through the regulation of plakoglobin, which could be a promising target for treatment of renal disease in the future.
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Podócitos/metabolismo , Ubiquitina Tiolesterase/metabolismo , Animais , Linhagem Celular , Citoesqueleto/metabolismo , Citoesqueleto/patologia , Humanos , Nefropatias/metabolismo , Nefropatias/patologia , Camundongos , Proteínas dos Microfilamentos/metabolismo , Podócitos/patologia , Ubiquitinação , gama Catenina/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
A collaborative system of carboxymethyl cellulose stabilized nanosized zero-valent iron (CMC-nFe0) and microorganisms was set up to enhance the stabilization of Cr(VI) in soil. In comparison with an aqueous-bound Cr(VI) removal of 18.9% in the nFe0 system, a higher Cr(VI) removal of 68.9% was achieved in the nFe0 and microorganisms system after 14 d remediation because the microorganisms on the nFe0 surface promoted nFe0 corrosion and enhanced abiotic and biotic Cr(VI) stabilization by generating highly active minerals such as magnetite, lepidocrocite and green rust on the nFe0 surface. As a stabilizing agent for nFe0 and an organic substrate for microorganisms, CMC on the nFe0 surface not only enhanced the dispersion of nFe0, but also boosted the activity of microorganisms, resulting in a promotion of 0.9 and 0.5 times higher aqueous-bound Cr(VI) removal via the improvement of nFe0 and microorganisms respectively, thus a total 4 times higher aqueous-bound Cr(VI) removal of 95.3% was achieved in the CMC-nFe0 and microorganisms system as compared to the nFe0 system. After 14 d remediation, easily available species of Cr(VI) and Crtotal, such as water soluble (WS), exchangeable (EX) and bounded to carbonates (CB), were mainly transformed to less available Fe-Mn oxides-bounded (OX) and residual (RS) species because of the production of ferrochrome precipitates (CrxFe1-xOOH or CrxFe1-x(OH)3). Besides, the stabilization of Cr(VI) in the CMC-nFe0 and microorganisms system was pH-dependent and it increased with CMC-nFe0 dosage. Due to excellent Cr(VI) stabilization and Cr immobilization, coupled CMC-nFe0 and anaerobic microorganisms process is of great potential in remediating Cr(VI)-containing soil.
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Solo , Poluentes Químicos da Água , Anaerobiose , Carboximetilcelulose Sódica , CromoRESUMO
SAMHD1 (Sterile Alpha Motif and Histidine-aspartate Domain containing protein 1) has been documented as a host factor that restricts HIV-1 and some DNA viruses. In this work, we attempted to explore possible effects of SAMHD1 on exogenous DNA and show that SAMHD1 exerts a general inhibition on the expression of exogenous DNA in vitro and in mice. This inhibition is achieved through repressing transcription of exogenous DNA. Intriguingly, unlike SAMHD1's restriction of HIV-1, such restriction does not require the dNTPase or RNase activities, or T592 phosphorylation of SAMHD1. Mechanistically, SAMHD1 enhances the expression of interferon regulatory factor-1 (IRF1), while IRF1 upregulation was demonstrated to inhibit exogenous DNA expression in a similar fashion as SAMHD1. IFNλ1, whose induction has been associated with IRF1 activation, is dispensable for SAMHD1/IRF1-mediated restriction of exogenous DNA, and neither type I nor II interferons appear to be involved. We also demonstrate that SAMHD1/IRF1-mediated restriction can effectively inhibit hepatitis B virus (HBV) antigen expression and progeny virus production in mouse models. In conclusion, these data support restriction of exogenous DNA as a novel function of SAMHD1.
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Diabetic nephrology (DN) is attributed largely to the depletion of podocytes, which is closely associated to apoptosis. However, the complex mechanism of podocyte loss in DN pathogenesis remains unclear. Recently, necroptosis has emerged as an important cell death model in many pathological conditions, which is regulated through RIPK1/RIPK3 pathway. In addition, necroptosis was found to share several upstream signaling pathways with apoptosis. Therefore, it was speculated that both apoptosis and necroptosis may occur in podocytes during the process of podocyte injury in DN. Herein, necroptosis and apoptosis were shown to be involved in podocyte injury induced by high glucose (HG), both in vitro and in vivo, with a high level of positive signaling markers RIPK1 (298.4⯱â¯17.35), cleaved caspase 3 (497.1⯱â¯23.09), RIPK3 (108.4⯱â¯14.92), and MLKL (470.4⯱â¯15.73) than the control groups. Scaning electron microscopy examination revealed the morphological characteristics of necroptotic and apoptotic cells, which differed remarkably. z-VAD-fmk, a pan-inhibitor of apoptosis, could block apoptosis and enhance necroptosis. Furthermore, UCHL1 was found to play a major role in promoting podocyte necroptosis by regulating the ubiquitination state of the RIPK1/RIPK3 pathway. The half-life of RIPK1 and RIPK3 proteins reduced and the expression of RIPK1, RIPK3, and MLKL decreased significantly after the knockdown of UCHL1. It was shown that UCHL1 exerted a more regulatory response to necroptosis. These data suggested that necroptosis may have more effect on the loss of podocytes than apoptosis in DN with the regulation of UCHL1. Thus, inhibiting UCHL1 to downregulate the RIPK1/RIPK3 pathway may be a novel strategy to protect the podocytes in DN patients.
