RESUMO
INTRODUCTION: Several studies have reported kidney injury caused by immune checkpoint inhibitors, and proteinuria caused by vascular endothelial growth factor inhibitors for unresectable hepatocellular carcinoma (u-HCC). We investigated the relationship between renal function and prognosis in patients with u-HCC receiving atezolizumab and bevacizumab (AB) and lenvatinib (LEN) therapy. METHODS: Fifty-one patients who received AB and 50 patients who received LEN therapy were included. We analyzed prognostic factors related to the overall survival (OS), and characteristics related to renal function. RESULTS: In patients with AB therapy, OS was shorter in patients with baseline proteinuria of 1+ or higher, as assessed by urine dipstick test, compared to those with -/± (p = 0.024). There were many cases with two or more drugs with a high risk of renal dysfunction (p = 0.019) in patients with 1+ or higher. Furthermore, OS was shorter in the group with estimated glomerular filtration rate (eGFR) grade deterioration without urinary protein-creatinine ratio (UPCR) of 2 g/g·Cre or higher than in the other groups (p = 0.027). In the group where eGFR worsened without an increase in UPCR, there were many cases with a daily salt intake of 10 g or more (p = 0.027), three or more drugs with a high risk of renal dysfunction (p = 0.021), and a history of arteriosclerosis (p = 0.021). On the other hand, in patients with LEN therapy, OS tends to be shorter in patients with proteinuria of ± or higher, compared to those without (p = 0.074). There were many cases with a daily salt intake of 10 g or more in patients with ± or higher (p = 0.002). CONCLUSION: In patients receiving AB and LEN therapy, baseline proteinuria was associated with OS. Renal function deterioration without proteinuria was associated with a poor prognosis in AB therapy. Excessive salt intake, preexisting atherosclerotic disease, and drug with a high risk of renal dysfunction were risk factors for renal deterioration.
Assuntos
Carcinoma Hepatocelular , Nefropatias , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/tratamento farmacológico , Bevacizumab/efeitos adversos , Cloreto de Sódio na Dieta , Fator A de Crescimento do Endotélio Vascular , Neoplasias Hepáticas/tratamento farmacológico , Prognóstico , Rim/fisiologiaRESUMO
Since 2015, Japanese researchers have made great progress in developing a method to differentiate human pluripotent stem cells (hPSCs) into kidney organoids. Protocols have been established to produce increasingly complex three-dimensional (3D) structures, which are used as a human kidney disease model and adapted for high-throughput screening. During this period, single-cell RNA sequencing (scRNA-seq) technology was developed to perform a comprehensive analysis at the single-cell level. We have performed a comprehensive analysis using scRNA-seq to define how kidney organoids can be applied to understand kidney development and pathology. The structure of kidney organoids is complex and contains many cell types of varying maturity. Since only a few proteins and mRNAs can be identified by immunostaining and other techniques, we performed scRNA-seq, which is an unbiased technology that can comprehensively categorize all cell types present in organoids. The aim of this study is to review the problems of kidney organoids based on scRNA-seq and the efforts to address the problems and predict future applications with this powerful technique.
Assuntos
Nefropatias , Rim , Humanos , Diferenciação Celular , Nefropatias/genética , Organoides , Análise de Sequência de RNARESUMO
After acute kidney injury (AKI), patients either recover or alternatively develop fibrosis and chronic kidney disease. Interactions between injured epithelia, stroma, and inflammatory cells determine whether kidneys repair or undergo fibrosis, but the molecular events that drive these processes are poorly understood. Here, we use single nucleus RNA sequencing of a mouse model of AKI to characterize cell states during repair from acute injury. We identify a distinct proinflammatory and profibrotic proximal tubule cell state that fails to repair. Deconvolution of bulk RNA-seq datasets indicates that this failed-repair proximal tubule cell (FR-PTC) state can be detected in other models of kidney injury, increasing during aging in rat kidney and over time in human kidney allografts. We also describe dynamic intercellular communication networks and discern transcriptional pathways driving successful vs. failed repair. Our study provides a detailed description of cellular responses after injury and suggests that the FR-PTC state may represent a therapeutic target to improve repair.
