RESUMEN
BACKGROUND: Recent findings support a connection between mitochondrial dysfunction and activation of inflammatory pathways in articular cells. This study investigates in vivo in an acute model whether intra-articular administration of oligomycin, an inhibitor of mitochondrial function, induces an oxidative and inflammatory response in rat knee joints. METHODS: Oligomycin was injected into the rat left knee joint on days 0, 2, and 5 before joint tissues were obtained on day 6. The right knee joint served as control. Results were evaluated by macroscopy and histopathology and by measuring cellular and mitochondrial reactive oxygen species (ROS), 4-hydroxy-2-nonenal (4-HNE, a marker of lipid peroxidation), nuclear factor erythroid 2-related factor 2 (Nrf2), and CD68 (macrophages) and chemokine levels. The marker of mitochondrial mass COX-IV was also evaluated. RESULTS: The macroscopic findings showed significantly greater swelling in oligomycin-injected knees than in control knees. Likewise, the histological score of synovial damage was also increased significantly. Immunohistochemical studies showed high expression of IL-8, coinciding with a marked infiltration of polymorphonuclears and CD68+ cells in the synovium. Mitochondrial mass was increased in the synovium of oligomycin-injected joints, as well as cellular and mitochondrial ROS production, and 4-HNE. Relatedly, expression of the oxidative stress-related transcription factor Nrf2 was also increased. As expected, no histological differences were observed in the cartilage; however, cytokine-induced neutrophil chemoattractant-1 mRNA and protein expression were up-regulated in this tissue. CONCLUSIONS: Mitochondrial failure in the joint is able to reproduce the oxidative and inflammatory status observed in arthritic joints.
Asunto(s)
Artritis Experimental/patología , Inhibidores Enzimáticos/farmacología , Articulación de la Rodilla/patología , Mitocondrias/efectos de los fármacos , Osteoartritis/patología , Anciano , Anciano de 80 o más Años , Aldehídos/metabolismo , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/metabolismo , Artritis Experimental/inducido químicamente , Cartílago Articular/patología , Quimiocina CXCL1/metabolismo , Complejo IV de Transporte de Electrones/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Inyecciones Intraarticulares , Interleucina-8/metabolismo , Macrófagos/metabolismo , Persona de Mediana Edad , Mitocondrias/metabolismo , ATPasas de Translocación de Protón Mitocondriales/antagonistas & inhibidores , Factor 2 Relacionado con NF-E2/metabolismo , Oligomicinas/farmacología , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Membrana Sinovial/patologíaRESUMEN
OBJECTIVE: The present study aimed to determine the protective effects of dietary supplementation with resveratrol (RSV) in an acute antigen-induced arthritis (AIA) model. METHODS: Rats were randomly divided into three groups: control, AIA and RSV-treated AIA group. RSV (12.5 mg/kg/day) was given orally for 8 weeks before induction of AIA and until the end of the experiment (48 h after intra-articular injection). The control and AIA animals were administered 100 µl of water. Results were evaluated by macroscopic observation, histopathology and immunohistochemistry for anti-PCNA, macrophages (CD68), T lymphocytes (CD3), monocyte chemoattractant protein-1 and 8-oxo-7,8-dihydro-2'-deoxyguanine (a marker of DNA damage). Cytokine-induced neutrophil chemoattractant-1 in serum and peroxidase activity in synovial tissue were measured using commercial kits. RESULTS: At the end of the study, RSV significantly reduced knee swelling. Likewise, the histological score of synovial tissue also reduced significantly. The arthritis-protective effects were associated with a significant decrease in PCNA, CD68, CD3 and monocyte chemoattractant protein-1 staining, as well as a reduction in serum concentrations of cytokine-induced neutrophil chemoattractant-1. RSV treatment also decreased the level of the marker of DNA damage, 8-oxo-7,8-dihydro-2'-deoxyguanine. Accordingly, peroxidase activity in the synovial tissue was up-regulated. CONCLUSION: Dietary supplementation with RSV lowers the main pathological hallmarks of RA disease in an acute model of AIA. RSV may represent a promising strategy in controlling the severity of RA.
