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1.
Kidney Int ; 98(6): 1502-1518, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33038424

RESUMO

COVID-19 morbidity and mortality are increased via unknown mechanisms in patients with diabetes and kidney disease. SARS-CoV-2 uses angiotensin-converting enzyme 2 (ACE2) for entry into host cells. Because ACE2 is a susceptibility factor for infection, we investigated how diabetic kidney disease and medications alter ACE2 receptor expression in kidneys. Single cell RNA profiling of kidney biopsies from healthy living donors and patients with diabetic kidney disease revealed ACE2 expression primarily in proximal tubular epithelial cells. This cell-specific localization was confirmed by in situ hybridization. ACE2 expression levels were unaltered by exposures to renin-angiotensin-aldosterone system inhibitors in diabetic kidney disease. Bayesian integrative analysis of a large compendium of public -omics datasets identified molecular network modules induced in ACE2-expressing proximal tubular epithelial cells in diabetic kidney disease (searchable at hb.flatironinstitute.org/covid-kidney) that were linked to viral entry, immune activation, endomembrane reorganization, and RNA processing. The diabetic kidney disease ACE2-positive proximal tubular epithelial cell module overlapped with expression patterns seen in SARS-CoV-2-infected cells. Similar cellular programs were seen in ACE2-positive proximal tubular epithelial cells obtained from urine samples of 13 hospitalized patients with COVID-19, suggesting a consistent ACE2-coregulated proximal tubular epithelial cell expression program that may interact with the SARS-CoV-2 infection processes. Thus SARS-CoV-2 receptor networks can seed further research into risk stratification and therapeutic strategies for COVID-19-related kidney damage.


Assuntos
/metabolismo , Nefropatias Diabéticas/metabolismo , Túbulos Renais Proximais/metabolismo , /metabolismo , Adulto , Idoso , Antagonistas de Receptores de Angiotensina/farmacologia , Antagonistas de Receptores de Angiotensina/uso terapêutico , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , /virologia , Estudos de Casos e Controles , Nefropatias Diabéticas/tratamento farmacológico , Feminino , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Interações Hospedeiro-Patógeno , Humanos , Túbulos Renais Proximais/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade
2.
Life Sci ; 263: 118672, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33121990

RESUMO

AIMS: Cisplatin is an anticancer agent marred by nephrotoxicity. Limiting this adverse effect may allow the use of higher doses to improve its efficacy. The Wnt/ß-catenin signaling pathway plays a critical role in nephrogenesis and repair of renal diseases. BIO, a small molecule agonist of this pathway, exerted a protective effect in adriamycin nephropathy and promoted nephrogenesis. The aim of this study, therefore, was to investigate whether Wnt/ß-catenin agonist BIO could protect against cisplatin-induced nephrotoxicity in vivo and in vitro, as well as its possible mechanism. MAIN METHODS: Male mice and human renal proximal tubular cells (HK-2) were subjected to cisplatin to study reno-protective effect of BIO. Renal function, cell viability, tubular apoptosis, production of reactive oxygen species (ROS) and proliferative level were analyzed respectively. Additionally, xenograft model was induced to investigate if BIO would impair the antitumor effect of cisplatin. KEY FINDINGS: Cisplatin increased serum creatinine levels and promoted histological renal injury as well as oxidative stress levels. Besides, renal apoptotic level and the expression of pro-apoptotic proteins, Bax/bcl-2 and cleaved-caspase3 included, in the kidney were increased. All these features were decreased by BIO, which also activated Wnt/ß-catenin pathway in cisplatin-induced nephrotoxicity. Similarly, accompanied by the motivation of Wnt/ß-catenin pathway, BIO exerted a positively protective effect on HK-2 challenged cisplatin. Last, the chemotherapeutic effects of cisplatin in xenograft mice of ovary tumor models and in lung cancer cells weren't compromised by BIO. SIGNIFICANCE: Wnt/ß-catenin agonist BIO has the potential to prevent cisplatin nephrotoxicity without compromising its anti-proliferation efficacy.


Assuntos
Antineoplásicos/toxicidade , Cisplatino/toxicidade , Indóis/farmacologia , Nefropatias/prevenção & controle , Neoplasias Ovarianas/tratamento farmacológico , Oximas/farmacologia , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Feminino , Humanos , Nefropatias/induzido quimicamente , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Estresse Oxidativo/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Metabolism ; 113: 154405, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33069809

