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Sodium-glucose cotransporter 2 inhibitors (SGLT2is), initially developed for type 2 diabetes (T2D) treatment, have demonstrated significant cardiovascular and renal benefits in heart failure (HF) and chronic kidney disease (CKD), irrespective of T2D. This review provides an analysis of the multifaceted mechanisms underlying the cardiorenal benefits of SGLT2i in HF and CKD outside of the T2D context. Eight major aspects of the protective effects of SGLT2i beyond glycemic control are explored: 1) the impact on renal hemodynamics and tubuloglomerular feedback; 2) the natriuretic effects via proximal tubule Na+/H+ exchanger NHE3 inhibition; 3) the modulation of neurohumoral pathways with evidence of attenuated sympathetic activity; 4) the impact on erythropoiesis, not only in the context of local hypoxia but also systemic inflammation and iron regulation; 5) the uricosuria and mitigation of the hyperuricemic environment in cardiorenal syndromes; 6) the multiorgan metabolic reprogramming including the potential induction of a fasting-like state, improvement in glucose and insulin tolerance, and stimulation of lipolysis and ketogenesis; 7) the vascular endothelial growth factor A (VEGF-A) upregulation and angiogenesis, and 8) the direct cardiac effects. The intricate interplay between renal, neurohumoral, metabolic, and cardiac effects underscores the complexity of SGLT2i actions and provides valuable insights into their therapeutic implications for HF and CKD. Furthermore, this review sets the stage for future research to evaluate the individual contributions of these mechanisms in diverse clinical settings.
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Insuficiência Cardíaca , Insuficiência Renal Crônica , Inibidores do Transportador 2 de Sódio-Glicose , Humanos , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Insuficiência Cardíaca/tratamento farmacológico , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/fisiopatologia , Animais , Rim/efeitos dos fármacos , Rim/metabolismo , Trocador 3 de Sódio-Hidrogênio/metabolismo , Trocador 3 de Sódio-Hidrogênio/antagonistas & inibidoresRESUMO
Tubular proteinuria is a common feature in COVID-19 patients, even in the absence of established acute kidney injury. SARS-CoV-2 spike protein (S protein) was shown to inhibit megalin-mediated albumin endocytosis in proximal tubule epithelial cells (PTECs). Angiotensin-converting enzyme type 2 (ACE2) was not directly involved. Since Toll-like receptor 4 (TLR4) mediates S protein effects in various cell types, we hypothesized that TLR4 could be participating in the inhibition of PTECs albumin endocytosis elicited by S protein. Two different models of PTECs were used: porcine proximal tubule cells (LLC-PK1) and human embryonic kidney cells (HEK-293). S protein reduced Akt activity by specifically inhibiting of threonine 308 (Thr308) phosphorylation, a process mediated by phosphoinositide-dependent kinase 1 (PDK1). GSK2334470, a PDK1 inhibitor, decreased albumin endocytosis and megalin expression mimicking S protein effect. S protein did not change total TLR4 expression but decreased its surface expression. LPS-RS, a TLR4 antagonist, also counteracted the effects of the S protein on Akt phosphorylation at Thr308, albumin endocytosis, and megalin expression. Conversely, these effects of the S protein were replicated by LPS, an agonist of TLR4. Incubation of PTECs with a pseudovirus containing S protein inhibited albumin endocytosis. Null or VSV-G pseudovirus, used as control, had no effect. LPS-RS prevented the inhibitory impact of pseudovirus containing the S protein on albumin endocytosis but had no influence on virus internalization. Our findings demonstrate that the inhibitory effect of the S protein on albumin endocytosis in PTECs is mediated through TLR4, resulting from a reduction in megalin expression.
