RESUMO
Fibroblast growth factor (FGF) 23 is a phosphate-regulating hormone that is elevated in patients with chronic kidney disease and associated with cardiovascular mortality. Experimental studies showed that elevated FGF23 levels induce cardiac hypertrophy by targeting cardiac myocytes via FGF receptor isoform 4 (FGFR4). A recent structural analysis revealed that the complex of FGF23 and FGFR1, the physiologic FGF23 receptor in the kidney, includes soluble α-klotho (klotho) and heparin, which both act as co-factors for FGF23/FGFR1 signaling. Here, we investigated whether soluble klotho, a circulating protein with cardio-protective properties, and heparin, a factor that is routinely infused into patients with kidney failure during the hemodialysis procedure, regulate FGF23/FGFR4 signaling and effects in cardiac myocytes. We developed a plate-based binding assay to quantify affinities of specific FGF23/FGFR interactions and found that soluble klotho and heparin mediate FGF23 binding to distinct FGFR isoforms. Heparin specifically mediated FGF23 binding to FGFR4 and increased FGF23 stimulatory effects on hypertrophic growth and contractility in isolated cardiac myocytes. When repetitively injected into two different mouse models with elevated serum FGF23 levels, heparin aggravated cardiac hypertrophy. We also developed a novel procedure for the synthesis and purification of recombinant soluble klotho, which showed anti-hypertrophic effects in FGF23-treated cardiac myocytes. Thus, soluble klotho and heparin act as independent FGF23 co-receptors with opposite effects on the pathologic actions of FGF23, with soluble klotho reducing and heparin increasing FGF23-induced cardiac hypertrophy. Hence, whether heparin injections during hemodialysis in patients with extremely high serum FGF23 levels contribute to their high rates of cardiovascular events and mortality remains to be studied.
Assuntos
Fator de Crescimento de Fibroblastos 23 , Heparina , Proteínas Klotho , Insuficiência Renal Crônica , Animais , Cardiomegalia , Glucuronidase/metabolismo , Heparina/metabolismo , Humanos , Proteínas Klotho/metabolismo , Camundongos , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/terapiaRESUMO
RATIONALE & OBJECTIVE: Pulmonary hypertension (PH) contributes to cardiovascular disease and mortality in patients with chronic kidney disease (CKD), but the pathophysiology is mostly unknown. This study sought to estimate the prevalence and consequences of PH subtypes in the setting of CKD. STUDY DESIGN: Observational retrospective cohort study. SETTING & PARTICIPANTS: We examined 12,618 patients with a right heart catheterization in the Duke Databank for Cardiovascular Disease from January 1, 2000, to December 31, 2014. EXPOSURES: Baseline kidney function stratified by CKD glomerular filtration rate category and PH subtype. OUTCOMES: All-cause mortality. ANALYTICAL APPROACH: Multivariable Cox proportional hazards analysis. RESULTS: In this cohort, 73.4% of patients with CKD had PH, compared with 56.9% of patients without CKD. Isolated postcapillary PH (39.0%) and combined pre- and postcapillary PH (38.3%) were the most common PH subtypes in CKD. Conversely, precapillary PH was the most common subtype in the non-CKD cohort (35.9%). The relationships between mean pulmonary artery pressure, pulmonary capillary wedge pressure, and right atrial pressure with mortality were similar in both the CKD and non-CKD cohorts. Compared with those without PH, precapillary PH conferred the highest mortality risk among patients without CKD (HR, 2.27; 95% CI, 2.00-2.57). By contrast, in those with CKD, combined pre- and postcapillary PH was associated with the highest risk for mortality in CKD in adjusted analyses (compared with no PH, HRs of 1.89 [95% CI, 1.57-2.28], 1.87 [95% CI, 1.52-2.31], 2.13 [95% CI, 1.52-2.97], and 1.63 [95% CI, 1.12-2.36] for glomerular filtration rate categories G3a, G3b, G4, and G5/G5D). LIMITATIONS: The cohort referred for right heart catheterization may not be generalizable to the general population. Serum creatinine data in the 6 months preceding catheterization may not reflect true baseline CKD. Observational design precludes assumptions of causality. CONCLUSIONS: In patients with CKD referred for right heart catheterization, PH is common and associated with poor survival. Combined pre- and postcapillary PH was common and portended the worst survival for patients with CKD.