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Apoptose/efeitos dos fármacos , Glucose/toxicidade , Necroptose/efeitos dos fármacos , Podócitos/patologia , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Transdução de Sinais , Ubiquitina Tiolesterase/metabolismo , Adulto , Clorometilcetonas de Aminoácidos/farmacologia , Caspase 3/metabolismo , Forma Celular/efeitos dos fármacos , Células Cultivadas , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Feminino , Humanos , Imidazóis/farmacologia , Indóis/farmacologia , Masculino , Pessoa de Meia-Idade , Podócitos/efeitos dos fármacos , Podócitos/ultraestrutura , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Quinases/metabolismo , Proteólise/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Ubiquitina/metabolismo , Ubiquitinação/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
In this study, activated carbon (AC) was modified with micro-sized geothite (mFeOOH) using a facile and cost-effective impregnation method for enhanced Cr(VI) removal from aqueous solutions. X-ray diffraction (XRD) and scanning electron microscope (SEM) analysis showed that FeOOH particles with a diameter of 0.1-1⯵m were dispersed homogeneously on the surfaces and pores of the AC. Fourier transform infrared spectrum (FTIR) and X-ray photoelectron spectra (XPS) analysis indicated that Cr(VI) was easily adsorbed onto the mFeOOH and reduced to Cr(III) by the AC, eventually deposited as Cr(III)-Fe(III) hydroxides (e.g., (CrxFe1-x)(OH)3). Hence, the mFeOOH@AC achieved a significantly higher Cr(VI) removal efficiency of 90.4%, 4.5 times of that the AC. The adsorption of Cr(VI) onto the mFeOOH@AC agreed well with the Langmuir adsorption model, demonstrating that the adsorption process was controlled by monolayer adsorption. This adsorption process also followed the pseudo second-order kinetics and the adsorption rate constant K2 was determined to be 0.013â¯g/mg·min. The Cr(VI) removal efficiency decreased with pH values as the adsorption process was highly pH-dependent. After the desorption-adsorption process by 0.1â¯M HCl solution for 4â¯cycles, the removal efficiency of Cr(VI) was still kept up to 75.1%, indicating that the mFeOOH@AC has a good stability and can be easily regenerated. In addition, the mFeOOH@AC also exhibited a promising potential for reutilization since a Cr(VI) removal efficiency of 85.4% was achieved after stripping all the mFeOOH and Cr(III)-Fe(III) hydroxides by 1â¯M HCl solution and regeneration with mFeOOH. We demonstrate that the modified AC with micro-sized goethite can remarkably enhance its ability for Cr(VI) removal in water treatment.
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In this study, a sequential Fe0/H2O2 reaction and biological process was employed as a low-cost depth treatment method to remove recalcitrant compounds from coal-chemical engineering wastewater after regular biological treatment. First of all, a chemical oxygen demand (COD) and color removal efficiency of 66 and 63% was achieved at initial pH of 6.8, 25 mmol L-1 of H2O2, and 2 g L-1 of Fe0 in the Fe0/H2O2 reaction. According to the gas chromatography-mass spectrometer (GC-MS) and gas chromatography-flame ionization detector (GC-FID) analysis, the recalcitrant compounds were effectively decomposed into short-chain organic acids such as acetic, propionic, and butyric acids. Although these acids were resistant to the Fe0/H2O2 reaction, they were effectively eliminated in the sequential air lift reactor (ALR) at a hydraulic retention time (HRT) of 2 h, resulting in a further decrease of COD and color from 120 to 51 mg L-1 and from 70 to 38 times, respectively. A low operational cost of 0.35 $ m-3 was achieved because pH adjustment and iron-containing sludge disposal could be avoided since a total COD and color removal efficiency of 85 and 79% could be achieved at an original pH of 6.8 by the above sequential process with a ferric ion concentration below 0.8 mg L-1 after the Fe0/H2O2 reaction. It indicated that the above sequential process is a promising and cost-effective method for the depth treatment of coal-chemical engineering wastewaters to satisfy discharge requirements.