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Endocitose , Túbulos Renais Proximais , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Receptor 4 Toll-Like , Receptor 4 Toll-Like/metabolismo , Endocitose/efeitos dos fármacos , Humanos , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/virologia , Animais , Glicoproteína da Espícula de Coronavírus/metabolismo , SARS-CoV-2/metabolismo , Células HEK293 , Suínos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosforilação , COVID-19/metabolismo , COVID-19/virologia , COVID-19/patologia , Albuminas/metabolismo , Células LLC-PK1 , Células Epiteliais/metabolismo , Células Epiteliais/virologiaRESUMO
BACKGROUND: Diabetic kidney disease (DKD) is a severe complication of diabetes mellitus (DM). It has been proposed that modifications in the function of proximal tubule epithelial cells (PTECs) precede glomerular damage during the onset of DKD. This study aimed to identify modifications in renal sodium handling in the early stage of DM and its molecular mechanism. METHODS: Streptozotocin (STZ)-induced diabetic BALB/c mice (STZ group) and LLC-PK1 cells, a model of PTECs, were used. All parameters were assessed in the 4th week after an initial injection of STZ. RESULTS: Early stage of DKD was characterized by hyperfiltration and PTEC dysfunction. STZ group exhibited increased urinary sodium excretion due to impairment of tubular sodium reabsorption. This was correlated to a decrease in cortical (Na++K+)ATPase (NKA) α1 subunit expression and enzyme activity and an increase in O-GlcNAcylation. RNAseq analysis of patients with DKD revealed an increase in expression of the glutamine-fructose aminotransferase (GFAT) gene, a rate-limiting step of hexosamine biosynthetic pathway, and a decrease in NKA expression. Incubation of LLC-PK1 cells with 10 µM thiamet G, an inhibitor of O-GlcNAcase, reduced the expression and activity of NKA and increased O-GlcNAcylation. Furthermore, 6-diazo-5-oxo-L-norleucine (DON), a GFAT inhibitor, or dapagliflozin, an SGLT2 inhibitor, avoided the inhibitory effect of HG on expression and activity of NKA associated with the decrease in O-GlcNAcylation. CONCLUSION: Our results show that the impairment of tubular sodium reabsorption, in the early stage of DM, is due to SGLT2-mediated HG influx in PTECs, increase in O-GlcNAcylation and reduction in NKA expression and activity.
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Diabetes Mellitus , Nefropatias Diabéticas , Camundongos , Suínos , Animais , Humanos , Túbulos Renais Proximais/metabolismo , Rim/metabolismo , Nefropatias Diabéticas/metabolismo , Sódio/metabolismo , Adenosina Trifosfatases/metabolismo , Diabetes Mellitus/metabolismoRESUMO
Introduction: Rapamycin is an immunosuppressor that acts by inhibiting the serine/threonine kinase mechanistic target of rapamycin complex 1. Therapeutic use of rapamycin is limited by its adverse effects. Proteinuria is an important marker of kidney damage and a risk factor for kidney diseases progression and has been reported in patients and animal models treated with rapamycin. However, the mechanism underlying proteinuria induced by rapamycin is still an open matter. In this work, we investigated the effects of rapamycin on parameters of renal function and structure and on protein handling by proximal tubule epithelial cells (PTECs). Methods: Healthy BALB/c mice were treated with 1.5 mg/kg rapamycin by oral gavage for 1, 3, or 7 days. At the end of each treatment, the animals were kept in metabolic cages and renal function and structural parameters were analyzed. LLC-PK1 cell line was used as a model of PTECs to test specific effect of rapamycin. Results: Rapamycin treatment did not change parameters of glomerular structure and function. Conversely, there was a transient increase in 24-h proteinuria, urinary protein to creatinine ratio (UPCr), and albuminuria in the groups treated with rapamycin. In accordance with these findings, rapamycin treatment decreased albumin-fluorescein isothiocyanate uptake in the renal cortex. This effect was associated with reduced brush border expression and impaired subcellular distribution of megalin in PTECs. The effect of rapamycin seems to be specific for albumin endocytosis machinery because it did not modify renal sodium handling or (Na++K+)ATPase activity in BALB/c mice and in the LLC-PK1 cell line. A positive Pearson correlation was found between megalin expression and albumin uptake while an inverse correlation was shown between albumin uptake and UPCr or 24-h proteinuria. Despite its effect on albumin handling in PTECs, rapamycin treatment did not induce tubular injury measured by interstitial space and collagen deposition. Conclusion: These findings suggest that proteinuria induced by rapamycin could have a tubular rather than a glomerular origin. This effect involves a specific change in protein endocytosis machinery. Our results open new perspectives on understanding the undesired effect of proteinuria generated by rapamycin.