Assuntos
Injúria Renal Aguda/metabolismo , Túbulos Renais Proximais/metabolismo , Rim/metabolismo , Transcriptoma , Injúria Renal Aguda/genética , Injúria Renal Aguda/patologia , Aloenxertos , Animais , Modelos Animais de Doenças , Fibrose , Redes Reguladoras de Genes , Humanos , Rim/lesões , Túbulos Renais Proximais/lesões , Túbulos Renais Proximais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Análise de Sequência de RNA , Células Estromais/metabolismo , Células Estromais/patologiaRESUMO
Thyroid hormones are critical regulators of vertebrate development and metabolism. Under hyperthyroid conditions, excess thyroid hormones induce expression of several enzymes and activities via activation of ligand-bound thyroid hormone receptors (TRs). Arginase (ARG) is downstream of a ligand-bound TR and overexpression of ARG2 induces the production of reactive oxygen species and subsequent exacerbation of kidney ischemia/reperfusion (I/R) injury. To clarify the association between I/R-induced kidney injury and hyperthyroidism, mice were pretreated with L-thyroxine (LT4) or vehicle alone, then subjected to I/R. Proximal tubular cell-specific conditional knockout of thyroid hormone receptor ß (TRßcKO) mice was generated and the effects of I/R were analyzed. Hyperthyroidism enhanced tubular damage and fibrosis in the kidneys of mice after I/R. Hyperthyroidism induced tubular cell necroptosis following inflammatory cell accumulation in the kidney after I/R. ARG2 expressions and reactive oxygen species accumulated in the kidneys of hyperthyroid mice after I/R, but these changes were ameliorated in the kidneys of TRßcKO mice. Hyperthyroidism-enhanced kidney injury was ameliorated in the kidney of TRßcKO mice after I/R. These results suggest that excess thyroid hormones are disadvantageous for the kidney under ischemic stress. Overt hypothyroidism represents a severe thyroid hormone deficiency disease that requires LT4 treatment, while overreplacement or iatrogenic thyrotoxicosis might cause kidney injury.
Assuntos
Hipertireoidismo , Hipotireoidismo , Traumatismo por Reperfusão , Animais , Hipertireoidismo/complicações , Hipertireoidismo/genética , Hipotireoidismo/metabolismo , Rim/metabolismo , Camundongos , Traumatismo por Reperfusão/complicações , Traumatismo por Reperfusão/metabolismo , Hormônios Tireóideos/metabolismoRESUMO
Inteletin1 (Itln1) is an adipokine that is abundantly expressed in intestine, ovary, and lung. The expression levels of ITLN1 are decreased in the presence of diabetes or obesity, but the mechanisms of its production and function are still controversial. The aim of this study is to elucidate the mechanisms of ITLN1 synthesis and ITLN1-associated macrophage activation. To analyze the effects of high fat and high-carbohydrate diet (HFHCD) on the expression of ITLN1 in the intestine, the mice were fed a HFHCD for 8 weeks. HFHCD feeding enhanced the endoplasmic reticulum (ER)-stress in the intestine and inhibited the expression of Itln1 in the intestinal endocrine cells and lowered circulating ITLN1 levels. In contrast, treatment with a chemical chaperone and reduction of ER-stress restored the expression of Itln1 in the intestine of HFHCD-fed mice. Furthermore, in vitro studies indicated that ITLN1 physically interacts with adiponectin receptor 1 and suppresses lipopolysaccharide-induced mRNA expressions of pro-inflammatory cytokines and phagocytosis activities via inhibition of the nuclear factor kappa B-signaling pathway in macrophages. These results suggest that diet-induced ER-stress decreases circulating ITLN1 via inhibition of its synthesis in the intestine, and a reduction of circulating ITLN1 might enhanced the expression of proinflammatory cytokines and macrophage activation, following exacerbate the chronic inflammation of metabolic syndrome.
Assuntos
Citocinas/metabolismo , Proteínas Ligadas por GPI/metabolismo , Lectinas/metabolismo , Ativação de Macrófagos , NF-kappa B , Animais , Estresse do Retículo Endoplasmático , Feminino , Inflamação , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Transdução de SinaisRESUMO
Diabetic nephropathy is characterized by damage to both the glomerulus and tubulointerstitium, but relatively little is known about accompanying cell-specific changes in gene expression. We performed unbiased single-nucleus RNA sequencing (snRNA-seq) on cryopreserved human diabetic kidney samples to generate 23,980 single-nucleus transcriptomes from 3 control and 3 early diabetic nephropathy samples. All major cell types of the kidney were represented in the final dataset. Side-by-side comparison demonstrated cell-type-specific changes in gene expression that are important for ion transport, angiogenesis, and immune cell activation. In particular, we show that the diabetic thick ascending limb, late distal convoluted tubule, and principal cells all adopt a gene expression signature consistent with increased potassium secretion, including alterations in Na+/K+-ATPase, WNK1, mineralocorticoid receptor, and NEDD4L expression, as well as decreased paracellular calcium and magnesium reabsorption. We also identify strong angiogenic signatures in glomerular cell types, proximal convoluted tubule, distal convoluted tubule, and principal cells. Taken together, these results suggest that increased potassium secretion and angiogenic signaling represent early kidney responses in human diabetic nephropathy.