Assuntos
Hipertensão Pulmonar/classificação , Insuficiência Renal Crônica/epidemiologia , Idoso , Cateterismo Cardíaco , Causas de Morte , Comorbidade , Feminino , Hemodinâmica , Humanos , Hipertensão Pulmonar/complicações , Hipertensão Pulmonar/epidemiologia , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Mortalidade , Prevalência , Modelos de Riscos Proporcionais , Estudos RetrospectivosRESUMO
Circulating levels of fibroblast growth factor (FGF)23 are associated with systemic inflammation and increased mortality in chronic kidney disease. α-Klotho, a co-receptor for FGF23, is downregulated in chronic obstructive pulmonary disease (COPD). However, whether FGF23 and Klotho-mediated FGF receptor (FGFR) activation delineates a pathophysiological mechanism in COPD remains unclear. We hypothesised that FGF23 can potentiate airway inflammation via Klotho-independent FGFR4 activation.FGF23 and its effect were studied using plasma and transbronchial biopsies from COPD and control patients, and primary human bronchial epithelial cells isolated from COPD patients as well as a murine COPD model.Plasma FGF23 levels were significantly elevated in COPD patients. Exposure of airway epithelial cells to cigarette smoke and FGF23 led to a significant increase in interleukin-1ß release via Klotho-independent FGFR4-mediated activation of phospholipase Cγ/nuclear factor of activated T-cells signalling. In addition, Klotho knockout mice developed COPD and showed airway inflammation and elevated FGFR4 expression in their lungs, whereas overexpression of Klotho led to an attenuation of airway inflammation.Cigarette smoke induces airway inflammation by downregulation of Klotho and activation of FGFR4 in the airway epithelium in COPD. Inhibition of FGF23 or FGFR4 might serve as a novel anti-inflammatory strategy in COPD.
Assuntos
Fatores de Crescimento de Fibroblastos/sangue , Glucuronidase/metabolismo , Pulmão/patologia , Doença Pulmonar Obstrutiva Crônica/sangue , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Adulto , Idoso , Animais , Células Epiteliais/metabolismo , Feminino , Fator de Crescimento de Fibroblastos 23 , Glucuronidase/genética , Humanos , Inflamação/patologia , Proteínas Klotho , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Doença Pulmonar Obstrutiva Crônica/metabolismo , Fumaça/efeitos adversosRESUMO
CD63 is a ubiquitously expressed member of the tetraspanin superfamily. Using a mating-based split-ubiquitin-yeast 2-hybrid system, pull-down experiments, total internal reflection fluorescence microscopy, Förster resonance energy transfer, and biotinylation assays, we found that CD63 interacts with human organic cation transporter 2 (hOCT2), which transports endogenous and exogenous substrates, such as neurotransmitters and drugs in several epithelial cells. CD63 overexpression affects cellular localization of hOCT2 expressed in human embryonic kidney (HEK)293 cells. Studies with CD63-knockout mice indicate that in renal proximal tubules, CD63 determines the insertion of the mouse ortholog of the transporter into the proper membrane domain and mediates transporter regulation by trafficking processes. In polarized Madin-Darby kidney canine kidney (MDCK) epithelial cells, CD63 and hOCT2 colocalize with the small GTPase Rab4, which controls the rapid recycling from sorting endosomes back to the cell surface. Suitable negative and positive control experiments were performed for each experimental approach. Empty vector transfected cells and wild-type mice were used as control. CD63 seems to play a role in the recycling of hOCT2 from endosomes to the basolateral membrane in polarized epithelia. These data indicate that CD63 has a previously uncharacterized function in regulating trafficking of specific membrane proteins in polarized cells.-Schulze, U., Brast, S., Grabner, A., Albiker, C., Snieder, B., Holle, S., Schlatter, E., Schröter, R., Pavenstädt, H., Herrmann, E., Lambert, C., Spoden, G. A., Florin, L., Saftig, P., Ciarimboli, G. Tetraspanin CD63 controls basolateral sorting of organic cation transporter 2 in renal proximal tubules.