Asunto(s)
Antioxidantes/farmacología , Artritis Experimental/tratamiento farmacológico , Estilbenos/farmacología , Membrana Sinovial/patología , Animales , Artritis Experimental/inmunología , Artritis Experimental/patología , Proliferación Celular/efectos de los fármacos , Citocinas/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Hiperplasia/inmunología , Hiperplasia/prevención & control , Inmunidad Celular , Peroxidasa/antagonistas & inhibidores , Distribución Aleatoria , Ratas Endogámicas Lew , Resveratrol , Membrana Sinovial/inmunologíaRESUMEN
OBJECTIVES: In RA, synoviocytes cause increased oxidative stress, leading to mitochondrial alterations that may participate in the pathogenesis of RA. Here we investigated whether mitochondrial dysfunction induces inflammatory responses in cultured normal human synoviocytes, a hallmark of RA. METHODS: Mitochondrial dysfunction was induced with the inhibitor oligomycin. The effects of mitochondrial dysfunction on cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2) and IL-8 expression; cellular and mitochondrial reactive oxygen species (ROS) production; nuclear factor-κB (NF-κB) activation and p65 translocation were studied. ROS scavengers (N-acetylcysteine and mitoTEMPO) and an NF-κB inhibitor (BAY-117085) were used to investigate the pathways involved. The natural anti-inflammatory antioxidant resveratrol was also tested. RESULTS: Mitochondrial dysfunction per se significantly stimulated mitochondrial ROS production as well as low-grade expressions of COX-2, PGE2 and IL-8. Interestingly, mitochondrial dysfunction induced by pretreatment of synoviocytes with oligomycin synergized with IL-1ß to increase the expression of these inflammatory mediators. The inflammatory effects of mitochondrial damage appeared to be dependent on ROS production and NF-κB activation since the inflammatory response was counteracted by both N-acetylcysteine and mitoTEMPO and it was also reduced by BAY-117085. Antimycin A and paraquat (inhibitors of mitochondrial function) also induced inflammatory responses. Furthermore, resveratrol significantly reduced the inflammatory response by decreasing ROS production and NF-κB activation. CONCLUSION: These data suggest that mitochondrial dysfunction could induce an inflammatory response in normal human synoviocytes and sensitize these cells, causing a significant amplification of the inflammatory response induced by IL-1ß. Resveratrol may represent a promising strategy in controlling the synovial inflammatory response.
Asunto(s)
Inhibidores Enzimáticos/farmacología , Inflamación/fisiopatología , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiología , Oligomicinas/farmacología , Membrana Sinovial/patología , Membrana Sinovial/fisiopatología , Anciano , Artritis Reumatoide/metabolismo , Artritis Reumatoide/patología , Artritis Reumatoide/fisiopatología , Células Cultivadas , Ciclooxigenasa 2/metabolismo , Dinoprostona/metabolismo , Humanos , Inflamación/metabolismo , Interleucina-1beta/farmacología , Interleucina-8/metabolismo , Persona de Mediana Edad , FN-kappa B/metabolismo , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Especies Reactivas de Oxígeno/metabolismo , Resveratrol , Estilbenos/farmacología , Membrana Sinovial/efectos de los fármacosRESUMEN
During peritoneal dialysis (PD), mesothelial cells undergo mesothelial-to-mesenchymal transition (MMT), a process associated with peritoneal-membrane dysfunction. Because TGF-ß1 can induce MMT, we evaluated the efficacy of TGF-ß1-blocking peptides in modulating MMT and ameliorating peritoneal damage in a mouse model of PD. Exposure of the peritoneum to PD fluid induced fibrosis, angiogenesis, functional impairment, and the accumulation of fibroblasts. In addition to expressing fibroblast-specific protein-1 (FSP-1), some fibroblasts co-expressed cytokeratin, indicating their mesothelial origin. These intermediate-phenotype (Cyto(+)/FSP-1(+)) fibroblasts had features of myofibroblasts with fibrogenic capacity. PD fluid treatment triggered the appearance of CD31(+)/FSP-1(+) and CD45(+)/FSP-1(+) cells, suggesting that fibroblasts also originate from endothelial cells and from cells recruited from bone marrow. Administration of blocking peptides significantly ameliorated fibrosis and angiogenesis, improved peritoneal function, and reduced the number of FSP-1(+) cells, especially in the Cyto(+)/FSP-1(+) subpopulation. Conversely, overexpression of TGF-ß1 in the peritoneum by adenovirus-mediated gene transfer led to a marked accumulation of fibroblasts, most of which derived from the mesothelium. Taken together, these results demonstrate that TGF-ß1 drives the peritoneal deterioration induced by dialysis fluid and highlights a role of TGF-ß1-mediated MMT in the pathophysiology of peritoneal-membrane dysfunction.