Assuntos
Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Idoso , Cálcio/metabolismo , Cálcio/urina , Diabetes Mellitus/metabolismo , Nefropatias Diabéticas/fisiopatologia , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Rim/metabolismo , Glomérulos Renais/metabolismo , Túbulos Renais Distais/metabolismo , Túbulos Renais Proximais/metabolismo , Magnésio/metabolismo , Magnésio/urina , Masculino , Pessoa de Meia-Idade , Potássio/metabolismo , Potássio/urina , Análise de Sequência de RNA , Análise de Célula Única/métodos , Transcriptoma/genéticaRESUMO
Metabolic syndrome (MetS) is associated with chronic kidney disease and proteinuria. Previously, we reported that a synthetic serine protease inhibitor, camostat mesilate (CM), mitigated hypertension and proteinuria in rodent disease models. The present study evaluated the anti-hypertensive and anti-proteinuric effects of CM in MetS model rats (SHR/ND mcr-cp). Rats were divided into normal salt-fed (NS), high salt-fed (HS), HS and CM-treated (CM), and HS and hydralazine-treated (Hyd) groups. Rats were sacrificed after four weeks of treatment. Severe hypertension and proteinuria were observed in the HS group. Although CM and Hyd equally alleviated hypertension, CM suppressed proteinuria and glomerular sclerosis more efficiently than Hyd. The HS group revealed a decrease in podocyte number and podocyte-specific molecules, together with an increase in glomerular apoptotic cells and apoptosis-related proteins in the kidney. These changes were significantly attenuated by CM, but not by Hyd. Furthermore, CM ameliorated the apoptotic signals in murine cultured podocytes stimulated with the high glucose and aldosterone medium. In conclusion, CM could exert renoprotective effects in MetS model rats, together with the inhibition of podocyte apoptosis. Our study suggests that serine protease inhibition may become a new therapeutic strategy against MetS-related hypertension and renal injuries.
Assuntos
Apoptose/efeitos dos fármacos , Ésteres/farmacologia , Guanidinas/farmacologia , Síndrome Metabólica/patologia , Podócitos/patologia , Inibidores de Proteases/farmacologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Hipertensão/tratamento farmacológico , Hipertensão/etiologia , Masculino , Síndrome Metabólica/complicações , Camundongos , Proteinúria/tratamento farmacológico , Proteinúria/etiologia , Ratos Endogâmicos SHR , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/etiologiaRESUMO
Hypoxia occurs in the kidneys of chronic kidney disease (CKD) patients, inducing interstitial fibrosis and tubule cell death. Renal tubule cell death is an important determinant of mortality in CKD. We focused on the regulation of cell-cycle-mediated protein expression to prevent cell death under chronic hypoxia in the kidneys of CKD patients. Paraffin-embedded kidney sections from patients with CKD (diabetes nephropathy, nephrosclerosis, or IgA nephropathy) were analyzed for the expression of hypoxia-inducible factor (HIF), thyroid hormone receptor (TR) ß, or p21 and levels of interstitial fibrosis. Human renal proximal tubule cells were exposed to hypoxia and analyzed for the expression of HIF, TRß, or p21 and the cell-cycle stage. TRß expression was enhanced early on when fibrosis was not fully developed in the tubule cells of CKD patients. HIF1α bound to the TRß promoter and directly induced its transcription. Further, HIF1α expression induced the expression of TRß and inhibited cell-cycle progression. In the early stage of kidney injury, TRß might act as a guardian to prepare and organize cell-cycle proliferation and prevent cell death. While the molecular mechanism that regulates the expression of cell-cycle regulators in renal tubule cells remains controversial, TRß has strong potential as a new therapeutic target.