Assuntos
Túbulos Renais Proximais/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Tetraspanina 30/metabolismo , Animais , Membrana Celular/metabolismo , Cães , Endossomos/metabolismo , Células Epiteliais/metabolismo , Células HEK293 , Humanos , Túbulos Renais Proximais/citologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos C57BL , Transportador 2 de Cátion Orgânico , Ligação Proteica , Transporte Proteico , Tetraspanina 30/genética , Proteínas rab4 de Ligação ao GTP/metabolismoRESUMO
BACKGROUND: Vitamin D deficiency and excess of circulating fibroblast growth factor 23 (FGF23) contribute to cardiovascular mortality in patients with chronic kidney disease (CKD). FGF23 activates FGF receptor 4 and (FGFR4) calcineurin/nuclear factor of activated T cells (NFAT) signaling in cardiac myocytes, thereby causing left ventricular hypertrophy (LVH). Here, we determined if 1,25-dihydroxyvitamin D (calcitriol) inhibits FGF23-induced cardiac signaling and LVH. METHODS: 5/6 nephrectomized (5/6 Nx) rats were treated with different doses of calcitriol for 4 or 10 weeks and cardiac expression of FGF23/FGFR4 and activation of calcineurin/NFAT as well as LVH were analyzed. FGFR4 activation and hypertrophic cell growth were studied in cultured cardiac myocytes that were co-treated with FGF23 and calcitriol. RESULTS: In 5/6Nx rats with LVH, we detected elevated FGF23 expression in bone and myocardium, increased cardiac expression of FGFR4 and elevated cardiac activation of calcineurin/NFAT signaling. Cardiac expression levels of FGF23 and FGFR4 significantly correlated with the presence of LVH in uremic rats. Treatment with calcitriol reduced LVH as well as cardiac FGFR4 expression and calcineurin/NFAT activation. Bone and cardiac FGF23 expression were further stimulated by calcitriol in a dose-dependent manner, but levels of intact cardiac FGF23 protein were suppressed by high-dose calcitriol. In cultured cardiac myocytes, co-treatment with calcitriol blocked FGF23-induced activation of FGFR4 and hypertrophic cell growth. CONCLUSIONS: Our data suggest that in CKD, cardioprotective effects of calcitriol stem from its inhibitory actions on the cardiac FGF23/FGFR4 system, and based on their counterbalancing effects on cardiac myocytes, high FGF23 and low calcitriol synergistically contribute to cardiac hypertrophy.
Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Hipertrofia Ventricular Esquerda/prevenção & controle , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Insuficiência Renal Crônica/complicações , Vitamina D/administração & dosagem , Animais , Fator de Crescimento de Fibroblastos 23 , Humanos , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Vitaminas/administração & dosagemRESUMO
Patients with chronic kidney disease (CKD) develop increased levels of the phosphate-regulating hormone, fibroblast growth factor (FGF) 23, that are associated with a higher risk of mortality. Increases in inflammatory markers are another common feature that predicts poor clinical outcomes. Elevated FGF23 is associated with higher circulating levels of inflammatory cytokines in CKD, which can stimulate osteocyte production of FGF23. Here, we studied whether FGF23 can directly stimulate hepatic production of inflammatory cytokines in the absence of α-klotho, an FGF23 coreceptor in the kidney that is not expressed by hepatocytes. By activating FGF receptor isoform 4 (FGFR4), FGF23 stimulated calcineurin signaling in cultured hepatocytes, which increased the expression and secretion of inflammatory cytokines, including C-reactive protein. Elevating serum FGF23 levels increased hepatic and circulating levels of C-reactive protein in wild-type mice, but not in FGFR4 knockout mice. Administration of an isoform-specific FGFR4 blocking antibody reduced hepatic and circulating levels of C-reactive protein in the 5/6 nephrectomy rat model of CKD. Thus, FGF23 can directly stimulate hepatic secretion of inflammatory cytokines. Our findings indicate a novel mechanism of chronic inflammation in patients with CKD and suggest that FGFR4 blockade might have therapeutic anti-inflammatory effects in CKD.
Assuntos
Citocinas/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Hepatócitos/metabolismo , Inflamação/metabolismo , Insuficiência Renal Crônica/metabolismo , Animais , Calcineurina/metabolismo , Fator de Crescimento de Fibroblastos 23 , Glucuronidase/metabolismo , Humanos , Proteínas Klotho , Camundongos , Fatores de Transcrição NFATC/metabolismo , Fosfolipase C gama/metabolismo , Cultura Primária de Células , Ratos , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de SinaisRESUMO
PURPOSE OF REVIEW: In chronic kidney disease (CKD), multiple factors contribute to the development of cardiac hypertrophy by directly targeting the heart or indirectly by inducing systemic changes such as hypertension, anemia, and inflammation. Furthermore, disturbances in phosphate metabolism have been identified as nonclassical risk factors for cardiovascular mortality in these patients. With declining kidney function, the physiologic regulators of phosphate homeostasis undergo changes in their activity as well as their circulating levels, thus potentially contributing to cardiac hypertrophy once they are out of balance. Recently, two of these phosphate regulators, fibroblast growth factor 23 (FGF23) and Klotho, have been shown to affect cardiac remodeling, thereby unveiling a novel pathomechanism of cardiac hypertrophy in CKD. Here we discuss the potential direct versus indirect effects of FGF23 and the soluble form of Klotho on the heart, and their crosstalk in the regulation of cardiac hypertrophy. RECENT FINDINGS: In models of CKD, FGF23 can directly target cardiac myocytes via FGF receptor 4 and induce cardiac hypertrophy in a blood pressure-independent manner. Soluble Klotho may directly target the heart via an unknown receptor thereby protecting the myocardium from pathologic stress stimuli that are associated with CKD, such as uremic toxins or FGF23. SUMMARY: Elevated serum levels of FGF23 and reduced serum levels of soluble Klotho contribute to uremic cardiomyopathy in a synergistic manner.