RESUMO

AIMS: The crosstalk between sodium-glucose cotransporter 2 (SGLT2) inhibition and a membrane-associated endocytic receptor megalin function involved in renal proximal tubular protein overload in progressive diabetic nephropathy (DN) is uncertain. Here, we determined whether SGLT2 inhibition affects megalin endocytic function through suppressing its O-linked ß-N-acetylglucosamine modification (O-GlcNAcylation) and protects the diabetic kidney from protein overload. MATERIALS AND METHOD: We treated 8-week-old male non-obese and hypoinsulinemic KK/Ta-Ins2Akita (KK/Ta-Akita) mice which develop progressive DN with an SGLT2 inhibitor ipragliflozin or insulin for 6 weeks, and investigated the endocytic function (proximal tubular protein reabsorption), renal expression and O-GlcNAcylation of megalin along with their effects on renal phenotypes including histology and biochemical markers. RESULTS: The treatment with ipragliflozin, but not insulin, suppressed megalin O-GlcNAcylation and accelerated its internalization, resulting in reduction in proximal tubular reabsorption of the highly filtered plasma proteins such as albumin and neutrophil gelatinase-associated lipocalin. These alterations following the ipragliflozin treatment contributed to amelioration of proximal tubular protein overload, mitochondrial morphological abnormality, and renal oxidative stress and tubulointerstitial fibrosis. CONCLUSIONS: The present study provides a novel crosstalk mechanism between SGLT2 inhibition and megalin underlying the potential renal benefits of SGLT2 inhibition in DN.


Assuntos
Acetilglucosamina/metabolismo , Nefropatias Diabéticas/metabolismo , Túbulos Renais Proximais/metabolismo , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteínas/metabolismo , Transportador 2 de Glucose-Sódio/metabolismo , Acilação , Animais , Nefropatias Diabéticas/urina , Progressão da Doença , Masculino , Camundongos , Camundongos Endogâmicos C57BL
4.
Cell Prolif ; 53(10): e12902, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32945585

RESUMO

OBJECTIVES: Calcium oxalate (CaOx) crystals can activate inflammatory cytokines by triggering inflammasomes, which cause damage to the adhered epithelium, a dysfunctional microenvironment and even renal failure. However, a comprehensive and in-depth understanding of the mechanisms underlying the effects of these crystals on damage and cytokine function in renal tubular epithelial cells (TECs) remains limited and to be explored. MATERIALS AND METHODS: We detected the pyroptosis of TECs induced after exposure to CaOx crystals and demonstrated the significance of cytokine activation in the subsequent inflammatory processes through a proteomic study. We then conducted animal and cell experiments to verify relevant mechanisms through morphological, protein, histological and biochemical approaches. Human serum samples were further tested to help explain the pathophysiological mechanism of H3 relaxin. RESULTS: We verified that crystal-induced extracellular adenosine triphosphate (ATP) upregulation via the membrane purinergic 2X7 receptor (P2X7 R) promotes ROS generation and thereby activates NLRP3 inflammasome-mediated interleukin-1ß/18 maturation and gasdermin D cleavage. Human recombinant relaxin-3 (H3 relaxin) can act on the transmembrane receptor RXFP1 to produce cAMP and subsequently improves crystal-derived damage via ATP consumption. Additionally, endogenous relaxin-3 was found to be elevated in patients with renal calculus and can thus serve as a biomarker. CONCLUSIONS: Our results provide previously unidentified mechanistic insights into CaOx crystal-induced inflammatory pyroptotic damage and H3 relaxin-mediated anti-inflammatory protection and thus suggest a series of potential therapeutic targets and methods for but not limited to nephrocalcinosis.


Assuntos
Anti-Inflamatórios/farmacologia , Oxalato de Cálcio/farmacologia , Piroptose/efeitos dos fármacos , Relaxina/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Oxalato de Cálcio/química , Linhagem Celular , AMP Cíclico/metabolismo , Humanos , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Masculino , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Receptores de Peptídeos/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Relaxina/sangue
5.
Autophagy ; 16(12): 2287-2288, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32954913

RESUMO

The kidney, similar to many other organs, has to face shear stress induced by biological fluids. How epithelial kidney cells respond to shear stress is poorly understood. Recently we showed in vitro and in vivo that proximal tubule epithelial cells use lipophagy to fuel mitochondria with fatty acids. Lipophagy is stimulated by a primary cilium-dependent signaling that converges at AMP kinase. AMP kinase is a central signaling hub to trigger lipophagy and also to stimulate mitochondrial biogenesis. These two pathways contribute to generate ATP needed to support energy-consuming cellular processes such as glucose reabsorption, gluconeogenesis. These findings demonstrate the role of the primary cilium and selective macroautophagy/autophagy to integrate shear stress and to sustain the execution of a specific cellular program.


Assuntos
Autofagia , Cílios , Cílios/metabolismo , Células Epiteliais/metabolismo , Rim , Túbulos Renais Proximais/metabolismo , Mitocôndrias , Biogênese de Organelas
6.
Am J Physiol Renal Physiol ; 319(4): F697-F711, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32865013

