RESUMO
Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) is a member of the TNF superfamily. It has been suggested that it plays a pivotal role in various physiological and pathological conditions due to its proinflammatory properties. Fibroblast growth-inducible 14 (Fn14) has been identified as a TWEAK receptor. A number of studies have suggested that TWEAK-Fn14 interaction results in the promotion of apoptosis, cell growth as well as angiogenesis. Although recent studies have indicated that TWEAK and Fn14 are expressed in a number of tumor lines and tissues, the therapeutic potential of this pathway has yet to be elucidated. This study investigated the potential of TWEAK and Fn14 in esophageal and pancreatic cancer as novel molecular targets for anti-cancer therapy. TWEAK and Fn14 protein expression was evaluated in 43 patients with esophageal cancer and 51 patients with pancreatic cancer by immunohistochemistry. As a result, either TWEAK or Fn14 expression was observed in 58.1% of the cases with esophageal cancer and 74.5% of the cases with pancreatic cancer. Furthermore, TWEAK/Fn14 gene expression was identified in the majority of the human esophageal and pancreatic cancer cell lines. Therapeutic efficacies of blocking TWEAK and Fn14 were evaluated by tumor growth inhibition assay in TWEAK- and Fn14-expressing human esophageal and pancreatic cancer cell lines. Coculture with anti-TWEAK or -Fn14 mAb was found to induce a 22-65% cell growth inhibition of these cells. Finally, the significant therapeutic effect of targeting this pathway under in vivo physiological conditions was confirmed using a murine gastrointestinal cancer model. In conclusion, the TWEAK/Fn14 pathway may be functional and critical in intractable gastrointestinal cancers. Therefore, TWEAK and/or Fn14 may be novel molecular targets for anti-cancer therapy.
RESUMO
BACKGROUND: Clinical studies suggest a role for angiogenesis in the development and persistence of chronic asthma, but whether angiogenic mediators contribute to acute asthma has not been fully studied. OBJECTIVE: The aim of this study was to investigate a role of vascular endothelial growth factor (VEGF), a major angiogenic and proinflammatory mediator, in allergen-induced acute asthma and to determine whether endostatin/Fc, a potent antiangiogenic factor can attenuate allergic airway responses. METHODS: We sensitized BALB/c mice with ovalbumin. We measured serum VEGF and examined immunoreactive VEGF around the airways 48 hours after the last challenge with either aerosolized PBS or ovalbumin once per day for 3 days. We also treated ovalbumin-sensitized mice with either endostatin/Fc or control fusion protein at the time of challenge with ovalbumin. We analyzed allergic airway responses 48 hours after the last ovalbumin challenge. RESULTS: Ovalbumin challenge induced immunolocalization of numerous VEGF-positive cells around airways and increased serum VEGF levels. Treatment with endostatin/Fc inhibited the airway hyperresponsiveness, pulmonary allergic inflammation, production of ovalbumin-specific IgE, and lung inflammatory mediators. Both VEGF-dependent and independent mechanisms are indicated by results using antibody blockade of VEGF receptors, which caused decreased allergic pulmonary inflammation but did not alter airway hyperresponsiveness or serum IgE levels. CONCLUSION: These data demonstrate for the first time that recombinant endostatin can prevent the development of asthma features in a mouse model and suggest that this class of agents merits further study as novel therapeutics for asthma.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Asma/prevenção & controle , Endostatinas/uso terapêutico , Animais , Asma/imunologia , Asma/fisiopatologia , Feminino , Mediadores da Inflamação/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/sangue , Fator A de Crescimento do Endotélio Vascular/fisiologiaRESUMO
BACKGROUND: We have recently reported that vascular endothelial growth factor (VEGF) functions as a proinflammatory cytokine to regulate the trafficking of leukocytes into allografts in the early posttransplant period. VEGF binds two major VEGF receptors: VEGFR-1 (flt-1) and VEGFR-2 (flk-1/KDR). Here, we wished to investigate the expression and function of VEGF receptors in the process of acute allograft rejection in vivo. METHODS: We performed fully MHC-mismatched C57BL/6 (H-2b) into BALB/c (H-2d) vascularized heterotopic murine cardiac transplants and we examined the expression of VEGF and VEGF receptors by immunohistochemistry during acute allograft rejection. Next, we treated mice with specific neutralizing monoclonal antibodies against murine VEGFR-1 and VEGFR-2 and examined their effect on the development of acute allograft rejection by histology and by analysis of graft survival. The intragraft expression of cytokines and chemokines were also evaluated by quantitative real-time PCR analysis. RESULTS: The expression of VEGF, VEGFR-1 and VEGFR-2 were significantly up-regulated during allograft rejection as compared to isografts. Administration of either anti-VEGFR-1 or anti-VEGFR-2 alone failed to inhibit allograft rejection. However, coadministration of both antibodies together inhibited leukocyte infiltration of allografts and prolonged allograft survival. Furthermore, the effect of VEGFR blockade was associated with the downregulation of intragraft cytokine and chemokine expression. CONCLUSIONS: Our data suggest that VEGF-VEGFR interactions function in the alloimmune response in vivo. Targeting VEGFRs may represent a novel therapy to protect allografts following clinical transplantation.