Assuntos
Cardiomegalia/metabolismo , Cardiomiopatias/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Glucuronidase/metabolismo , Miocárdio/metabolismo , Insuficiência Renal Crônica/metabolismo , Uremia/metabolismo , Cardiomegalia/etiologia , Cardiomiopatias/etiologia , Fator de Crescimento de Fibroblastos 23 , Humanos , Proteínas Klotho , Fosfatos/metabolismo , Insuficiência Renal Crônica/complicações , Uremia/complicaçõesRESUMO
BACKGROUND: Activation of fibroblast growth factor receptor (FGFR)-dependent signalling by FGF23 may contribute to the complex pathogenesis of left ventricular hypertrophy (LVH) in chronic kidney disease (CKD). Pan FGFR blockade by PD173074 prevented development of LVH in the 5/6 nephrectomy rat model of CKD, but its ability to treat and reverse established LVH is unknown. METHODS: CKD was induced in rats by 5/6 nephrectomy. Two weeks later, rats began treatment with vehicle (0.9% NaCl) or PD173074, 1 mg/kg once-daily for 3 weeks. Renal function was determined by urine and blood analyses. Left ventricular (LV) structure and function were determined by echocardiography, histopathology, staining for myocardial fibrosis (Sirius-Red) and investigating cardiac gene expression profiles by real-time PCR. RESULTS: Two weeks after inducing CKD by 5/6 nephrectomy, rats manifested higher (mean ± SEM) systolic blood pressure (208 ± 4 versus 139 ± 3 mmHg; P < 0.01), serum FGF23 levels (1023 ± 225 versus 199 ± 9 pg/mL; P < 0.01) and LV mass (292 ± 9 versus 220 ± 3 mg; P < 0.01) when compared with sham-operated animals. Thereafter, 3 weeks of treatment with PD173074 compared with vehicle did not significantly change blood pressure, kidney function or metabolic parameters, but significantly reduced LV mass (230 ± 14 versus 341 ± 33 mg; P < 0.01), myocardial fibrosis (2.5 ± 0.7 versus 5.4 ± 0.95% staining/field; P < 0.01) and cardiac expression of genes associated with pathological LVH, while significantly increasing ejection fraction (18 versus 2.5% post-treatment increase; P < 0.05). CONCLUSIONS: FGFR blockade improved cardiac structure and function in 5/6 nephrectomy rats with previously established LVH. These data support FGFR activation as a potentially modifiable, blood pressure-independent molecular mechanism of LVH in CKD.
Assuntos
Ventrículos do Coração/diagnóstico por imagem , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Pirimidinas/uso terapêutico , Receptores de Fatores de Crescimento de Fibroblastos/antagonistas & inibidores , Insuficiência Renal Crônica/complicações , Função Ventricular Esquerda/fisiologia , Animais , Modelos Animais de Doenças , Ecocardiografia , Ventrículos do Coração/fisiopatologia , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/fisiopatologia , Injeções Intraperitoneais , Masculino , Pirimidinas/administração & dosagem , Ratos , Ratos Sprague-Dawley , Função Ventricular Esquerda/efeitos dos fármacosRESUMO
Cardiovascular disease is the leading cause of death in patients with chronic kidney disease (CKD). As CKD progresses, CKD-specific risk factors, such as disordered mineral homeostasis, amplify traditional cardiovascular risk factors. Fibroblast growth factor 23 (FGF23) regulates mineral homeostasis by activating complexes of FGF receptors and transmembrane klotho co-receptors. A soluble form of klotho also acts as a 'portable' FGF23 co-receptor in tissues that do not express klotho. In progressive CKD, rising circulating FGF23 levels in combination with decreasing kidney expression of klotho results in klotho-independent effects of FGF23 on the heart that promote left ventricular hypertrophy, heart failure, atrial fibrillation and death. Emerging data suggest that soluble klotho might mitigate some of these effects via several candidate mechanisms. More research is needed to investigate FGF23 excess and klotho deficiency in specific cardiovascular complications of CKD, but the pathophysiological primacy of FGF23 excess versus klotho deficiency might never be precisely resolved, given the entangled feedback loops that they share. Therefore, randomized trials should prioritize clinical practicality over scientific certainty by targeting disordered mineral homeostasis holistically in an effort to improve cardiovascular outcomes in patients with CKD.