Assuntos
Ciclo Celular/fisiologia , Células Epiteliais/metabolismo , Hipóxia/metabolismo , Túbulos Renais Proximais/patologia , Receptores dos Hormônios Tireóideos/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular , Proliferação de Células , Células Epiteliais/patologia , Feminino , Humanos , Hipóxia/genética , Hipóxia/patologia , Túbulos Renais Proximais/metabolismo , Masculino , Pessoa de Meia-Idade , Receptores dos Hormônios Tireóideos/genética , Adulto JovemRESUMO
The liver has a most indispensable role in glucose and lipid metabolism where we see some of the most serious worldwide health problems. The serine protease prostasin (PRSS8) cleaves toll-like receptor 4 (TLR4) and regulates hepatic insulin sensitivity under PRSS8 knockout condition. However, liver substrate proteins of PRSS8 other than TLR4 and the effect to glucose and lipid metabolism remain unclarified with hepatic elevation of PRSS8 expression. Here we show that high-fat-diet-fed liver-specific PRSS8 transgenic mice improved glucose tolerance and hepatic steatosis independent of body weight. PRSS8 amplified extracellular signal-regulated kinase phosphorylation associated with matrix metalloproteinase 14 activation in vivo and in vitro. Moreover, in humans, serum PRSS8 levels reduced more in type 2 diabetes mellitus (T2DM) patients than healthy controls and were lower in T2DM patients with increased maximum carotid artery intima media thickness (>1.1 mm). These results identify the regulatory mechanisms of PRSS8 overexpression over glucose and lipid metabolism, as well as excessive hepatic fat storage.
Assuntos
Diabetes Mellitus Tipo 2/patologia , Dieta Hiperlipídica/efeitos adversos , Fígado Gorduroso/patologia , Serina Endopeptidases/metabolismo , Animais , Peso Corporal , Estudos de Casos e Controles , Diabetes Mellitus Tipo 2/metabolismo , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Feminino , Metabolismo dos Lipídeos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Serina Endopeptidases/genéticaRESUMO
BACKGROUND: Patients undergoing hemodialysis treatment have a poor prognosis, as many develop premature aging. Systemic inflammatory conditions often underlie premature aging phenotypes in uremic patients. We investigated whether angiopoietin-like protein 2 (ANGPTL 2), a factor that accelerates the progression of aging-related and noninfectious inflammatory diseases, was associated with increased mortality risk in hemodialysis patients. METHODS: We conducted a multicenter prospective cohort study of 412 patients receiving maintenance hemodialysis and evaluated the relationship between circulating ANGPTL2 levels and the risk for all-cause mortality. Circulating ANGPTL2 levels were log-transformed to correct for skewed distribution and analyzed as a continuous variable. RESULTS: Of 412 patients, 395 were included for statistical analysis. Time-to-event data analysis showed high circulating ANGPTL2 levels were associated with an increased risk for all-cause mortality after adjustment for age, sex, hemodialysis vintage, nutritional status, metabolic parameters and circulating high-sensitivity C-reactive protein levels {hazard ratio [HR] 2.04 [95% confidence interval (CI) 1.10-3.77]}. High circulating ANGPTL2 levels were also strongly associated with an increased mortality risk, particularly in patients with a relatively benign prognostic profile [HR 3.06 (95% CI 1.86-5.03)]. Furthermore, the relationship between circulating ANGPTL2 levels and mortality risk was particularly strong in patients showing few aging-related phenotypes, such as younger patients [HR 7.99 (95% CI 3.55-18.01)], patients with a short hemodialysis vintage [HR 3.99 (95% CI 2.85-5.58)] and nondiabetic patients [HR 5.15 (95% CI 3.19-8.32)]. CONCLUSION: We conclude that circulating ANGPTL2 levels are positively associated with mortality risk in patients receiving maintenance hemodialysis and that ANGPTL2 could be a unique marker for the progression of premature aging and subsequent mortality risk in uremic patients, except those with significant aging-related phenotypes.