Asunto(s)
Transdiferenciación Celular/efectos de los fármacos , Diálisis Peritoneal/efectos adversos , Fibrosis Peritoneal/etiología , Peritoneo/patología , Factor de Crecimiento Transformador beta1/metabolismo , Animales , Biomarcadores/metabolismo , Células Cultivadas , Soluciones para Diálisis/efectos adversos , Femenino , Inyecciones Intraperitoneales , Queratinas/metabolismo , Ratones , Ratones Endogámicos C57BL , Miofibroblastos/metabolismo , Miofibroblastos/patología , Fragmentos de Péptidos/farmacología , Fragmentos de Péptidos/uso terapéutico , Péptidos/farmacología , Péptidos/uso terapéutico , Fibrosis Peritoneal/metabolismo , Fibrosis Peritoneal/patología , Fibrosis Peritoneal/prevención & control , Fenotipo , Receptores de Factores de Crecimiento Transformadores beta/uso terapéutico , Proteína de Unión al Calcio S100A4 , Proteínas S100/metabolismo , Factor de Crecimiento Transformador beta1/antagonistas & inhibidoresRESUMEN
Exposure to non-physiological solutions during peritoneal dialysis (PD) produces structural alterations to the peritoneal membrane and ultrafiltration dysfunction. The high concentration of glucose and glucose degradation products in standard PD fluids induce a local diabetic environment, which leads to the formation of advanced glycation end products (AGEs) that have an important role in peritoneal membrane deterioration. Peroxisome proliferator-activated receptor γ (PPAR-γ) agonists are used to treat type II diabetes and they have beneficial effects on inflammation, fibrosis, and angiogenesis. Hence, we evaluated the efficacy of the PPAR-γ agonist rosiglitazone (RSG) in ameliorating peritoneal membrane damage in a mouse PD model, and we analyzed the mechanisms underlying the protection offered by RSG. Exposure of the peritoneum to PD fluid resulted in AGEs accumulation, an inflammatory response, the loss of mesothelial cell monolayer and invasion of the compact zone by mesothelial cells, fibrosis, angiogenesis, and functional impairment of the peritoneum. Administration of RSG diminished the accumulation of AGEs, preserved the mesothelial monolayer, decreased the number of invading mesothelial cells, reduced fibrosis and angiogenesis, and improved peritoneal function. Interestingly, instead of reducing the leukocyte recruitment, RSG administration enhanced this process and specifically, the recruitment of CD3+ lymphocytes. Furthermore, RSG treatment augmented the levels of the anti-inflammatory cytokine interleukin (IL)-10 and increased the recruitment of CD4+ CD25+ FoxP3+ cells, suggesting that regulatory T cells mediated the protection of the peritoneal membrane. In cell-culture experiments, RSG did not prevent or reverse the mesothelial to mesenchymal transition, although it decreased mesothelial cells apoptosis. Accordingly, RSG appears to produce pleiotropic protective effects on the peritoneal membrane by reducing the accumulation of AGEs and inflammation, and by preserving the mesothelial cells monolayer, highlighting the potential of PPAR-γ activation to ameliorate peritoneal deterioration in PD patients.