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BACKGROUND: Fanconi-Debré-de Toni syndrome (also known as Fanconi renotubular syndrome, or FRST) profoundly increased the understanding of the functions of the proximal convoluted tubule (PCT) and provided important insights into the pathophysiology of several kidney diseases and drug toxicities. DATA SOURCES: We searched Pubmed and Scopus databases to find relevant articles about FRST. This review article focuses on the physiology of the PCT, as well as on the physiopathology of FRST in children, its diagnosis, and treatment. RESULTS: FRST encompasses a wide variety of inherited and acquired PCT alterations that lead to impairment of PCT reabsorption. In children, FRST often presents as a secondary feature of systemic disorders that impair energy supply, such as Lowe's syndrome, Dent's disease, cystinosis, hereditary fructose intolerance, galactosemia, tyrosinemia, Alport syndrome, and Wilson's disease. Although rare, congenital causes of FRST greatly impact the morbidity and mortality of patients and impose diagnostic challenges. Furthermore, its treatment is diverse and considers the ability of the clinician to identify the correct etiology of the disease. CONCLUSION: The early diagnosis and treatment of pediatric patients with FRST improve the prognosis and the quality of life.
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Cistinose , Síndrome de Fanconi , Nefropatias , Síndrome Oculocerebrorrenal , Humanos , Criança , Síndrome de Fanconi/diagnóstico , Síndrome de Fanconi/genética , Síndrome de Fanconi/terapia , Qualidade de Vida , Cistinose/complicações , Síndrome Oculocerebrorrenal/complicaçõesRESUMO
Subclinical acute kidney injury (subAKI) is characterized by tubule-interstitial injury without significant changes in glomerular function. SubAKI is associated with the pathogenesis and progression of acute and chronic kidney diseases. Currently, therapeutic strategies to treat subAKI are limited. The use of gold nanoparticles (AuNPs) has shown promising benefits in different models of diseases. However, their possible effects on subAKI are still unknown. Here, we investigated the effects of AuNPs on a mouse model of subAKI. Animals with subAKI showed increased functional and histopathologic markers of tubular injury. There were no changes in glomerular function and structure. The animals with subAKI also presented an inflammatory profile demonstrated by activation of Th1 and Th17 cells in the renal cortex. This phenotype was associated with decreased megalin-mediated albumin endocytosis and expression of proximal tubular megalin. AuNP treatment prevented tubule-interstitial injury induced by subAKI. This effect was associated with a shift to an anti-inflammatory Th2 response. Furthermore, AuNP treatment preserved megalin-mediated albumin endocytosis in vivo and in vitro. AuNPs were not nephrotoxic in healthy mice. These results suggest that AuNPs have a protective effect in the tubule-interstitial injury observed in subAKI, highlighting a promising strategy as a future antiproteinuric treatment.
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Injúria Renal Aguda , Nanopartículas Metálicas , Camundongos , Animais , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Ouro/farmacologia , Túbulos Renais Proximais , Modelos Animais de Doenças , Proteinúria/metabolismo , Proteinúria/patologia , Albuminas/metabolismo , Injúria Renal Aguda/metabolismoRESUMO
Diabetic kidney disease (DKD) is characterized by progressive impairment of kidney function. It has been postulated that tubule-interstitial injury, associated with tubular albuminuria, precedes glomerular damage in the early stage of DKD. Here, we wanted to determine if the development of tubule-interstitial injury at the early stage of DKD implies modulation of megalin-mediated protein reabsorption in proximal tubule epithelial cells (PTECs) by SGLT2-dependent high glucose influx. Rats with streptozotocin (STZ)-induced diabetes were treated or not with dapagliflozin (DAPA) for 8 weeks. Four experimental groups were generated: (1) CONT, control; (2) DAPA, rats treated with DAPA; (3) STZ, diabetic rats; (4) STZ + DAPA, diabetic rats treated with DAPA. No changes in glomerular structure and function were observed. The STZ group presented proteinuria and albuminuria associated with an increase in the fractional excretion of proteins. A positive correlation between glycemia and proteinuria was found. These phenomena were linked to a decrease in luminal and total megalin expression and, consequently, in albumin reabsorption in PTECs. We also observed tubule-interstitial injury characterized by an increase in urinary tubular injury biomarkers and changes in tubular histomorphometry parameters. In addition, inverse correlations were found between cortical albumin uptake and tubule-interstitial injury or glycemia. All these modifications were attenuated in the STZ + DAPA group. These results suggest that SGLT2-dependent high glucose influx into PTECs promotes a harmful effect on the PTECs, leading to the development of tubular albuminuria and tubule-interstitial injury preceding glomerular damage. These results expand current knowledge on the renoprotective effects of gliflozins.