Assuntos
Proteínas Semelhantes a Angiopoietina/sangue , Biomarcadores/sangue , Nefropatias/mortalidade , Diálise Renal/mortalidade , Idoso , Proteína 2 Semelhante a Angiopoietina , Proteína C-Reativa/análise , Progressão da Doença , Feminino , Humanos , Nefropatias/sangue , Nefropatias/terapia , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Fatores de Risco , Taxa de SobrevidaRESUMO
Background Single-cell genomics techniques are revolutionizing our ability to characterize complex tissues. By contrast, the techniques used to analyze renal biopsy specimens have changed little over several decades. We tested the hypothesis that single-cell RNA-sequencing can comprehensively describe cell types and states in a human kidney biopsy specimen.Methods We generated 8746 single-cell transcriptomes from a healthy adult kidney and a single kidney transplant biopsy core by single-cell RNA-sequencing. Unsupervised clustering analysis of the biopsy specimen was performed to identify 16 distinct cell types, including all of the major immune cell types and most native kidney cell types, in this biopsy specimen, for which the histologic read was mixed rejection.Results Monocytes formed two subclusters representing a nonclassical CD16+ group and a classic CD16- group expressing dendritic cell maturation markers. The presence of both monocyte cell subtypes was validated by staining of independent transplant biopsy specimens. Comparison of healthy kidney epithelial transcriptomes with biopsy specimen counterparts identified novel segment-specific proinflammatory responses in rejection. Endothelial cells formed three distinct subclusters: resting cells and two activated endothelial cell groups. One activated endothelial cell group expressed Fc receptor pathway activation and Ig internalization genes, consistent with the pathologic diagnosis of antibody-mediated rejection. We mapped previously defined genes that associate with rejection outcomes to single cell types and generated a searchable online gene expression database.Conclusions We present the first step toward incorporation of single-cell transcriptomics into kidney biopsy specimen interpretation, describe a heterogeneous immune response in mixed rejection, and provide a searchable resource for the scientific community.
Assuntos
Falência Renal Crônica/genética , Transplante de Rim/métodos , Rim/citologia , Rim/patologia , Transcriptoma/genética , Aloenxertos , Biomarcadores/análise , Biópsia por Agulha , Comunicação Celular , Análise por Conglomerados , Perfilação da Expressão Gênica/métodos , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Humanos , Imuno-Histoquímica , Falência Renal Crônica/patologia , Falência Renal Crônica/cirurgia , Masculino , Valores de Referência , Análise de Sequência de RNA , Adulto JovemRESUMO
Emerging evidence has suggested that aldosterone has direct deleterious effects on the kidney independently of its hemodynamic effects. However, the detailed mechanisms of these direct effects remain to be elucidated. We have previously reported that camostat mesilate (CM), a synthetic serine protease inhibitor, attenuated kidney injuries in Dahl salt-sensitive rats, remnant kidney rats, and unilateral ureteral obstruction rats, suggesting that some serine proteases would be involved in the pathogenesis of kidney injuries. The current study was conducted to investigate the roles of serine proteases and the beneficial effects of CM in aldosterone-related kidney injuries. We observed a serine protease that was activated by aldosterone/salt in rat kidney lysate, and identified it as plasmin with liquid chromatography-tandem mass spectrometry. Plasmin increased pro-fibrotic and inflammatory gene expressions in rat renal fibroblast cells. CM inhibited the protease activity of plasmin and suppressed cell injury markers induced by plasmin in the fibroblast cells. Furthermore, CM ameliorated glomerulosclerosis and interstitial fibrosis in the kidney of aldosterone/salt-treated rats. Our findings indicate that plasmin has important roles in kidney injuries that are induced by aldosterone/salt, and that serine protease inhibitor could provide a new strategy for the treatment of aldosterone-associated kidney diseases in humans.
Assuntos
Injúria Renal Aguda/tratamento farmacológico , Injúria Renal Aguda/metabolismo , Aldosterona/toxicidade , Antifibrinolíticos/uso terapêutico , Fibrinolisina/metabolismo , Inibidores de Serina Proteinase/uso terapêutico , Injúria Renal Aguda/induzido quimicamente , Animais , Antifibrinolíticos/farmacologia , Fibrinolisina/antagonistas & inibidores , Masculino , Ratos , Ratos Sprague-Dawley , Inibidores de Serina Proteinase/farmacologiaRESUMO