Asunto(s)
Soluciones para Diálisis/efectos adversos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , PPAR gamma/agonistas , Diálisis Peritoneal/efectos adversos , Peritoneo/efectos de los fármacos , Peritoneo/patología , Tiazolidinedionas/farmacología , Animales , Células Epiteliales/metabolismo , Transición Epitelial-Mesenquimal/efectos de los fármacos , Epitelio/efectos de los fármacos , Epitelio/patología , Fibrosis , Glucosa/metabolismo , Productos Finales de Glicación Avanzada/metabolismo , Inmunidad Celular/efectos de los fármacos , Inflamación , Ratones , PPAR gamma/metabolismo , Peritoneo/inmunología , Peritoneo/metabolismo , Rosiglitazona , Linfocitos T/inmunologíaRESUMEN
BACKGROUND: During peritoneal dialysis (PD), mesothelial cells (MC) undergo an epithelial-to-mesenchymal transition (EMT), and this process is associated with peritoneal membrane (PM) damage. Bone morphogenic protein-7 (BMP-7) antagonizes transforming growth factor (TGF)-beta1, modulates EMT and protects against fibrosis. Herein, we analysed the modulating role of BMP-7 on EMT of MC in vitro and its protective effects in a rat PD model. METHODS: Epitheliod or non-epitheliod MC were analysed for the expression of BMP-7, TGF-beta1, activated Smads, epithelial cadherin (E-cadherin), collagen I, alpha smooth muscle cell actin (alpha-SMA) and vascular endothelial growth factor (VEGF) using standard procedures. Rats were daily instilled with PD fluid with or without BMP-7 during 5 weeks. Histological analyses were carried out in parietal peritoneum. Fibrosis was quantified with van Gieson or Masson's trichrome staining. Vasculature, activated macrophages and invading MC were quantified by immunofluorescence analysis. Quantification of infiltrating leukocytes and MC density in liver imprints was performed by May-Grünwald-Giemsa staining. Hyaluronic acid levels were determined by ELISA. RESULTS: MC constitutively expressed BMP-7, and its expression was downregulated during EMT. Treatment with recombinant BMP-7 resulted in blockade of TGF-beta1-induced EMT of MC. We provide evidence of a Smad-dependent mechanism for the blockade of EMT. Exposure of rat peritoneum to PD fluid resulted in inflammatory and regenerative responses, invasion of the compact zone by MC, fibrosis and angiogenesis. Administration of BMP-7 decreased the number of invading MC and reduced fibrosis and angiogenesis. In contrast, BMP-7 had no effect on inflammatory and regenerative responses, suggesting that these are EMT-independent, and probably upstream, processes. CONCLUSIONS: Data point to a balance between BMP-7 and TGF-beta1 in the control of EMT and indicate that blockade of EMT may be a therapeutic approach to ameliorate peritoneal membrane damage during PD.
Asunto(s)
Proteína Morfogenética Ósea 7/fisiología , Epitelio/metabolismo , Mesodermo/metabolismo , Diálisis Peritoneal , Peritoneo/metabolismo , Actinas/metabolismo , Animales , Western Blotting , Cadherinas/metabolismo , Diferenciación Celular , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Fibrosis , Humanos , Técnicas para Inmunoenzimas , Masculino , Ratas , Ratas Wistar , Proteínas Smad/metabolismo , Factor de Crecimiento Transformador beta1/antagonistas & inhibidoresRESUMEN
During peritoneal dialysis (PD), exposure of the peritoneal membrane to nonphysiologic solutions causes inflammation, ultimately leading to altered structure and function. Myofibroblasts, one of the cell types that contribute to dysfunction of the peritoneal membrane, can originate from mesothelial cells (MCs) by epithelial-to-mesenchymal transition (EMT), a process that has been associated with an increased rate of peritoneal transport. Because cyclooxygenase-2 (COX-2) is induced by inflammation, we studied the role of COX-2 in the deterioration of the peritoneal membrane. We observed that nonepithelioid MCs found in peritoneal effluent expressed higher levels of COX-2 than epithelioid MCs. The mass transfer coefficient for creatinine correlated with MC phenotype and with COX-2 levels. Although COX-2 was upregulated during EMT of MCs in vitro, COX-2 inhibition did not prevent EMT. In a mouse model of PD, however, COX-2 inhibition with Celecoxib resulted in reduced fibrosis and in partial recovery of ultrafiltration, outcomes that were associated with a reduction of inflammatory cells. Furthermore, PD fluid with a low content of glucose degradation products did not induce EMT or COX-2; the peritoneal membranes of mice treated with this fluid showed less worsening than mice exposed to standard fluid. In conclusion, upregulation of COX-2 during EMT may mediate peritoneal inflammation, suggesting COX-2 inhibition as a potential strategy to ameliorate peritoneal deterioration in PD patients.