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Diabetes Mellitus Experimental , Nefropatias Diabéticas , Ratos , Animais , Nefropatias Diabéticas/metabolismo , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Albuminúria , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/induzido quimicamente , Transportador 2 de Glucose-Sódio/metabolismo , Proteínas/metabolismo , Albuminas/metabolismo , Glucose/efeitos adversosRESUMO
Titatinum dioxide nanoparticles (TiO2-NPs) are frequently used in several areas. Titanium alloys are employed in orthopedic and odontological surgery (such as hip, knee, and teeth implants). To evaluate the potential acute toxic effects of titanium pieces implantations and in other sources that allow the systemic delivery of titanium, parenteral routes of TiO2-NPs administration should be taken into account. The present study evaluated the impact of subcutaneous administration of TiO2-NPs on renal function and structure in rats. Animals were exposed to a dose of 50 mg/kg b.w., s.c. and sacrificed after 48 h. Titanium levels were detected in urine (135 ± 6 ηg/mL) and in renal tissue (502 ± 40 ηg/g) employing inductively coupled plasma mass spectrometry. An increase in alkaline phosphatase activity, total protein levels, and glucose concentrations was observed in urine from treated rats suggesting injury in proximal tubule cells. In parallel, histopathological studies showed tubular dilatation and cellular desquamation in these nephron segments. In summary, this study demonstrates that subcutaneous administration of TiO2-NPs causes acute nephrotoxicity evidenced by functional and histological alterations in proximal tubule cells. This fact deserves to be mainly considered when humans are exposed directly or indirectly to TiO2-NPs sources that cause the systemic delivery of titanium.
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Nanopartículas Metálicas , Nanopartículas , Humanos , Ratos , Animais , Titânio/toxicidade , Titânio/química , Nanopartículas/toxicidade , Nanopartículas Metálicas/toxicidade , Nanopartículas Metálicas/químicaRESUMO
Since the outbreak of COVID-19 disease, a bidirectional interaction between kidney disease and the progression of COVID-19 has been demonstrated. Kidney disease is an independent risk factor for mortality of patients with COVID-19 as well as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection leading to the development of acute kidney injury (AKI) and chronic kidney disease (CKD) in patients with COVID-19. However, the detection of kidney damage in patients with COVID-19 may not occur until an advanced stage based on the current clinical blood and urinary examinations. Some studies have pointed out the development of subclinical acute kidney injury (subAKI) syndrome with COVID-19. This syndrome is characterized by significant tubule interstitial injury without changes in the estimated glomerular filtration rate. Despite the complexity of the mechanism(s) underlying the development of subAKI, the involvement of changes in the protein endocytosis machinery in proximal tubule (PT) epithelial cells (PTECs) has been proposed. This paper focuses on the data relating to subAKI and COVID-19 and the role of PTECs and their protein endocytosis machinery in its pathogenesis.
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Injúria Renal Aguda , COVID-19 , Insuficiência Renal Crônica , Humanos , COVID-19/complicações , SARS-CoV-2 , Injúria Renal Aguda/metabolismo , Insuficiência Renal Crônica/metabolismo , Túbulos Renais Proximais/metabolismoRESUMO
Patients with COVID-19 have high prevalence of albuminuria which is used as a marker of progression of renal disease and is associated with severe COVID-19. We hypothesized that SARS-CoV-2 spike protein (S protein) could modulate albumin handling in proximal tubule epithelial cells (PTECs) and, consequently contribute to the albuminuria observed in patients with COVID-19. In this context, the possible effect of S protein on albumin endocytosis in PTECs was investigated. Two PTEC lines were used: HEK-293A and LLC-PK1. Incubation of both cell types with S protein for 16 h inhibited albumin uptake at the same magnitude. This effect was associated with canonical megalin-mediated albumin endocytosis because: (1) DQ-albumin uptake, a marker of the lysosomal degradation pathway, was reduced at a similar level compared with fluorescein isothiocyanate (FITC)-albumin uptake; (2) dextran-FITC uptake, a marker of fluid-phase endocytosis, was not changed; (3) cell viability and proliferation were not changed. The inhibitory effect of S protein on albumin uptake was only observed when it was added at the luminal membrane, and it did not involve the ACE2/Ang II/AT1R axis. Although both cells uptake S protein, it does not seem to be required for modulation of albumin endocytosis. The mechanism underlying the inhibition of albumin uptake by S protein encompasses a decrease in megalin expression without changes in megalin trafficking and stability. These results reveal a possible mechanism to explain the albuminuria observed in patients with COVID-19.