RESUMO

Praliciguat, a clinical-stage soluble guanylate cyclase (sGC) stimulator, increases cGMP via the nitric oxide-sGC pathway. Praliciguat has been shown to be renoprotective in rodent models of hypertensive nephropathy and renal fibrosis. In the present study, praliciguat alone and in combination with enalapril attenuated proteinuria in the obese ZSF1 rat model of diabetic nephropathy. Praliciguat monotherapy did not affect hemodynamics. In contrast, enalapril monotherapy lowered blood pressure but did not attenuate proteinuria. Renal expression of genes in pathways involved in inflammation, fibrosis, oxidative stress, and kidney injury was lower in praliciguat-treated obese ZSF1 rats than in obese control rats; fasting glucose and cholesterol were also lower with praliciguat treatment. To gain insight into how tubular mechanisms might contribute to its pharmacological effects on the kidneys, we studied the effects of praliciguat on pathological processes and signaling pathways in cultured human primary renal proximal tubular epithelial cells (RPTCs). Praliciguat inhibited the expression of proinflammatory cytokines and secretion of monocyte chemoattractant protein-1 in tumor necrosis factor-α-challenged RPTCs. Praliciguat treatment also attenuated transforming growth factor-ß-mediated apoptosis, changes to a mesenchyme-like cellular phenotype, and phosphorylation of SMAD3 in RPTCs. In conclusion, praliciguat improved proteinuria in the ZSF1 rat model of diabetic nephropathy, and its actions in human RPTCs suggest that tubular effects may contribute to its renal benefits, building upon strong evidence for the role of cGMP signaling in renal health.


Assuntos
Apoptose/efeitos dos fármacos , Nefropatias Diabéticas/tratamento farmacológico , Agonistas da Guanilil Ciclase C/farmacologia , Túbulos Renais Proximais/efeitos dos fármacos , Nefrite/tratamento farmacológico , Pirazóis/farmacologia , Pirimidinas/farmacologia , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Animais , Linhagem Celular , Citocinas/metabolismo , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Progressão da Doença , Enalapril/farmacologia , Humanos , Mediadores da Inflamação/metabolismo , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Masculino , Nefrite/metabolismo , Nefrite/patologia , Fosforilação , Ratos Zucker , Transdução de Sinais , Proteína Smad3/metabolismo
7.
Am J Physiol Renal Physiol ; 319(4): F712-F728, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32893663

RESUMO

Inhibitors of proximal tubular Na+-glucose cotransporter 2 (SGLT2) are natriuretic, and they lower blood pressure. There are reports that the activities of SGLT2 and Na+-H+ exchanger 3 (NHE3) are coordinated. If so, then part of the natriuretic response to an SGLT2 inhibitor is mediated by suppressing NHE3. To examine this further, we compared the effects of an SGLT2 inhibitor, empagliflozin, on urine composition and systolic blood pressure (SBP) in nondiabetic mice with tubule-specific NHE3 knockdown (NHE3-ko) and wild-type (WT) littermates. A single dose of empagliflozin, titrated to cause minimal glucosuria, increased urinary excretion of Na+ and bicarbonate and raised urine pH in WT mice but not in NHE3-ko mice. Chronic empagliflozin treatment tended to lower SBP despite higher renal renin mRNA expression and lowered the ratio of SBP to renin mRNA, indicating volume loss. This effect of empagliflozin depended on tubular NHE3. In diabetic Akita mice, chronic empagliflozin enhanced phosphorylation of NHE3 (S552/S605), changes previously linked to lesser NHE3-mediated reabsorption. Chronic empagliflozin also increased expression of genes involved with renal gluconeogenesis, bicarbonate regeneration, and ammonium formation. While this could reflect compensatory responses to acidification of proximal tubular cells resulting from reduced NHE3 activity, these effects were at least in part independent of tubular NHE3 and potentially indicated metabolic adaptations to urinary glucose loss. Moreover, empagliflozin increased luminal α-ketoglutarate, which may serve to stimulate compensatory distal NaCl reabsorption, while cogenerated and excreted ammonium balances urine losses of this "potential bicarbonate." The data implicate NHE3 as a determinant of the natriuretic effect of empagliflozin.


Assuntos
Compostos Benzidrílicos/farmacologia , Diabetes Mellitus/tratamento farmacológico , Glucosídeos/farmacologia , Túbulos Renais Proximais/efeitos dos fármacos , Natriurese/efeitos dos fármacos , Natriuréticos/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Transportador 2 de Glucose-Sódio/metabolismo , Trocador 3 de Sódio-Hidrogênio/metabolismo , Equilíbrio Ácido-Base/efeitos dos fármacos , Animais , Glicemia/metabolismo , Pressão Sanguínea/efeitos dos fármacos , Diabetes Mellitus/metabolismo , Diabetes Mellitus/fisiopatologia , Modelos Animais de Doenças , Glicosúria/metabolismo , Glicosúria/fisiopatologia , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Trocador 3 de Sódio-Hidrogênio/deficiência , Trocador 3 de Sódio-Hidrogênio/genética
8.
Nat Commun ; 11(1): 4467, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32948751

RESUMO

Recent studies have reported that upregulation of disulfide-bond A oxidoreductase-like protein (DsbA-L) prevented lipid-induced renal injury in diabetic nephropathy (DN). However, the role and regulation of proximal tubular DsbA-L for renal tubulointerstitial fibrosis (TIF) remains unclear. In current study, we found that a proximal tubules-specific DsbA-L knockout mouse (PT-DsbA-L-KO) attenuated UUO-induced TIF, renal cell apoptosis and inflammation. Mechanistically, the DsbA-L interacted with Hsp90 in mitochondria of BUMPT cells which activated the signaling of Smad3 and p53 to produce connective tissue growth factor (CTGF) and then resulted in accumulation of ECM of BUMPT cells and mouse kidney fibroblasts. In addition, the progression of TIF caused by UUO, ischemic/reperfusion (I/R), aristolochic acid, and repeated acute low-dose cisplatin was also alleviated in PT-DsbA-L-KO mice via the activation of Hsp90 /Smad3 and p53/CTGF axis. Finally, the above molecular changes were verified in the kidney biopsies from patients with obstructive nephropathy (Ob). Together, these results suggest that DsbA-L in proximal tubular cells promotes TIF via activation of the Hsp90 /Smad3 and p53/CTGF axis.