We previously reported that camostat mesilate (CM) had renoprotective and antihypertensive effects in rat CKD models. In this study, we examined if CM has a distinct renoprotective effect from telmisartan (TE), a renin-angiotensin-aldosterone system (RAS) inhibitor, on the progression of CKD. We evaluated the effect of CM (400 mg/kg/day) and/or TE (10 mg/kg/day) on renal function, oxidative stress, renal fibrosis, and RAS components in the adenine-induced rat CKD model following 5-weeks treatment period. The combination therapy with CM and TE significantly decreased the adenine-induced increase in serum creatinine levels compared with each monotherapy, although all treatment groups showed similar reduction in blood pressure. Similarly, adenine-induced elevation in oxidative stress markers and renal fibrosis markers were significantly reduced by the combination therapy relative to each monotherapy. Furthermore, the effect of the combination therapy on plasma renin activity (PRA) and plasma aldosterone concentration (PAC) was similar to that of TE monotherapy, and CM had no effect on both PRA and PAC, suggesting that CM has a distinct pharmacological property from RAS inhibition. Our findings indicate that CM could be a candidate drug for an add-on therapy for CKD patients who had been treated with RAS inhibitors.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/administração & dosagem , Anti-Hipertensivos/administração & dosagem , Benzimidazóis/administração & dosagem , Benzoatos/administração & dosagem , Gabexato/análogos & derivados , Insuficiência Renal Crônica/tratamento farmacológico , Inibidores de Serina Proteinase/administração & dosagem , Aldosterona/sangue , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Anti-Hipertensivos/farmacologia , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Creatinina/sangue , Modelos Animais de Doenças , Quimioterapia Combinada , Ésteres , Fibrose/tratamento farmacológico , Gabexato/administração & dosagem , Gabexato/farmacologia , Guanidinas , Rim/patologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Ratos Sprague-Dawley , Renina/sangue , Sistema Renina-Angiotensina/efeitos dos fármacos , Inibidores de Serina Proteinase/farmacologia , TelmisartanRESUMO
Serine proteases play pivotal roles in many biological processes including clotting, digestion, and immune system. A glycosylphosphatidylinositol-anchored serine protease prostasin(PRSS8) is ubiquitously expressed in many tissues such as kidney, prostate, skin, liver, lung, and colon. However, the physiological role for PRSS8 has not been fully understood. Recently, we have identified a novel role for PRSS8 in the regulation of glucose homeostasis via Toll-like receptor 4(TLR4)-mediated signaling in the liver and demonstrated new insight into the development of diabetes resulting from obesity-induced inflammation or metabolic endotoxemia. In this review, we provide an overview of current understanding of PRSS8 as a new pathological factor for human diseases.
Assuntos
Serina Endopeptidases/fisiologia , Animais , Diabetes Mellitus Tipo 2/etiologia , Canais Epiteliais de Sódio , Humanos , Inflamação/fisiopatologia , Fígado/metabolismo , CamundongosRESUMO
Interstitial fibrosis is a final common pathway for the progression of chronic kidney diseases. Activated fibroblasts have an extremely important role in the progression of renal fibrosis, and transforming growth factor (TGF)-ß1 is a major activator of fibroblasts. Since previous reports have indicated that serine protease inhibitors have a potential to inhibit TGF-ß1 signaling in vitro, we hypothesized that a synthetic serine protease inhibitor, camostat mesilate (CM), could slow the progression of renal fibrosis. TGF-ß1 markedly increased the phosphorylation of TGF-ß type I receptor, ERK 1/2, and Smad2/3 and the levels of profibrotic markers, such as α-smooth muscle actin (α-SMA), connective tissue growth factor (CTGF), and plasminogen activator inhibitor-1, in renal fibroblasts (NRK-49F cells), and they were all significantly reduced by CM. In protocol 1, 8-wk-old male Sprague-Dawley rats were subjected to unilateral ureteral obstruction (UUO) and were concurrently treated with a slow-release pellet of CM or vehicle for 14 days. Protocol 2 was similar to protocol 1 except that CM was administered 7 days after UUO. CM substantially improved renal fibrosis as determined by sirius red staining, collagen expression, and hydroxyproline levels. The phosphorylation of ERK1/2 and Smad2/3 and the levels of α-SMA, CTGF, promatrix metalloproteinase-2, and matrix metalloproteinase-2 were substantially increased by UUO, and they were all significantly attenuated by CM. These antifibrotic effects of CM were also observed in protocol 2. Our present results suggest the possibility that CM might represent a new class of therapeutic drugs for the treatment of renal fibrosis through the suppression of TGF-ß1 signaling.