Asunto(s)
Ciclooxigenasa 2/metabolismo , Soluciones para Diálisis/efectos adversos , Diálisis Peritoneal/efectos adversos , Peritoneo/enzimología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Transporte Biológico Activo , Celecoxib , Células Cultivadas , Ciclooxigenasa 2/genética , Inhibidores de la Ciclooxigenasa 2/farmacología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/enzimología , Células Epiteliales/patología , Epitelio/efectos de los fármacos , Epitelio/enzimología , Epitelio/patología , Femenino , Humanos , Inflamación/prevención & control , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Modelos Animales , Peritoneo/efectos de los fármacos , Peritoneo/patología , Peritoneo/fisiopatología , Pirazoles/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción de la Familia Snail , Sulfonamidas/farmacología , Factores de Transcripción/genética , Regulación hacia Arriba , Adulto JovenRESUMEN
BACKGROUND: Peritoneal dialysis (PD) is associated with functional and structural alterations of the peritoneal membrane, particularly new vessel formation and fibrosis. In addition to anticoagulant effects, heparin displays anti-inflammatory and angiostatic properties. Therefore, the effects of administration of heparins on function and morphology of the peritoneal membrane were studied in a rat PD model. METHODS: Rats received 10 mL conventional PD fluid (PDF) daily, with or without the addition of unfractionated heparin (UFH) or low molecular weight heparin (LMWH) in the PDF (1 mg/10 mL intraperitoneally) via a mini access port. Untreated rats served as controls. After 5 weeks, a 90-minute functional peritoneal transport test was performed and tissues and peritoneal leukocytes were taken. RESULTS: PD treatment induced loss of ultrafiltration (p<0.01), a twofold increase in glucose absorption (p<0.03), increased urea transport (p<0.02), and loss of sodium sieving (p<0.03), which were also found in the PDF+heparin groups. Increased peritoneal cell influx and hyaluronan production (p<0.02) as well as an exchange of mast cells and eosinophils for neutrophils after PD treatment were observed in PD rats; addition of heparin did not affect those changes. Mesothelial regeneration, submesothelial blood vessel and matrix formation, and accumulation of tissue macrophages were seen in PD animals. Spindle-shaped vimentin-positive and cytokeratin-negative cells indicated either partial injury and denudation of mesothelial cells or epithelial-to-mesenchymal transition. Neither UFH nor LMWH affected any of these morphological changes. CONCLUSION: Within 5 weeks, PD treatment induces a chronic inflammatory condition in the peritoneum, evidenced by high transport, leukocyte recruitment, tissue remodeling, and induction of spindle-shaped cells in the mesothelium. Addition of LMWH or UFH to the PDF did not prevent these adverse PDF-induced peritoneal changes.
Asunto(s)
Anticoagulantes/administración & dosificación , Heparina/administración & dosificación , Diálisis Peritoneal/métodos , Peritoneo/metabolismo , Animales , Anticoagulantes/farmacocinética , Transporte Biológico/fisiología , Modelos Animales de Enfermedad , Fibrosis/etiología , Fibrosis/patología , Fibrosis/prevención & control , Estudios de Seguimiento , Heparina/farmacocinética , Fallo Renal Crónico/metabolismo , Fallo Renal Crónico/terapia , Masculino , Mesenterio/efectos de los fármacos , Mesenterio/metabolismo , Mesenterio/patología , Neovascularización Patológica/patología , Neovascularización Patológica/prevención & control , Epiplón/efectos de los fármacos , Epiplón/metabolismo , Epiplón/patología , Diálisis Peritoneal/efectos adversos , Peritoneo/efectos de los fármacos , Peritoneo/patología , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
Animal models of peritoneal dialysis fluid (PDF) exposure are key tools in the study of mechanisms involved in alterations of the peritoneal membrane and in the design of therapies. We recently developed a mouse model of chronic peritoneal exposure to high glucose dialysate. Herein, we make a sequential analysis of the effects of glucose-based PDF on mouse peritoneal membrane and on mesothelium. We demonstrate that chronic exposure to PDF induces thickness and fibrosis of the peritoneal membrane in a time-dependent manner. We also show that mesothelial cells progressively detach and lose cytokeratin expression. In addition, we demonstrate that some mesothelial cells invade the submesothelial space, where they appear as cytokeratin- and alpha-smooth muscle actin-positive cells. These findings demonstrate that epithelial-to-mesenchymal transition (EMT) of mesothelial cells takes place in mouse peritoneum exposed to PDF, validating this model for the study of effects of drugs on the EMT process as a therapy for peritoneal deterioration.