Assuntos
Fibrose/genética , Predisposição Genética para Doença/genética , Glutationa Transferase/genética , Glutationa Transferase/metabolismo , Nefropatias/genética , Idoso , Animais , Apoptose , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Nefropatias Diabéticas , Modelos Animais de Doenças , Feminino , Fibrose/patologia , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Inflamação , Rim/lesões , Nefropatias/patologia , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Transdução de Sinais , Proteína Smad3/metabolismo , Proteína Supressora de Tumor p53/metabolismo
9.
Am J Physiol Renal Physiol ; 319(5): F941-F953, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32865015

RESUMO

Tubulointerstitial fibrosis has been regarded as a critical event in the pathogenesis of chronic kidney disease. The soluble form of (pro)renin receptor (sPRR), generated by site-1 protease (S1P) cleavage of full-length PRR, can be detected in biological fluid and elevated under certain pathological conditions. The present study was designed to evaluate the potential role of sPRR in the regulation of the fibrotic response in a cultured human renal proximal tubular cell line (HK-2 cells) in the setting of transforming growth factor (TGF)-ß or sPRR-His treatment. The TGF-ß-induced fibrotic response of HK-2 cells was indicated by upregulation of fibronectin (FN) expression; meanwhile, TGF-ß could also induce the generation of sPRR, due to enhanced cleavage of full-length PRR. To explore the role of sPRR in the fibrotic response of HK-2 cells, we blocked the production of sPRR with a the S1P inhibitor PF429242 and found that PF429242 remarkably suppressed TGF-ß-induced sPRR generation and FN expression in HK-2 cells. Administration of sPRR-His restored the PF429242-attenuated FN expression in HK-2 cells, indicating that sPRR could promote the TGF-ß-induced fibrotic response. Furthermore, sPRR-His alone also increased the abundance of FN in HK-2 cells. These data suggested that sPRR was sufficient and necessary for the TGF-ß-induced fibrotic response of HK-2 cells. Mechanistically, sPRR activated the AKT and ß-catenin pathway in HK-2 cells, and blockade of the AKT or ß-catenin pathway significantly abrogated sPRR-induced FN and Snail expression. Taking together, sPRR promoted the fibrotic response of HK-2 cells by activating Akt/ß-catenin/Snail signaling, and it may serve as a potential therapeutic target in renal fibrosis.


Assuntos
Células Epiteliais/metabolismo , Túbulos Renais Proximais/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , beta Catenina/metabolismo , Humanos , Rim/metabolismo , Receptores de Superfície Celular/metabolismo , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia , Transdução de Sinais/fisiologia , Regulação para Cima/fisiologia
10.
Am J Physiol Renal Physiol ; 319(5): F773-F779, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32954853

RESUMO

Diabetes is a prevalent metabolic disease that contributes to ∼50% of all end-stage renal disease and has limited treatment options. We previously demonstrated that the ß2-adrenergic receptor agonist formoterol induced mitochondrial biogenesis and promoted recovery from acute kidney injury. Here, we assessed the effects of formoterol on mitochondrial dysfunction and dynamics in renal proximal tubule cells (RPTCs) treated with high glucose and in a mouse model of type 2 diabetes. RPTCs exposed to 17 mM glucose exhibited increased electron transport chain (ETC) complex I, II, III, and V protein levels and reduced ATP levels and uncoupled oxygen consumption rate compared with RPTCs cultured in the absence of glucose or osmotic controls after 96 h. ETC proteins, ATP, and oxygen consumption rate were restored in RPTCs treated with formoterol. RPTCs exposed to high glucose had increased phospho-dynamin-related protein 1 (Drp1), a mitochondrial fission protein, and decreased mitofusin 1 (Mfn1), a mitochondrial fusion protein. Formoterol treatment restored phospho-Drp1 and Mfn1 to control levels. Db/db and nondiabetic (db/m) mice (10 wk old) were treated with formoterol or vehicle for 3 wk and euthanized. Db/db mice showed increased renal cortical ETC protein levels in complexes I, III, and V and decreased ATP; these changes were prevented by formoterol. Phospho-Drp1 was increased and Mfn1 was decreased in db/db mice, and formoterol restored both to control levels. Together, these findings demonstrate that hyperglycemic conditions in vivo and exposure of RPTCs to high glucose similarly alter mitochondrial bioenergetic and dynamics profiles and that treatment with formoterol can reverse these effects. Formoterol may be a promising strategy for treating early stages of diabetic kidney disease.