Assuntos
Fibroblastos/metabolismo , Gabexato/análogos & derivados , Nefroesclerose/terapia , Inibidores de Serina Proteinase/uso terapêutico , Fator de Crescimento Transformador beta1/metabolismo , Animais , Linhagem Celular , Quimiocina CCL2/metabolismo , Ésteres , Gabexato/farmacologia , Gabexato/uso terapêutico , Guanidinas , Macrófagos/efeitos dos fármacos , Nefroesclerose/etiologia , Nefroesclerose/metabolismo , Fosforilação/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Inibidores de Serina Proteinase/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Obstrução Ureteral/complicaçõesRESUMO
Prostasin (PRSS8) is a serine protease that metabolizes and moderates the effect of specific substrates. Epidermal growth factor receptor (EGFR), which modulates insulin secretion and pancreatic ß-cell proliferation, is regulated via proteolytic shedding by PRSS8. We first detected PRSS8 expression in ß-cells of pancreatic islets of mice. To better understand the molecular processes involved in PRSS8-associated insulin secretion, pancreatic ß-cell-specific PRSS8 knockout (ßKO) and PRSS8-overexpressing (ßTG) male mice were generated. We found that glucose intolerance and reduction in glucose-stimulated insulin secretion developed in ßKO mice compared with the control subjects. A higher response to glucose was noted in islets retrieved from ßTG mice. Erlotinib, a specific blocker of EGFR, blocks EGF- and glucose-stimulated secretion of insulin among MIN6 cells, and glucose improves EGF release from ß-cells. After silencing PRSS8 in MIN6 cells, glucose-stimulated insulin secretion decreased, and EGFR signaling was impaired. Conversely, overexpression of PRSS8 in MIN6 cells induced higher concentrations of both basal and glucose-stimulated insulin secretion and increased phospho-EGFR concentrations. Furthermore, short-term exposure to glucose improved the concentration of endogenous PRSS8 in MIN6 cells through inhibition of intracellular degradation. These findings suggest that PRSS8 is involved in glucose-dependent physiological regulation of insulin secretion via the EGF-EGFR signaling pathway in pancreatic ß-cells.
Assuntos
Células Secretoras de Insulina , Peptídeo Hidrolases , Masculino , Animais , Camundongos , Secreção de Insulina , Peptídeo Hidrolases/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Glucose/metabolismo , Receptores ErbB/metabolismoRESUMO
Aldosterone plays an important role in the regulation of blood pressure by modulating the activity of the epithelial sodium channel (ENaC) that consists of α-, ß-, and γ-subunits. Aldosterone induces a molecular weight shift of γENaC from 85 to 70 kDa that is necessary for the channel activation. In vitro experiments demonstrated that a dual cleavage mechanism is responsible for this shift. It has been postulated that furin executes the primary cleavage in the Golgi and that the second cleavage is provided by other serine proteases such as prostasin or plasmin at the plasma membrane. However, the in vivo contribution of serine proteases to this cleavage remains unclear. To address this issue, we administered the synthetic serine protease inhibitor camostat mesilate (CM) to aldosterone-infused rats. CM decreased the abundance of the 70-kDa form of ENaC and led to a new 75-kDa form with a concomitant increase in the urinary Na-to-K ratio. Because CM inhibits the protease activity of serine proteases such as prostasin and plasmin, but not furin, our findings strongly indicate that CM inhibited the second cleavage of γENaC and subsequently suppressed ENaC activity. The results of our current studies also suggest the possibility that the synthetic serine protease inhibitor CM might represent a new strategy for the treatment of salt-sensitive hypertension in humans.
Assuntos
Aldosterona/farmacologia , Pressão Sanguínea/fisiologia , Canais Epiteliais de Sódio/metabolismo , Serina Proteases/metabolismo , Animais , Pressão Sanguínea/efeitos dos fármacos , Cloretos/metabolismo , Ésteres , Gabexato/análogos & derivados , Gabexato/farmacologia , Guanidinas , Masculino , Potássio/metabolismo , Ratos , Ratos Sprague-Dawley , Inibidores de Serina Proteinase/farmacologia , Sódio/metabolismoRESUMO
The number of the chronic renal failure (CRF) patients is increasing explosively. Hypertension, proteinuria, inflammation, fibrosis, and oxidative stress are intertwined in a complicated manner that leads to the progression of CRF. However, the therapeutic strategies to delay its progression are limited. Since serine proteases are involved in many processes that contribute to these risk factors, we investigated the effects of a synthetic serine protease inhibitor, camostat mesilate (CM), on the progression of CRF in 5/6 nephrectomized (Nx) rats. Eighteen male Sprague-Dawley rats were divided into three groups: a sham-operated group (n = 6), a vehicle-treated Nx group (n = 6), and a CM-treated Nx group (n = 6). Following the 9-wk study period, both proteinuria and serum creatinine levels were substantially increased in the vehicle-treated Nx group, and treatment with CM significantly reduced proteinuria and serum creatinine levels. The levels of podocyte-associated proteins in glomeruli, such as nephrin and synaptopodin, were markedly decreased by 5/6 nephrectomy, and this was significantly ameliorated by CM. CM also suppressed the levels of inflammatory and fibrotic marker mRNAs including transforming growth factor-ß1, TNF-α, collagen types I, III, and IV, and reduced glomerulosclerosis, glomerular hypertrophy, and interstitial fibrosis in histological studies. Furthermore, CM decreased the expression of NADPH oxidase component mRNAs, as well as reactive oxygen species generation and advanced oxidative protein product levels. Our present results strongly suggest the possibility that CM could be a useful therapeutic agent against the progression of CRF.