Asunto(s)
Soluciones para Diálisis/farmacología , Células Epiteliales/efectos de los fármacos , Glucosa/farmacología , Mesodermo/efectos de los fármacos , Diálisis Peritoneal/efectos adversos , Peritoneo/efectos de los fármacos , Animales , Diferenciación Celular/efectos de los fármacos , Soluciones para Diálisis/administración & dosificación , Soluciones para Diálisis/química , Modelos Animales de Enfermedad , Femenino , Glucosa/administración & dosificación , Mesodermo/patología , Ratones , Ratones Endogámicos C57BL , Peritoneo/patología , Factores de TiempoRESUMEN
Ageing processes play a major contributing role for the development of Osteoarthritis (OA). This prototypic degenerative condition of ageing is the most common form of arthritis and is accompanied by a general decline, chronic pain and mobility deficits. The disease is primarily characterized by articular cartilage degradation, followed by subchondral bone thickening, osteophyte formation, synovial inflammation and joint degeneration. In the early stages, osteoarthritic chondrocytes undergo phenotypic changes that increase cell proliferation and cluster formation and enhance the production of matrix-remodelling enzymes. In fact, chondrocytes exhibit differentiation plasticity and undergo phenotypic changes during the healing process. Current studies are focusing on unravelling whether OA is a consequence of an abnormal wound healing response. Recent investigations suggest that alterations in different proteins, such as TGF-ß/BMPs, NF-Kß, Wnt, and Cx43, or SASP factors involved in signalling pathways in wound healing response, could be directly implicated in the initiation of OA. Several findings suggest that osteoarthritic chondrocytes remain in an immature state expressing stemness-associated cell surface markers. In fact, the efficacy of new disease-modifying OA drugs that promote chondrogenic differentiation in animal models indicates that this may be a drug-sensible state. In this review, we highlight the current knowledge regarding cellular plasticity in chondrocytes and OA. A better comprehension of the mechanisms involved in these processes may enable us to understand the molecular pathways that promote abnormal repair and cartilage degradation in OA. This understanding would be advantageous in identifying novel targets and designing therapies to promote effective cartilage repair and successful joint ageing by preventing functional limitations and disability.
Asunto(s)
Envejecimiento/metabolismo , Cartílago Articular/fisiología , Diferenciación Celular/fisiología , Plasticidad de la Célula/fisiología , Osteoartritis/metabolismo , Regeneración/fisiología , Envejecimiento/patología , Animales , Cartílago Articular/patología , Proliferación Celular/fisiología , Condrocitos/metabolismo , Condrocitos/patología , Humanos , Osteoartritis/patología , Transducción de Señal/fisiologíaRESUMEN
Peritoneal dialysis (PD) is a form of renal replacement therapy whose repeated use can alter dialytic function through induction of epithelial-mesenchymal transition (EMT) and fibrosis, eventually leading to PD discontinuation. The peritoneum from Cav1-/- mice showed increased EMT, thickness, and fibrosis. Exposure of Cav1-/- mice to PD fluids further increased peritoneal membrane thickness, altered permeability, and increased the number of FSP-1/cytokeratin-positive cells invading the sub-mesothelial stroma. High-throughput quantitative proteomics revealed increased abundance of collagens, FN, and laminin, as well as proteins related to TGF-ß activity in matrices derived from Cav1-/- cells. Lack of Cav1 was associated with hyperactivation of a MEK-ERK1/2-Snail-1 pathway that regulated the Smad2-3/Smad1-5-8 balance. Pharmacological blockade of MEK rescued E-cadherin and ZO-1 inter-cellular junction localization, reduced fibrosis, and restored peritoneal function in Cav1-/- mice. Moreover, treatment of human PD-patient-derived MCs with drugs increasing Cav1 levels, as well as ectopic Cav1 expression, induced re-acquisition of epithelial features. This study demonstrates a pivotal role of Cav1 in the balance of epithelial versus mesenchymal state and suggests targets for the prevention of fibrosis during PD.
Asunto(s)
Caveolina 1/deficiencia , Transición Epitelial-Mesenquimal , Diálisis Peritoneal/efectos adversos , Peritoneo/fisiopatología , Factores de Transcripción/metabolismo , Animales , Caveolina 1/genética , Células Epiteliales/metabolismo , Fibrosis , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , Ratones Noqueados , Proteína Quinasa 1 Activada por Mitógenos/genética , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Peritoneo/enzimología , Peritoneo/metabolismo , Peritoneo/patología , Proteínas Smad/genética , Proteínas Smad/metabolismo , Factores de Transcripción de la Familia Snail , Factores de Transcripción/genética , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
Preservation of peritoneal membrane (PM) is essential for long-term survival in peritoneal dialysis (PD). Continuous presence of PD fluids (PDF) in the peritoneal cavity generates chronic inflammation and promotes changes of the PM, such as fibrosis, angiogenesis, and lymphangiogenesis. Mesothelial-to-mesenchymal transition (MMT) and endothelial-to-mesenchymal transition (Endo-MT) seem to play a central role in this pathogenesis. We speculated that Rapamycin, a potent immunosuppressor, could be beneficial by regulating blood and lymphatic vessels proliferation. We demonstrate that mice undergoing a combined PD and Rapamycin treatment (PDF + Rapa group) presented a reduced PM thickness and lower number of submesothelial blood and lymphatic vessels, as well as decreased MMT and Endo-MT, comparing with their counterparts exposed to PD alone (PDF group). Peritoneal water transport in the PDF + Rapa group remained at control level, whereas PD effluent levels of VEGF, TGF-ß, and TNF-α were lower than in the PDF group. Moreover, the treatment of mesothelial cells with Rapamycin in vitro significantly decreased VEGF synthesis and selectively inhibited the VEGF-C and VEGF-D release when compared with control cells. Thus, Rapamycin has a protective effect on PM in PD through an antifibrotic and antiproliferative effect on blood and lymphatic vessels. Moreover, it inhibits Endo-MT and, at least partially, MMT.