Assuntos
Doenças Cardiovasculares , Insuficiência Renal Crônica , Humanos , Glucuronidase/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Rim , Insuficiência Renal Crônica/complicações , Minerais/metabolismoRESUMO
Human organic cation transporter 2 (hOCT2) is involved in transport of many endogenous and exogenous organic cations, mainly in kidney and brain cells. Because the quaternary structure of transmembrane proteins plays an essential role for their cellular trafficking and function, we investigated whether hOCT2 forms oligomeric complexes, and if so, which part of the transporter is involved in the oligomerization. A yeast 2-hybrid mating-based split-ubiquitin system (mbSUS), fluorescence resonance energy transfer, Western blot analysis, cross-linking experiments, immunofluorescence, and uptake measurements of the fluorescent organic cation 4-(4-(dimethylamino)styryl)-N-methylpyridinium were applied to human embryonic kidney 293 (HEK293) cells transfected with hOCT2 and partly also to freshly isolated human proximal tubules. The role of cysteines for oligomerization and trafficking of the transporter to the plasma membranes was investigated in cysteine mutants of hOCT2. hOCT2 formed oligomers both in the HEK293 expression system and in native human kidneys. The cysteines of the large extracellular loop are important to enable correct folding, oligomeric assembly, and plasma membrane insertion of hOCT2. Mutation of the first and the last cysteines of the loop at positions 51 and 143 abolished oligomer formation. Thus, the cysteines of the extracellular loop are important for correct trafficking of the transporter to the plasma membrane and for its oligomerization.
Assuntos
Membrana Celular/metabolismo , Cisteína/metabolismo , Túbulos Renais Proximais/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Transporte Biológico , Western Blotting , Cisteína/química , Cisteína/genética , Transferência Ressonante de Energia de Fluorescência , Células HEK293 , Células HeLa , Humanos , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Microscopia de Fluorescência , Mutação , Proteínas de Transporte de Cátions Orgânicos/química , Proteínas de Transporte de Cátions Orgânicos/genética , Transportador 2 de Cátion Orgânico , Ligação Proteica , Multimerização Proteica , Compostos de Piridínio/farmacocinética , Técnicas de Cultura de Tecidos , Transfecção , Técnicas do Sistema de Duplo-HíbridoRESUMO
Chronic kidney disease (CKD) is a global health epidemic that significantly increases mortality due to cardiovascular disease. Left ventricular hypertrophy (LVH) is an important mechanism of cardiac injury in CKD. High serum levels of fibroblast growth factor (FGF) 23 in patients with CKD may contribute mechanistically to the pathogenesis of LVH by activating FGF receptor (FGFR) 4 signaling in cardiac myocytes. Mitochondrial dysfunction and cardiac metabolic remodeling are early features of cardiac injury that predate development of hypertrophy, but these mechanisms of disease have been insufficiently studied in models of CKD. Wild-type mice with CKD induced by adenine diet developed LVH that was preceded by morphological changes in mitochondrial structure and evidence of cardiac mitochondrial and metabolic dysfunction. In bioengineered cardio-bundles and neonatal rat ventricular myocytes grown in vitro, FGF23-mediated activation of FGFR4 caused a mitochondrial pathology, characterized by increased bioenergetic stress and increased glycolysis, that preceded the development of cellular hypertrophy. The cardiac metabolic changes and associated mitochondrial alterations in mice with CKD were prevented by global or cardiac-specific deletion of FGFR4. These findings indicate that metabolic remodeling and eventually mitochondrial dysfunction are early cardiac complications of CKD that precede structural remodeling of the heart. Mechanistically, FGF23-mediated activation of FGFR4 causes mitochondrial dysfunction, suggesting that early pharmacologic inhibition of FGFR4 might serve as novel therapeutic intervention to prevent development of LVH and heart failure in patients with CKD.
RESUMO
AIMS: Chronic kidney disease is directly associated with cardiovascular complications. Heart remodelling, including fibrosis, hypertrophy, and decreased vascularization, is frequently present in renal diseases. Our objective was to investigate the impact of calcineurin inhibitors (CNI) on cardiac remodelling and function in a rat model of renal disease. METHODS AND RESULTS: Male Sprague Dawley rats were divided into six groups: sham-operated rats, 5/6 nephrectomized rats (Nx) treated with vehicle, CNI (cyclosporine A 5.0 or 7.5, or tacrolimus 0.5 mg/kg/day) or hydralazine (20 mg/kg twice a day) for 14 days, starting on the day of surgery. Creatinine clearance was significantly lower and blood pressure significantly higher in Nx rats when compared with controls. Morphological and echocardiographic analyses revealed increased left ventricular hypertrophy and decreased number of capillaries in Nx rats. Treatment with CNI affected neither the renal function nor the blood pressure, but prevented the development of cardiac hypertrophy and improved vascularization. In addition, regional blood volume improved as confirmed by contrast agent-based echocardiography. Hydralazine treatment did not avoid heart remodelling in this model. Gene expression analysis verified a decrease in hypertrophic genes in the heart of CNI-treated rats, while pro-angiogenic and stem cell-related genes were upregulated. Moreover, mobilization of stem/progenitor cells was increased through manipulation of the CD26/SDF-1 system. CONCLUSION: We conclude from our studies that CNI-treatment significantly prevented cardiac remodelling and improved heart function in Nx rats without affecting renal function and blood pressure. This sheds new light on possible therapeutic strategies for renal patients at high cardiovascular risk.