Assuntos
Agonistas Adrenérgicos/farmacologia , Fumarato de Formoterol/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Dinâmica Mitocondrial/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Rim/efeitos dos fármacos , Rim/metabolismo , Túbulos Renais Proximais/metabolismo , Camundongos , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Biogênese de Organelas
11.
Cell Prolif ; 53(11): e12909, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32975326

RESUMO

OBJECTIVES: In diabetic nephropathy (DN), hypoxia-inducible factor-1α (HIF-1α) activation in tubular cells plays an important protective role against kidney injury. The effects may occur via the target genes of HIF-1α, such as haem oxygenase-1 (HO-1), but the exact mechanisms are incompletely understood. MATERIALS AND METHODS: Mice with proximal tubule-specific knockout of HIF-1α (PT-HIF-1α-/- mice) were generated, and diabetes was induced in these mice by streptozotocin (STZ) injection. In addition, to mimic a hypoxic state, cobaltous chloride (CoCl2 ) was applied to HK-2 cells. RESULTS: Our study first verified that conditional knockout of HIF-1α worsened tubular injury in DN; additionally, aggravated kidney dysfunction, renal histopathological alterations, mitochondrial fragmentation, ROS accumulation and apoptosis were observed in diabetic PT-HIF-1α-/- mice. In vitro study showed that compared to control group, HK-2 cells cultured under hypoxic ambiance displayed increased mitochondrial fragmentation, ROS production, mitochondrial membrane potential loss and apoptosis. These increases were reversed by overexpression of HIF-1α or treatment with a HO-1 agonist. Importantly, cotreatment with a HIF-1α inhibitor and a HO-1 agonist rescued the HK-2 cells from the negative impacts of the HIF-1α inhibitor. CONCLUSIONS: These data revealed that HIF-1α exerted a protective effect against tubular injury in DN, which could be mediated via modulation of mitochondrial dynamics through HO-1 upregulation.


Assuntos
Nefropatias Diabéticas/patologia , Heme Oxigenase-1/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Túbulos Renais Proximais/patologia , Dinâmica Mitocondrial , Animais , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Deleção de Genes , Técnicas de Inativação de Genes , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia
12.
Cell Rep ; 32(12): 108175, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32946807

RESUMO

To predict the tropism of human coronaviruses, we profile 28 SARS-CoV-2 and coronavirus-associated receptors and factors (SCARFs) using single-cell transcriptomics across various healthy human tissues. SCARFs include cellular factors both facilitating and restricting viral entry. Intestinal goblet cells, enterocytes, and kidney proximal tubule cells appear highly permissive to SARS-CoV-2, consistent with clinical data. Our analysis also predicts non-canonical entry paths for lung and brain infections. Spermatogonial cells and prostate endocrine cells also appear to be permissive to SARS-CoV-2 infection, suggesting male-specific vulnerabilities. Both pro- and anti-viral factors are highly expressed within the nasal epithelium, with potential age-dependent variation, predicting an important battleground for coronavirus infection. Our analysis also suggests that early embryonic and placental development are at moderate risk of infection. Lastly, SCARF expression appears broadly conserved across a subset of primate organs examined. Our study establishes a resource for investigations of coronavirus biology and pathology.


Assuntos
Infecções por Coronavirus/patologia , Mucosa Nasal/metabolismo , Pneumonia Viral/patologia , Receptores Virais/genética , Tropismo Viral/genética , Internalização do Vírus , Células A549 , Animais , Betacoronavirus/crescimento & desenvolvimento , Linhagem Celular , Chlorocebus aethiops , Enterócitos/metabolismo , Perfilação da Expressão Gênica , Células Caliciformes/metabolismo , Células HEK293 , Humanos , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Mucosa Nasal/virologia , Pandemias , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Análise de Célula Única , Células Vero
13.
Cancer Sci ; 111(11): 4276-4287, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32860304

RESUMO

End-stage renal disease (ESRD) patients on dialysis therapy have a higher incidence of renal cell carcinomas (RCCs), which consist of 2 major histopathological types: clear-cell RCCs (ESRD-ccRCCs) and acquired cystic disease (ACD)-associated RCCs. However, their genetic and epigenetic alterations are still poorly understood. Here, we investigated somatic mutations, copy number alterations (CNAs), and DNA methylation profiles in 9 ESRD-ccRCCs and 7 ACD-associated RCCs to identify their molecular alterations and cellular origins. Targeted sequencing of 409 cancer-related genes, including VHL, PBRM1, SETD2, BAP1, KDM5C, MET, KMT2C (MLL3), and TP53, showed ESRD-ccRCCs harbored frequent VHL mutations, while ACD-associated RCCs did not. CNA analysis showed that ESRD-ccRCCs had a frequent loss of chromosome 3p while ACD-associated RCCs had a gain of chromosome 16. Beadarray methylation analysis showed that ESRD-ccRCCs had methylation profiles similar to those of sporadic ccRCCs, while ACD-associated RCCs had profiles similar to those of papillary RCCs. Expression analysis of genes whose expression levels are characteristic to individual segments of a nephron showed that ESRD-ccRCCs and ACD-associated RCCs had high expression of proximal tubule cell marker genes, while chromophobe RCCs had high expression of distal tubule cell/collecting duct cell marker genes. In conclusion, ESRD-ccRCCs and ACD-associated RCCs had mutation and methylation profiles similar to those of sporadic ccRCCs and papillary RCCs, respectively, and these 2 histopathological types of RCCs were indicated to have originated from proximal tubule cells of the nephron.