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COVID-19 , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Albuminas/metabolismo , Albuminas/farmacologia , Albuminúria/metabolismo , Enzima de Conversão de Angiotensina 2 , Células Cultivadas , Dextranos/farmacologia , Endocitose/fisiologia , Células Epiteliais/metabolismo , Fluoresceína-5-Isotiocianato/metabolismo , Fluoresceína-5-Isotiocianato/farmacologia , Humanos , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , SARS-CoV-2 , Glicoproteína da Espícula de CoronavírusRESUMO
Renal proximal tubule cells (PTECs) act as urine gatekeepers, constantly and efficiently avoiding urinary protein waste through receptor-mediated endocytosis. Despite its importance, little is known about how this process is modulated in physiologic conditions. Data suggest that the phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) pathway regulates PTEC protein reabsorption. Here, we worked on the hypothesis that the physiologic albumin concentration and PI3K/AKT pathway form a positive feedback loop to expand endocytic capacity. Using LLC-PK1 cells, a model of PTECs, we showed that the PI3K/AKT pathway is required for megalin recycling and surface expression, affecting albumin uptake. Inhibition of this pathway stalls megalin at EEA1+ endosomes. Physiologic albumin concentration (0.01 mg/mL) activated AKT; this depends on megalin-mediated albumin endocytosis and requires previous activation of PI3K/mTORC2. This effect is correlated to the increase in albumin endocytosis, a phenomenon that we refer to as "albumin-induced albumin endocytosis". Mice treated with L-lysine present decreased albumin endocytosis leading to proteinuria and albuminuria associated with inhibition of AKT activity. Renal cortex explants obtained from control mice treated with MK-2206 decreased albumin uptake and promoted megalin internalization. Our data highlight the mechanism behind the capacity of PTECs to adapt albumin reabsorption to physiologic fluctuations in its filtration, avoiding urinary excretion.
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Células Epiteliais/metabolismo , Retroalimentação Fisiológica , Túbulos Renais Proximais/metabolismo , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Albuminas/metabolismo , Animais , Biomarcadores , Endocitose , Células Epiteliais/efeitos dos fármacos , Imunofluorescência , Expressão Gênica , Túbulos Renais Proximais/citologia , Masculino , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Transporte Proteico , Transdução de Sinais/efeitos dos fármacosRESUMO
Hypertensive nephrosclerosis is the second most common cause of end-stage renal disease after diabetes. For years, hypertensive kidney disease has been focused on the afferent arterioles and glomeruli damage and the involvement of the renin angiotensin system (RAS). Nonetheless, in recent years, novel evidence has demonstrated that persistent high blood pressure injures tubular cells, leading to epithelial-mesenchymal transition (EMT) and tubulointerstitial fibrosis. Injury primarily determined at the glomerular level by hypertension causes changes in post-glomerular peritubular capillaries that in turn induce endothelial damage and hypoxia. Microvasculature dysfunction, by inducing hypoxic environment, triggers inflammation, EMT with epithelial cells dedifferentiation and fibrosis. Hypertensive kidney disease also includes podocyte effacement and loss, leading to disruption of the filtration barrier. This review highlights the molecular mechanisms and histologic aspects involved in the pathophysiology of hypertensive kidney disease incorporating knowledge about EMT and tubulointerstitial fibrosis. The role of the Hsp70 chaperone on the angiotensin II-induced EMT after angiotensin II type 1 receptor (AT1R) blockage, as a possible molecular target for therapeutic strategy against hypertensive renal damage is discussed.