Assuntos
Inibidores de Calcineurina , Ciclosporina/uso terapêutico , Cardiopatias/prevenção & controle , Imunossupressores/uso terapêutico , Nefropatias/complicações , Tacrolimo/uso terapêutico , Animais , Doença Crônica , Hipertrofia Ventricular Esquerda/prevenção & controle , Masculino , Nefrectomia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Remodelação Ventricular/efeitos dos fármacosRESUMO
Fibroblast growth factor (FGF) 21, a hormone that increases insulin sensitivity, has shown promise as a therapeutic agent to improve metabolic dysregulation. Here we report that FGF21 directly targets cardiac myocytes by binding ß-klotho and FGF receptor (FGFR) 4. In combination with high glucose, FGF21 induces cardiac myocyte growth in width mediated by extracellular signal-regulated kinase 1/2 (ERK1/2) signaling. While short-term FGF21 elevation can be cardio-protective, we find that in type 2 diabetes (T2D) in mice, where serum FGF21 levels are elevated, FGFR4 activation induces concentric cardiac hypertrophy. As T2D patients are at risk for heart failure with preserved ejection fraction (HFpEF), we propose that induction of concentric hypertrophy by elevated FGF21-FGFR4 signaling may constitute a novel mechanism promoting T2D-associated HFpEF such that FGFR4 blockade might serve as a cardio-protective therapy in T2D. In addition, potential adverse cardiac effects of FGF21 mimetics currently in clinical trials should be investigated.
Assuntos
Diabetes Mellitus Tipo 2 , Insuficiência Cardíaca , Animais , Cardiomegalia/metabolismo , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Fatores de Crescimento de Fibroblastos/metabolismo , Insuficiência Cardíaca/metabolismo , Humanos , Camundongos , Miócitos Cardíacos/metabolismo , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Volume SistólicoRESUMO
In the context of transplantation, complement activation is associated with poor prognosis and outcome. While complement activation in antibody-mediated rejection is well-known, less is known about complement activation in acute T cell-mediated rejection (TCMR). There is increasing evidence that complement contributes to the clearance of apoptotic debris and tissue repair. In this regard, we have analysed published human kidney biopsy transcriptome data clearly showing upregulated expression of complement factors in TCMR. To clarify whether and how the complement system is activated early during acute TCMR, experimental syngeneic and allogeneic renal transplantations were performed. Using an allogeneic rat renal transplant model, we also observed upregulation of complement factors in TCMR in contrast to healthy kidneys and isograft controls. While staining for C4d was positive, staining with a C3d antibody showed no C3d deposition. FACS analysis of blood showed the absence of alloantibodies that could have explained the C4d deposition. Gene expression pathway analysis showed upregulation of pro-apoptotic factors in TCMR, and apoptotic endothelial cells were detected by ultrastructural analysis. Monocytes/macrophages were found to bind to and phagocytise these apoptotic cells. Therefore, we conclude that early C4d deposition in TCMR may be relevant to the clearance of apoptotic cells.
Assuntos
Apoptose , Complemento C4b/metabolismo , Transplante de Rim , Fragmentos de Peptídeos/metabolismo , Animais , Biópsia , Modelos Animais de Doenças , Regulação da Expressão Gênica , Rim/patologia , Rim/ultraestrutura , Masculino , Ratos Endogâmicos BN , Receptores de Morte Celular/metabolismo , Transdução de Sinais , Transcriptoma/genética , Transplante HomólogoRESUMO
The rapid rise in circulating fibroblast growth factor 23 (FGF23) associated with kidney injury results in calcitriol deficiency, altered calcium homeostasis, and secondary hyperparathyroidism, and may contribute to cardiovascular complications and death. However, the mechanisms of increased FGF23 in states of kidney injury remain unclear. In this issue of the JCI, Simic et al. screened plasma taken from the renal vein of patients undergoing cardiac catheterization and identified glycerol-3-phosphate (G-3-P) as the most significant correlate of simultaneous arterial FGF23 levels. When G-3-P was administered to mice, FGF23 production increased in bone. In a series of elegant mouse studies, the authors discovered a pathway linking increased G-3-P to increased FGF23 via increases in lysophosphatidic acid (LPA), which activates the LPA receptor 1 in FGF23-secreting cells in the bone and bone marrow. Although the authors present human data that broadly support the results from the mouse models, further research is needed to determine whether targeting the G-3-P/FGF23 pathway has the potential to modify FGF23-related complications in the clinic.