Assuntos
Gabexato/análogos & derivados , Falência Renal Crônica/tratamento farmacológico , Rim/efeitos dos fármacos , Proteinúria/tratamento farmacológico , Inibidores de Serina Proteinase/uso terapêutico , Animais , Creatinina/sangue , Progressão da Doença , Ésteres , Gabexato/farmacologia , Gabexato/uso terapêutico , Guanidinas , Rim/metabolismo , Rim/patologia , Falência Renal Crônica/metabolismo , Falência Renal Crônica/patologia , Masculino , Proteínas de Membrana/metabolismo , Proteínas dos Microfilamentos/metabolismo , Nefrectomia , Estresse Oxidativo/efeitos dos fármacos , Proteinúria/metabolismo , Proteinúria/patologia , Ratos , Ratos Sprague-Dawley , Inibidores de Serina Proteinase/farmacologia , Resultado do TratamentoRESUMO
Diabetic kidney disease (DKD) is a critical complication associated with diabetes; however, there are only a few animal models that can be used to explore its pathogenesis. In the present study, we established a mouse model of DKD using a technique based on the Developmental Origins of Health and Disease theory, i.e., by manipulating the embryonic environment, and investigated whether a dietary intervention could ameliorate the model's pathology. Two-cell embryos were cultured in vitro in α-minimum essential medium (MEM; MEM mice) or in standard potassium simplex-optimized medium (KSOM) as controls (KSOM mice) for 48 h, and the embryos were reintroduced into the mothers. The MEM and KSOM mice born were fed a high-fat, high-sugar diet for 58 days after they were 8 weeks old. Subsequently, half of the MEM mice and all KSOM mice were fed a diet containing rice powder (control diet), and the remaining MEM mice were fed a diet containing barley powder (barley diet) for 10 weeks. Glomerulosclerosis and pancreatic exhaustion were observed in MEM mice, but not in control KSOM mice. Renal arteriolar changes, including intimal thickening and increase in the rate of hyalinosis, were more pronounced in MEM mice fed a control diet than in KSOM mice. Immunostaining showed the higher expression of transforming growth factor beta (TGFB) in the proximal/distal renal tubules of MEM mice fed a control diet than in those of KSOM mice. Pathologies, such as glomerulosclerosis, renal arteriolar changes, and higher TGFB expression, were ameliorated by barley diet intake in MEM mice. These findings suggested that the MEM mouse is an effective DKD animal model that shows glomerulosclerosis and renal arteriolar changes, and barley intake can improve these pathologies in MEM mice.
Assuntos
Nefropatias Diabéticas , Dieta , Técnicas de Cultura Embrionária/métodos , Hordeum , Ração Animal , Animais , Animais Recém-Nascidos , Células Cultivadas , Meios de Cultura/farmacologia , Nefropatias Diabéticas/dietoterapia , Nefropatias Diabéticas/embriologia , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Ingestão de Alimentos/fisiologia , Embrião de Mamíferos , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Masculino , Camundongos , Camundongos Endogâmicos ICR , Compostos Orgânicos/farmacologia , GravidezRESUMO
A serine protease prostasin has been demonstrated to have a pivotal role in the activation of the epithelial sodium channel. Systemic administration of adenovirus carrying human prostasin gene in rats resulted in an increase in plasma prostasin and aldosterone levels. However, the mechanism by which the elevation of prostasin levels in the systemic circulation stimulated the plasma aldosterone levels remains unknown. Therefore, we examined if prostasin increases the aldosterone synthesis in a human adrenocortical cell line (H295R cells). Luciferase assay using CYP11B2 promoter revealed that prostasin significantly increased the transcriptional activity of CYP11B2. Prostasin significantly increased both CYP11B2 mRNA expression and aldosterone production in a dose-dependent manner. Surprisingly, treatment with camostat mesilate, a potent prostasin inhibitor, had no effect on the aldosterone synthesis by prostasin and also a protease-dead mutant of prostasin significantly stimulated the aldosterone production. A T-type/L-type calcium channel blocker and a protein kinase C (PKC) inhibitor significantly reduced the aldosterone synthesis by prostasin. Our findings suggest a stimulatory effect of prostasin on the aldosterone synthesis by adrenal gland through the nonproteolytic action and indicate a new role of prostasin in the systemic circulation.