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Proteínas de Choque Térmico HSP70/metabolismo , Hipertensão Renal/tratamento farmacológico , Rim/patologia , Losartan/uso terapêutico , Nefrite/tratamento farmacológico , Substâncias Protetoras/uso terapêutico , Animais , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Rim/efeitos dos fármacos , Losartan/farmacologia , Substâncias Protetoras/farmacologiaRESUMO
The essential oil of Croton zehntneri (EOCZ) and its major compounds are known to have several biological activities. However, some evidence shows potential toxic effects of high doses of EOCZ (>300 mg/kg) in amphibian and human kidneys. The aim of the present work was to investigate the effects on renal function of EOCZ at 300 mg/kg/day in healthy Swiss mice and a subclinical acute kidney injury (subAKI) animal model, which presents tubule-interstitial injury (TII). Four experimental groups were generated: (1) CONT group (control); (2) EOCZ, mice treated with EOCZ; (3) subAKI; (4) subAKI+EOCZ, subAKI treated simultaneously with EOCZ. EOCZ treatment induced TII measured by increases in (1) proteinuria; (2) cortical tubule-interstitial space; (3) macrophage infiltration; (4) collagen deposition. A decrease in tubular sodium reabsorption was also observed. These results were similar and nonadditive to those observed in the subAKI group. These data suggest that treatment with EOCZ at higher concentrations induces TII in mice, which could be mediated by protein overload in the proximal tubule.
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This study investigated the molecular mechanisms underlying the antiproteinuric effect of DPP4 inhibition in 5/6 renal ablation rats and tested the hypothesis that the urinary activity of DPP4 correlates with chronic kidney disease (CKD) progression. Experiments were conducted in male Wistar rats who underwent 5/6 nephrectomy (Nx) or sham operation followed by 8 wk of treatment with the DPP4 inhibitor (DPP4i) sitagliptin or vehicle. Proteinuria increased progressively in Nx rats throughout the observation period. This increase was remarkably mitigated by sitagliptin. Higher levels of proteinuria in Nx rats compared to control rats were accompanied by higher urinary excretion of retinol-binding protein 4, a marker of tubular proteinuria, as well as higher urinary levels of podocin, a marker of glomerular proteinuria. Retinol-binding protein 4 and podocin were not detected in the urine of Nx + DPP4i rats. Tubular and glomerular proteinuria was associated with the reduced expression of megalin and podocin in the renal cortex of Nx rats. Sitagliptin treatment partially prevented this decrease. Besides, the angiotensin II renal content was significantly reduced in the Nx rats that received sitagliptin compared to vehicle-treated Nx rats. Interestingly, both urinary DPP4 activity and abundance increased progressively in Nx rats. Additionally, urinary DPP4 activity correlated positively with serum creatinine levels, proteinuria, and blood pressure. Collectively, these results suggest that DPP4 inhibition ameliorated both tubular and glomerular proteinuria and prevented the reduction of megalin and podocin expression in CKD rats. Furthermore, these findings suggest that urinary DPP4 activity may serve as a biomarker of renal disease and progression.
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Dipeptidil Peptidase 4/urina , Inibidores da Dipeptidil Peptidase IV/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Rim/efeitos dos fármacos , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteínas de Membrana/metabolismo , Proteinúria/prevenção & controle , Insuficiência Renal Crônica/prevenção & controle , Fosfato de Sitagliptina/farmacologia , Angiotensina II/metabolismo , Animais , Biomarcadores/urina , Modelos Animais de Doenças , Fibrose , Rim/enzimologia , Rim/patologia , Masculino , Proteinúria/enzimologia , Proteinúria/patologia , Proteinúria/urina , Ratos Wistar , Insuficiência Renal Crônica/enzimologia , Insuficiência Renal Crônica/patologia , Insuficiência Renal Crônica/urina , Proteínas Plasmáticas de Ligação ao Retinol/urina , Transdução de SinaisRESUMO
Angiotensin II exerts a cardinal role in the pathogenesis of hypertension and renal injury via action of angiotensin II type 1 (AT1) receptors. Local renin-angiotensin system (RAS) activity is essential for the mechanisms mediating pathophysiological functions. Proximal tubular angiotensinogen and tubular AT1 receptors are augmented by intrarenal angiotensin II. Caveolin 1 plays an important role as a regulatory molecule for the compartmentalization of redox signaling events through angiotensin II-induced NADPH oxidase activation in the kidney. A role for the renin-angiotensin system in the development and/or maintenance of hypertension has been demonstrated in spontaneously hypertensive rats (SHRs). Many effects of angiotensin II are dependent on the AT1 stimulation of reactive oxygen species (ROS) production by NADPH oxidase. Angiotensin II upregulation stimulates oxidative stress in proximal tubules from SHR. The NADPH oxidase 4 (Nox4) is abundantly expressed in kidney proximal tubule cells. Induction of the stress response includes synthesis of heat shock protein 70, a molecular chaperone that has a critical role in the recovery of cells from stress and in cytoprotection, guarding cells from subsequent insults. HSP70 chaperones function in part by driving the molecular triage decision, which determines whether proteins enter the productive folding pathway or result in client substrate ubiquitination and proteasomal degradation. This review examines regulation of losartan-mediated antioxidative stress responses by the chaperone HSP70 in proximal tubule cells of spontaneously hypertensive rats.