Assuntos
Fatores de Crescimento de Fibroblastos , Glicerol , Animais , Fator de Crescimento de Fibroblastos 23 , Fatores de Crescimento de Fibroblastos/genética , Humanos , Rim , Camundongos , FosfatosRESUMO
In chronic kidney disease (CKD), elevated serum levels of the phosphate regulating hormone fibroblast growth factor (FGF) 23 have emerged as powerful risk factors for cardiovascular disease and death. Mechanistically, FGF23 can bind and activate fibroblast growth factor receptor (FGFR) 4 independently of α-klotho, the canonical co-receptor for FGF23 in the kidney, which stimulates left ventricular hypertrophy and hepatic production of inflammatory cytokines. FGF23 has also been shown to independently predict progression of renal disease, however, whether FGF23 and FGFR4 also contribute to CKD remains unknown. Here, we generated a mouse model with dual deletions of FGFR4 and α-klotho, and we induced CKD in mice with either global deletion or constitutive activation of FGFR4. We demonstrate that FGF23 is not capable of inducing phosphaturia via FGFR4 and that FGFR4 does not promote or mitigate renal injury in animal models of CKD. Taken together our results suggest FGFR4 inhibition as a safe alternative strategy to target cardiovascular disease and chronic inflammation in patients with CKD without interrupting the necessary phosphaturic effects of FGF23.
Assuntos
Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Insuficiência Renal Crônica/patologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Fator de Crescimento de Fibroblastos 23 , Técnicas de Introdução de Genes , Glucuronidase/metabolismo , Humanos , Proteínas Klotho , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/fisiologia , Fatores de RiscoRESUMO
BACKGROUND: In uremic animals, vitamin D receptor (VDR) agonists like paricalcitol (Pc) attenuate cardiac hypertrophy, but this effect has not been replicated consistently in humans with chronic kidney disease. Elevated fibroblast growth factor 23 (FGF23) levels cause cardiac hypertrophy with activation of the myocardial calcineurin/nuclear factor of activated T cell (NFAT) axis and may antagonize the cardioprotective effects of VDR agonist therapy. We hypothesized that the effectiveness of Pc may depend on the prevailing circulating levels of FGF23 and could be potentiated by the combined administration of a pan-FGF23 receptor (FGFR) blocker agent (PD173074). METHODS: In rats with 5/6 nephrectomy treated with Pc or PD173074 or both agents concurrently, myocardial mRNA expression of renin-angiotensin system, VDR, FGFR4, and calcineurin/NFAT target genes was determined. In adolescents on hemodialysis, we analyzed sequential echocardiograms, blood pressures and serial FGF23 measurements, and their relations to the cumulative administered dose of parenteral Pc. RESULTS: The ratio of Pc dose/plasma levels of FGF23 correlated inversely (P < 0.005) with the cardiac mass in uremic rats and in hemodialysis patients, independently of hypertension. Despite persistently elevated FGF23 levels and myocardial FGFR4 activation, Pc suppressed upregulated myocardial calcineurin/NFAT target genes, and the effects were amplified by coadministration of PD173074. CONCLUSIONS: The beneficial effects of Pc on uremic cardiac hypertrophy are counterbalanced by the increased FGF23 levels. Blockade of FGF23-mediated signaling increased the Pc-induced suppression of the myocardial calcineurin/NFAT system. Higher doses of Pc should be considered in the treatment of patients with uremic cardiomyopathy.