Asunto(s)
Transición Epitelial-Mesenquimal/efectos de los fármacos , Linfangiogénesis/efectos de los fármacos , Membranas Artificiales , Neovascularización Fisiológica/efectos de los fármacos , Diálisis Peritoneal/efectos adversos , Sirolimus/farmacología , Animales , Citocinas/sangre , Femenino , RatonesRESUMEN
Mesothelial-to-mesenchymal transition (MMT) is an autoregulated physiological process of tissue repair that in uncontrolled conditions, such as peritoneal dialysis (PD), can lead to peritoneal fibrosis. The maximum expression of sclerotic peritoneal syndromes (SPS) is the encapsulating peritoneal sclerosis (EPS) for which no specific treatment exists. The SPS includes a wide range of peritoneal fibrosis that appears progressively and is considered as a reversible process, while EPS does not. EPS is a serious complication of PD characterized by a progressive intra-abdominal inflammatory process that results in bridles and severe fibrous tissue formation which cover and constrict the viscera. Recent studies show that transdifferentiated mesothelial cells isolated from the PD effluent correlate very well with the clinical events such as the number of hemoperitoneum and peritonitis, as well as with PD function (lower ultrafiltration and high Cr-MTC). In addition, in peritoneal biopsies from PD patients, the MMT correlates very well with anatomical changes (fibrosis and angiogenesis). However, the pathway to reach EPS from SPS has not been fully and completely established. Herein, we present important evidence pointing to the MMT that is present in the initial peritoneal fibrosis stages and it is perpetual over time, with at least theoretical possibility that MMT initiated the fibrosing process to reach EPS.
RESUMEN
Mesothelial-to-mesenchymal transition (MMT) is an auto-regulated physiological process of tissue repair that in uncontrolled conditions such as peritoneal dialysis (PD) can lead to peritoneal fibrosis. The maximum expression of peritoneal fibrosis induced by PD fluids and other peritoneal processes is the encapsulating peritoneal sclerosis (EPS) for which no specific treatment exists. Tamoxifen, a synthetic estrogen, has successfully been used to treat retroperitoneal fibrosis and EPS associated with PD. Hence, we used in vitro and animal model approaches to evaluate the efficacy of Tamoxifen to inhibit the MMT as a trigger of peritoneal fibrosis. In vitro studies were carried out using omentum-derived mesothelial cells (MCs) and effluent-derived MCs. Tamoxifen blocked the MMT induced by transforming growth factor (TGF)-ß1, as it preserved the expression of E-cadherin and reduced the expression of mesenchymal-associated molecules such as snail, fibronectin, collagen-I, α-smooth muscle actin, and matrix metalloproteinse-2. Tamoxifen-treatment preserved the fibrinolytic capacity of MCs treated with TGF-ß1 and decreased their migration capacity. Tamoxifen did not reverse the MMT of non-epitheliod MCs from effluents, but it reduced the expression of some mesenchymal molecules. In mice PD model, we demonstrated that MMT progressed in parallel with peritoneal membrane thickness. In addition, we observed that Tamoxifen significantly reduced peritoneal thickness, angiogenesis, invasion of the compact zone by mesenchymal MCs and improved peritoneal function. Tamoxifen also reduced the effluent levels of vascular endothelial growth factor and leptin. These results demonstrate that Tamoxifen is a therapeutic option to treat peritoneal fibrosis, and that its protective effect is mediated via modulation of the MMT process.