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Injúria Renal Aguda/tratamento farmacológico , Antioxidantes/farmacologia , Proteínas de Choque Térmico HSP70/metabolismo , Hipertensão/tratamento farmacológico , Losartan/farmacologia , Angiotensina II/metabolismo , Animais , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , NADPH Oxidase 4/metabolismo , Ratos , Ratos Endogâmicos SHRRESUMO
Polyuria is a hallmark symptom and the first clinical manifestation of diabetes mellitus (DM). The glucose that remains in renal tubules was proposed to produce an osmotic effect resulting in polyuria. Although water is reabsorbed in proximal tubules through an aquaporin-1 (AQP1) dependent mechanism, AQP1 role in the genesis of polyuria is unknown. AQP1 expression was studied in a rat model of Type-1 DM at 15-days and 5-months of evolution. A different AQP1 expression pattern was found in both experimental groups, with no changes in AQP1 localization, suggesting that changes in AQP1 may be involved in the development of polyuria.
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BACKGROUND/AIMS: Renal injury related to hypertension is characterized by glomerular and tubulointerstitial damage. The overactivation of the renin-angiotensin system mainly by angiotensin II (AII) seems to be a main contributor to progressive renal fibrosis. Epithelial to mesenchymal transition (EMT) is a mechanism that promotes renal fibrosis. Owing to heat shock protein 70 (Hsp70) cytoprotective properties, the chaperone exhibits an important potential as a therapeutic target. We investigate the role of Hsp70 on Angiotensin II induced epithelial mesenchymal transition within the Losartan effect in proximal tubule cells (PTCs) from a genetic model of hypertension in rats (SHR). METHODS: Primary cell culture of PTCs from SHR and Wistar Kyoto (WKY) rats were stimulated with AII, treated with Losartan (L), (L+AII) or untreated (Cc). The functional Hsp70 role in Losartan effect, after silencing its expression by cell transfection, was determined by Immunofluorescence; Western blotting; Gelatin Zymography assays; Scratch wound assays; flow cytometry; and Live Cell Time-lapse microscopy. RESULTS: (L) and (L+AII) treatments induced highly organized actin filaments and increased cortical actin in SHR PTCs. However, SHR PTCs (Cc) and (AII) treated cells showed disorganized actin. After Hsp72 knockdown in SHR PTCs, (L) was unable to stabilize the actin cytoskeleton. We demonstrated that (L) and (L+AII) increased E-cadherin levels and decreased vinculin, α-SMA, vimentin, pERK, p38 and Smad2-3 activation compared to (AII) and (Cc) SHR PTCs. Moreover, (L) inhibited MMP-2 and MMP-9 secretion, reduced migration and cellular displacement, stabilizing intercellular junctions. Notably, (L) treatment in shHsp72 knockdown SHR PTCs showed results similar to SHR PTCs (Cc). CONCLUSION: Our results demonstrate that Losartan through Hsp70 inhibits the EMT induced by AII in proximal tubule cells derived from SHR.
Assuntos
Angiotensina II/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Proteínas de Choque Térmico HSP70/metabolismo , Losartan/farmacologia , Citoesqueleto de Actina/efeitos dos fármacos , Animais , Caderinas/metabolismo , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Adesões Focais/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Proteínas de Choque Térmico HSP70/genética , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Vinculina/metabolismoRESUMO
Caveolin-2 (Cav-2) is expressed in a variety of cell tissue, and it has also been found in renal tissue. The expression of Cav-2 in proximal tubules is still unclear. The aim of this study was to carry out a complete evaluation of the expression pattern of Cav-2 in rat renal cortex to clarify and deepen the knowledge about the localization of Cav-2 in the proximal tubules and also to evaluate its presence in urine. Male Wistar rats were used to assess Cav-2 expression by Western blot analysis in homogenates, apical, and basolateral membranes from kidney cortex, in lysates and total plasma membranes from renal cortical cell suspensions, in urine, and in urinary exosomes. Cav-2 was clearly expressed in renal cortex homogenates and in both apical and basolateral membranes isolated from kidney cortex, with a greater expression on the former membranes. It was also observed in lysates and in plasma membranes from cortical cell suspensions. Moreover, Cav-2 was found in urine and in its exosomal fraction. These results confirmed the presence of Cav-2 in proximal tubule cells in the kidney of healthy rats, and showed for the first time its expression at the apical membrane of these cells and in urine. Besides, urinary exosomal pathway could be involved in Cav-2 urinary excretion under normal conditions. We observed an increase in the urinary abundance of Cav-2 in two models of acute kidney injury, and thus we proposed the urinary excretion of Cav-2 as a potential biomarker of kidney injury.