Assuntos
Síndrome Cardiorrenal/prevenção & controle , Cardiomiopatias/prevenção & controle , Ergocalciferóis/farmacologia , Fatores de Crescimento de Fibroblastos/sangue , Ventrículos do Coração/efeitos dos fármacos , Falência Renal Crônica/tratamento farmacológico , Pirimidinas/farmacologia , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptores de Calcitriol/agonistas , Adolescente , Animais , Síndrome Cardiorrenal/metabolismo , Síndrome Cardiorrenal/fisiopatologia , Cardiomiopatias/metabolismo , Cardiomiopatias/fisiopatologia , Criança , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Fator de Crescimento de Fibroblastos 23 , Ventrículos do Coração/metabolismo , Ventrículos do Coração/fisiopatologia , Humanos , Falência Renal Crônica/metabolismo , Falência Renal Crônica/fisiopatologia , Masculino , Ratos Sprague-Dawley , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Receptores de Calcitriol/metabolismo , Estudos Retrospectivos , Transdução de Sinais , Uremia/tratamento farmacológico , Uremia/metabolismo , Uremia/patologia , Uremia/fisiopatologia , Função Ventricular Esquerda/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacosRESUMO
Organic cation transporters (OCTs) are membrane proteins with relevant physiological (because they accept neurotransmitters as substrate) and pharmacological (because of their interaction with drugs) roles. The human OCTs hOCT1 (SLC22A1/hOCT1) and hOCT2 (SLC22A2/hOCT2) are highly expressed in hepatic (hOCT1) and in renal and neuronal tissue (hOCT2), suggesting a possible role in modulating neurotransmitter activity in the liver, kidney, and brain, and their clearance from the blood. Even though there are several data demonstrating that OCTs are regulated under various patho-physiological conditions, it remains largely unknown which proteins directly interact with OCTs and thereby influence their cellular processing, localization, and function. In this work, using a mating-based split-ubiquitin yeast two-hybrid system, we characterized the potential interactome of hOCT1 and 2. It became evident that these OCTs share some potential interaction partners, such as the tetraspanins CD63 and CD9. Moreover, we confirmed interaction of hOCT2 with CD9 by fluorescence-activated cell sorting coupled with Förster resonance energy transfer analysis. Together with other proteins, tetraspanins build "tetraspanins webs" in the plasma membrane, which are able to regulate cellular trafficking and compartmentalization of interacting partners. While CD63 was demonstrated to mediate the localization of the hOCT2 to the endosomal system, we show here that co-expression of hOCT2 and CD9 led to strong cell surface localization of the transporter. These data suggest that tetraspanins regulate the cellular localization and function of OCTs. Co-localization of CD9 and hOCT was confirmed in tissues endogenously expressing proteins, highlighting the potential biological relevance of this interaction.
Assuntos
Fator 1 de Transcrição de Octâmero/metabolismo , Transportador 2 de Cátion Orgânico/metabolismo , Tetraspanina 29/metabolismo , Tetraspaninas/metabolismo , Animais , Membrana Celular/metabolismo , Cães , Células HEK293 , Humanos , Células Madin Darby de Rim Canino , Proteínas de Membrana/metabolismo , Transporte Proteico/fisiologiaRESUMO
Immune evasion and the suppression of antitumor responses during cancer progression are considered hallmarks of cancer and are typically attributed to tumor-derived factors. Although the molecular basis for the crosstalk between tumor and immune cells is an area of active investigation, whether host-specific germline variants can dictate immunosuppressive mechanisms has remained a challenge to address. A commonly occurring germline mutation (c.1162G>A/rs351855 G/A) in the FGFR4 (CD334) gene enhances signal transducer and activator of transcription 3 (STAT3) signaling and is associated with poor prognosis and accelerated progression of multiple cancer types. Here, using rs351855 SNP-knockin transgenic mice and Fgfr4-knockout mice, we reveal the genotype-specific gain of immunological function of suppressing the CD8/CD4+FOXP3+CD25+ regulatory T cell ratio in vivo. Furthermore, using knockin transgenic mouse models for lung and breast cancers, we establish the host-specific, tumor-extrinsic functions of STAT3-enhancing germline variants in impeding the tumor infiltration of CD8 T cells. Thus, STAT3-enhancing germline receptor variants contribute to immune evasion through their pleiotropic functions in immune cells.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Mutação em Linhagem Germinativa/imunologia , Proteínas de Neoplasias/imunologia , Neoplasias Experimentais/imunologia , Fator de Transcrição STAT3/imunologia , Linfócitos T Reguladores/imunologia , Evasão Tumoral , Animais , Linfócitos T CD8-Positivos/patologia , Mutação em Linhagem Germinativa/genética , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/imunologia , Fator de Transcrição STAT3/genética , Linfócitos T Reguladores/patologiaRESUMO
The liver plays a decisive role in the regulation of systemic inflammation. In chronic kidney disease in particular, the liver reacts in response to the uremic milieu, oxidative stress, endotoxemia and the decreased clearance of circulating proinflammatory cytokines by producing a large number of acute-phase reactants. Experimental tools to study inflammation and the underlying role of hepatocytes are crucial to understand the regulation and contribution of hepatic cytokines to a systemic acute phase response and a prolonged pro-inflammatory scenario, especially in an intricate setting such as chronic kidney disease. Since studying complex mechanisms of inflammation in vivo remains challenging, resource-intensive and usually requires the usage of transgenic animals, primary isolated hepatocytes provide a robust tool to gain mechanistic insights into the hepatic acute-phase response. Since this in vitro technique features moderate costs, high reproducibility and common technical knowledge, primary isolated hepatocytes can also be easily used as a screening approach. Here, we describe an enzymatic-based method to isolate primary murine hepatocytes, and we describe the assessment of an inflammatory response in these cells using ELISA and quantitative real-time PCR.