Assuntos
Epigênese Genética , Predisposição Genética para Doença , Falência Renal Crônica/complicações , Falência Renal Crônica/genética , Neoplasias Renais/etiologia , Neoplasias Renais/patologia , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Adulto , Idoso , Carcinoma de Células Renais/etiologia , Carcinoma de Células Renais/patologia , Variações do Número de Cópias de DNA , Epigenoma , Feminino , Perfilação da Expressão Gênica , Genótipo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Mutação , Transcriptoma
14.
Mol Genet Genomic Med ; 8(10): e1442, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32744436

RESUMO

BACKGROUND: A novel coronavirus called SARS-Cov-2, which shared 82% similarity of genome sequence with SARS-CoV, was found in Wuhan in late December of 2019, causing an epidemic outbreak of novel coronavirus-induced pneumonia with dramatically increasing number of cases. Several organs are vulnerable to COVID-19 infection. Acute kidney injury (AKI) was reported in parts of case-studies reporting characteristics of COVID-19 patients. This study aimed at analyzing the potential route of SARS-Cov-2 entry and mechanism at cellular level. METHOD: Single-cell RNA sequencing (scRNA-seq) technology was used to obtain evidence of potential route and ACE2 expressing cell in renal system for underlying pathogenesis of kidney injury caused by COVID-19. The whole process was performed under R with Seurat packages. Canonical marker genes were used to annotate different types of cells. RESULTS: Ten different clusters were identified and ACE2 was mainly expressed in proximal tubule and glomerular parietal epithelial cells. From Gene Ontology (GO) & KEGG enrichment analysis, imbalance of ACE2 expression, renin-angiotensin system (RAS) activation, and neutrophil-related processes were the main issue of COVID-19 leading kidney injury. CONCLUSION: Our study provided the cellular evidence that SARS-Cov-2 invaded human kidney tissue via proximal convoluted tubule, proximal tubule, proximal straight tubule cells, and glomerular parietal cells by means of ACE2-related pathway and used their cellular protease TMPRSS2 for priming.


Assuntos
Lesão Renal Aguda/virologia , /patologia , Glomérulos Renais/metabolismo , Túbulos Renais Proximais/metabolismo , Receptores Virais/genética , Lesão Renal Aguda/patologia , Sequência de Bases , Humanos , Glomérulos Renais/patologia , Glomérulos Renais/virologia , Túbulos Renais Proximais/patologia , Túbulos Renais Proximais/virologia , Análise de Componente Principal , Análise de Sequência de RNA , Serina Endopeptidases/metabolismo , Análise de Célula Única
15.
Am J Physiol Renal Physiol ; 319(4): F579-F591, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32799673

RESUMO

Tubular atrophy is a common pathological feature of kidney fibrosis. Although fibroblasts play a predominant role in tissue fibrosis, the role of repairing tubular epithelia in tubular atrophy is unclear. We demonstrated the essential role of focal adhesion kinase (FAK)-mediated intratubular epithelial-mesenchymal transition (EMT) in the pathogenesis of tubular atrophy after severe ischemia-reperfusion injury (IRI). Actively proliferating tubular epithelia undergoing intratubular EMT were noted in the acute phase of severe IRI, resulting in tubular atrophy in the chronic phase, reflecting failed tubular repair. Furthermore, FAK was phosphorylated in the tubular epithelia in the acute phase of severe IRI, and its inhibition ameliorated both tubular atrophy and interstitial fibrosis in the chronic phase after injury. In vivo clonal analysis of single-labeled proximal tubular epithelial cells after IRI using proximal tubule reporter mice revealed substantial clonal expansion after IRI, reflecting active epithelial proliferation during repair. The majority of these proliferating epithelia were located in atrophic and nonfunctional tubules, and FAK inhibition was sufficient to prevent tubular atrophy. In vitro, transforming growth factor-ß induced FAK phosphorylation and an EMT phenotype, which was also prevented by FAK inhibition. In an in vitro tubular epithelia gel contraction assay, transforming growth factor-ß treatment accelerated gel contraction, which was suppressed by FAK inhibition. In conclusion, injury-induced intratubular EMT is closely related to tubular atrophy in a FAK-dependent manner.