Assuntos
Injúria Renal Aguda/urina , Caveolina 2/urina , Membrana Celular/genética , Exossomos/metabolismo , Túbulos Renais Proximais/metabolismo , Injúria Renal Aguda/patologia , Animais , Biomarcadores/urina , Membrana Celular/patologia , Exossomos/patologia , Túbulos Renais Proximais/patologia , Masculino , Ratos , Ratos WistarRESUMO
BACKGROUND: The calcium-sensing receptor (CaSR) is localized in the apical membrane of proximal tubules in close proximity to the transporters responsible for proton secretion. Therefore, the aim of the present study was to analyze the effects of CaSR stimulation on the biochemical activity of the vacuolar H+-ATPase in a cellular model of proximal tubule cells, OKP cells. METHODS: Biochemical activity of H+-ATPase was performed using cell homogenates, and the inorganic phosphate released was determined by a colorimetric method. Changes in cytosolic ionized calcium [Ca2+]i were also determined using Fluo-4. RESULTS: A significant increase of vacuolar H+-ATPase activity was observed when the CaSR was stimulated with agonists such as Gd3+ (300 µM) and neomycin (200 µM). This activity was also stimulated in a dose-dependent fashion by changes in extracellular Ca2+ (Ca2+o) between 10-4 and 2 mM. Gd3+ and neomycin produced a sustained rise of [Ca2+]i, an effect that disappears when extracellular calcium was removed in the presence of 0.1 µM thapsigargin. Inhibition of phospholipase C (PLC) activity with U73122 (5 × 10-8 M) reduced the increase in [Ca2+]i induced by neomycin. CONCLUSION: CaSR stimulation induces an increase in the vacuolar H+-ATPase activity of OKP cells, an effect that involves an increase in [Ca2+]i and require phospholipase C activity. The consequent decrease in intratubular pH could lead to increase ionization of luminal calcium, potentially enhancing its reabsorption in distal tubule segments and reducing the formation of calcium phosphate stones.
Assuntos
Túbulos Renais Proximais/metabolismo , Receptores de Detecção de Cálcio/fisiologia , ATPases Vacuolares Próton-Translocadoras/metabolismo , Animais , Cálcio/metabolismo , Células Cultivadas , Concentração de Íons de Hidrogênio , Neomicina/farmacologia , Gambás , Fosfolipases Tipo C/metabolismoRESUMO
Diabetic nephropathy (DN) occurs in around 40% of those with diabetes. Proteinuria is the main characteristic of DN and develops as a result of increased permeability of the glomerulus capillary wall and/or decreased proximal tubule endocytosis. The goal of this work was to evaluate renal function and the expression of megalin, cubilin, CFTR (cystic fibrosis transmembrane conductance regulator), and ClC-5 in the proximal tubule and renal cortex of rats with type 1 diabetes. Male Wistar rats were randomly assigned to control (CTRL) and diabetic (DM) groups for 4 weeks. Renal function was assessed in 24-h urine sample by calculating clearance and fractional excretion of solutes. The RNA and protein contents of ClC-5, CFTR, megalin, and cubilin were determined in the renal proximal tubule and cortex using real-time polymerase chain reaction and western blotting techniques, respectively. The results showed higher creatinine clearance and higher urinary excretion of proteins, albumin, and transferrin in the DM group than in the CTRL group. Furthermore, the renal cortex and proximal tubule of diabetic animals showed downregulation of megalin, cubilin, ClC-5, and CFTR, critical components of the endocytic apparatus. These data suggest dysfunction in proximal tubule low-molecular-weight endocytosis and protein glomerulus filtration in the kidney of diabetic rats.