Assuntos
Lesão Renal Aguda/patologia , Células Epiteliais/patologia , Transição Epitelial-Mesenquimal , Túbulos Renais Proximais/patologia , Lesão Renal Aguda/tratamento farmacológico , Lesão Renal Aguda/metabolismo , Animais , Atrofia , Linhagem Celular , Proliferação de Células , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fibrose , Quinase 1 de Adesão Focal/antagonistas & inibidores , Quinase 1 de Adesão Focal/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos Transgênicos , Fenótipo , Fosforilação , Ratos , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIa/genética , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIa/metabolismo
16.
Am J Physiol Renal Physiol ; 319(4): F603-F611, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32830538

RESUMO

The acyl-CoA synthetase medium-chain family member 2 (Acsm2) gene was first identified and cloned by our group as a kidney-specific "KS" gene. However, its expression pattern and function remain to be clarified. In the present study, we found that the Acsm2 gene was expressed specifically and at a high level in normal adult kidneys. Expression of Acsm2 in kidneys followed a maturational pattern: it was low in newborn mice and increased with kidney development and maturation. In situ hybridization and immunohistochemistry revealed that Acsm2 was expressed specifically in proximal tubular cells of adult kidneys. Data from the Encyclopedia of DNA Elements database revealed that the Acsm2 gene locus in the mouse has specific histone modifications related to the active transcription of the gene exclusively in kidney cells. Following acute kidney injury, partial unilateral ureteral obstruction, and chronic kidney diseases, expression of Acsm2 in the proximal tubules was significantly decreased. In human samples, the expression pattern of ACSM2A, a homolog of mouse Acsm2, was similar to that in mice, and its expression decreased with several types of renal injuries. These results indicate that the expression of Acsm2 parallels the structural and functional maturation of proximal tubular cells. Downregulation of its expression in several models of kidney disease suggests that Acms2 may serve as a novel marker of proximal tubular injury and/or dysfunction.


Assuntos
Coenzima A Ligases/metabolismo , Células Epiteliais/metabolismo , Túbulos Renais Proximais/metabolismo , Lesão Renal Aguda/enzimologia , Lesão Renal Aguda/genética , Lesão Renal Aguda/patologia , Animais , Coenzima A Ligases/genética , Modelos Animais de Doenças , Células Epiteliais/patologia , Fibrose , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Humanos , Integrina beta1/genética , Integrina beta1/metabolismo , Túbulos Renais Proximais/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Insuficiência Renal Crônica/enzimologia , Insuficiência Renal Crônica/genética , Insuficiência Renal Crônica/patologia , Renina/genética , Renina/metabolismo , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/patologia
17.
Biosci Rep ; 40(9)2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32808659

RESUMO

Endotoxemia-induced acute kidney injury (AKI) is a common clinical condition that lacks effective treatments. Elabela (ELA) is a recently discovered kidney peptide hormone, encoded by the gene apela, and has been reported to improve cardio-renal outcomes in sepsis. However, ELA is a small peptide and is largely unsuitable for clinical use because of its short in vivo half-life. In the present study, we evaluated the potential renoprotective effects of a long-acting constant fragment (Fc)-ELA fusion protein in liposaccharide (LPS)-induced AKI in mice. LPS administration in mice for 5 days greatly lowered the gene expression of apela and impaired kidney function, as evidenced by elevated serum creatinine and the ratio of urine protein to creatinine. In addition, renal inflammation and macrophage infiltration were apparent in LPS-challenged mice. Treatment with the Fc-ELA fusion protein partially restored apela expression and attenuated the kidney inflammation. Moreover, LPS treatment induced reactive oxygen species (ROS) production and apoptosis in kidney HK-2 cells as well as in the mouse kidney, which were mitigated by ELA or Fc-ELA treatment. Finally, we found that ELA promoted the survival of HK-2 cells treated with LPS, and this action was abolished by LY204002, a PI3K/Akt inhibitor. Collectively, we have demonstrated that the Fc-ELA fusion protein has significant renoprotective activities against LPS-induced AKI in mice.


Assuntos
Lesão Renal Aguda/tratamento farmacológico , Fragmentos Fc das Imunoglobulinas/genética , Lipopolissacarídeos/toxicidade , Hormônios Peptídicos/genética , Proteínas Recombinantes de Fusão/farmacologia , Lesão Renal Aguda/induzido quimicamente , Lesão Renal Aguda/genética , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Fragmentos Fc das Imunoglobulinas/farmacologia , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Camundongos Endogâmicos C57BL , Hormônios Peptídicos/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Substâncias Protetoras/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Recombinantes de Fusão/genética
18.
Biochim Biophys Acta Mol Basis Dis ; 1866(11): 165907, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32738302

RESUMO

High salt diet (HSD), considered a public health problem worldwide, is associated with chronic degenerative diseases including renal diseases. However, little is known about the effects of HSD on renal function independently of the development of hypertension. To address the hypothesis that HSD induces renal injuries even without changes in blood pressure, BALB/c mice were fed for 7 days with chow with a high salt content (0.3-8%). Blood pressure did not change and there was a decrease in cortical (Na+ + K+)ATPase and NHE3 exchanger and an increase in renal fractional excretion of sodium. Positive correlations between Na+ intake or urinary sodium excretion with proteinuria were found. HSD did not change glomerular function and structure, but induced tubule-interstitial injury measured by an increase in collagen deposition, interstitial space and γ-GT activity, a marker of tubular injury. These effects were associated with a decrease in cortical albumin reabsorption and megalin expression. Similarly, the addition of NaCl 20 mM to the incubation medium of LLC-PK1 cells reduced megalin expression and albumin endocytosis indicating that HSD could have a direct effect on proximal tubule cells. Furthermore, tubule-interstitial injury was associated with pro-inflammatory and pro-fibrotic phenotypes with an increase in Th1 and Th17 phenotypes and a decrease in Tregs followed by increases in IL-6, -17, -10, TNF-α, IFN-γ and TGF-ß. Our results reveal a complex network involved in renal injuries induced by HSD independently of changes in blood pressure. These findings strengthen the importance of restriction of salt intake for the general population even for salt-resistant individuals.


Assuntos
Hipertensão/metabolismo , Inflamação/metabolismo , Cloreto de Sódio na Dieta/efeitos adversos , Animais , Pressão Sanguínea/efeitos dos fármacos , Endocitose/efeitos dos fármacos , Imunofluorescência , Hipertensão/induzido quimicamente , Immunoblotting , Imuno-Histoquímica , Inflamação/induzido quimicamente , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-17/metabolismo , Interleucina-6/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão , Suínos , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
19.
Clin Sci (Lond) ; 134(16): 2223-2234, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32808649

RESUMO

Exosomes have been shown to effectively regulate the biological functions of target cells. Here, we investigated the protective effect and mechanism of hypoxia-induced renal tubular epithelial cells (TECs)-derived exosomes on acute tubular injury. We found that in vitro hypoxia-induced tubular exosomes (Hy-EXOs) were protective in acute tubular injury by promoting TECs proliferation and improving mitochondrial functions. By using exosome miRNA sequencing, we identified miR-20a-5p was abundant and was a key mechanism for the protective effect of Hy-EXOs on tubular injury as up-regulation of miR-20a-5p enhanced but down-regulation of miR-20a-5p inhibited the protective effect of Hy-EXOs on tubular injury under hypoxia conditions. Further study in a mouse model of ischemia-reperfusion-induced acute kidney injury (IRI-AKI) also confirmed this notion as pre-treating mice with the miR-20a-5p agomir 48 h prior to AKI induction was capable of inhibiting IRI-AKI by lowering serum levels of creatinine and urea nitrogen, and attenuating the severity of tubular necrosis, F4/80(+) macrophages infiltration and vascular rarefaction. Mechanistically, the protective effect of miR-20a-5p on acute kidney injury (AKI) was associated with inhibition of TECs mitochondrial injury and apoptosis in vitro and in vivo. In conclusion, miR-20a-5p is enriched in hypoxia-derived tubular exosomes and protects against acute tubular injury. Results from the present study also reveal that miR-20a-5p may represent as a novel therapy for AKI.


Assuntos
Lesão Renal Aguda/genética , Células Epiteliais/metabolismo , Exossomos/genética , Túbulos Renais Proximais/metabolismo , MicroRNAs/genética , Traumatismo por Reperfusão/genética , Animais , Linhagem Celular , Modelos Animais de Doenças , Regulação da Expressão Gênica , Humanos , Hipóxia , Túbulos Renais Proximais/citologia , Masculino , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Transmissão , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Espécies Reativas de Oxigênio/metabolismo
20.
Biochim Biophys Acta Biomembr ; 1862(11): 183434, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32781157

RESUMO

Guanidinoacetate (GAA), which is a precursor of creatine, is mainly biosynthesized in the renal proximal tubular epithelial cells (RPTECs). Plasma concentration of GAA has been reported to be reduced in patients with monocarboxylate transporter 12 (MCT12) mutation (p.Q215X). However, the mechanism underlying GAA release from the RPTECs remains unclear. Therefore, to elucidate the role of MCT12 in renal GAA release, MCT12-mediated GAA transport was evaluated using the human and rat MCT12-expressing Xenopus laevis oocytes and primary-cultured rat RPTECs. [14C]GAA uptake by the human and rat MCT12-expressing oocytes was significantly higher than that by the water-injected oocytes. Rat MCT12-mediated uptake of [14C]GAA by the oocytes was found to be sodium ion (Na+)-independent and exhibited saturable kinetics with a Michaelis-Menten constant of 3.38 mM. Transport activities of rat MCT12 tend to increase along with increasing of extracellular pH. In addition, the efflux transport of [14C]GAA from the human and rat MCT12-expressing oocytes was significantly higher than that from the water-injected oocytes. These results suggest that both the influx and efflux transport of GAA is mediated by MCT12. In the primary-cultured rat RPTECs, [14C]GAA efflux transport was significantly reduced by the transfection of MCT12-specific siRNAs, suggesting that MCT12 participates in GAA efflux transport in rat RPTECs. Therefore, it suggests that MCT12 is involved in GAA release from RPTECs to the circulating blood, since MCT12 is known to be localized on the basal membrane of RPTECs.


Assuntos
Células Epiteliais/metabolismo , Glicina/análogos & derivados , Túbulos Renais Proximais/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Sódio/metabolismo , Animais , Linhagem Celular , Células Epiteliais/citologia , Feminino , Glicina/metabolismo , Cobaias , Humanos , Transporte de Íons , Túbulos Renais Proximais/citologia , Masculino , Transportadores de Ácidos Monocarboxílicos/genética , Oócitos , Ratos , Ratos Wistar , Xenopus laevis
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