Assuntos
Rejeição de Enxerto/imunologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/imunologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Doença Aguda , Animais , Quimiocinas/genética , Citocinas/genética , Imuno-Histoquímica , Camundongos , RNA Mensageiro/genética , Transplante Homólogo , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Prostaglandin E2 (PGE2) mediates a variety of innate and adaptive immunity through four distinct receptors: EP1-EP4. It has been suggested that each EP plays a unique and pivotal role in various disease conditions. We investigated the pathophysiological role of EP receptors in hepatic ischemia/reperfusion (I/R) injury. In this study, a 70% hepatic ischemic model was used in male C57BL/6 mice. Selective EP agonists were used to clarify the function of each PGE2 receptor in I/R injury. Although all four receptors were expressed in the naïve liver, EP4 expression was significantly upregulated after hepatic I/R. Although EP1, 2, or 3 agonists did not show any protective effect on liver function, the EP4 agonist significantly inhibited hepatic I/R injury as determined by serological and histological analyses. Furthermore, the EP4 agonist downregulated the local expressions of several proinflammatory cytokines, chemokines, and adhesion molecules in the early phase of reperfusion. In contrast, it augmented the local expression of an anti-inflammatory cytokine, interleukin 10. Additionally, the neutrophil accumulation was also inhibited by EP4 agonist treatment. Finally, to confirm the therapeutic efficacy of the EP4 agonist in hepatic I/R injury, the nonischemic shunt liver was removed after 120 minutes of ischemia, resulting in the death of 86% of control mice within 48 hours. In sharp contrast, 80% of mice treated with the EP4 agonist survived. In conclusion, the PGE2-EP4 signaling pathway has an inhibitory role in hepatic I/R injury. An EP4 agonist effectively protects against ischemic injury.
Assuntos
Circulação Hepática , Fígado/irrigação sanguínea , Receptores de Prostaglandina E/fisiologia , Traumatismo por Reperfusão/prevenção & controle , Alanina Transaminase/metabolismo , Animais , Aspartato Aminotransferases/metabolismo , Regulação da Expressão Gênica/fisiologia , Testes de Função Hepática , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Prostaglandina E/agonistas , Receptores de Prostaglandina E/genética , Traumatismo por Reperfusão/fisiopatologiaRESUMO
BACKGROUND: Chemokines and chemokine receptors are critical in leukocyte recruitment, activation, and differentiation. Among them, CC chemokine receptor 5 (CCR5) and CXC chemokine receptor 3 (CXCR3) have been reported to play important roles in alloimmune responses and may be potential targets for posttransplant immunosuppression. METHODS: Fully major histocompatibility complex (MHC)-mismatched murine cardiac and islet transplant models were used to test the effect in vivo of a novel, small-molecule compound TAK-779 by targeting CCR5 and CXCR3 in acute allograft rejection. An MHC class II mismatched cardiac transplant model was used to evaluate its efficacy in chronic allograft rejection. Intragraft expression of cytokines, chemokines, and chemokine receptors was measured by quantitative real-time polymerase chain reaction and by histological analysis. RESULTS: Treatment of TAK-779 significantly prolonged allograft survival across the MHC barrier in two distinct transplant models. The treatment downregulated local immune activation as observed by the reduced expression of several chemokines, cytokines, and chemokine receptors. Thereby, the recruitment of CD4, CD8, and CD11c cells into transplanted allografts were inhibited. Furthermore, TAK-779 treatment significantly attenuated the development of chronic vasculopathy, fibrosis, and cellular infiltration. CONCLUSIONS: Antagonism of CCR5 and CXCR3 has a substantial therapeutic effect on inhibiting both acute and chronic allograft rejection. CCR5 and CXCR3 are functional in the process of allograft rejection and may be potential targets in clinical transplantation in the future.
Assuntos
Amidas/farmacologia , Antagonistas dos Receptores CCR5 , Rejeição de Enxerto , Sobrevivência de Enxerto , Transplante de Coração/métodos , Transplante das Ilhotas Pancreáticas/métodos , Compostos de Amônio Quaternário/farmacologia , Receptores CCR5/química , Receptores de Quimiocinas/química , Amidas/química , Animais , Antígeno CD11c/biossíntese , Antígenos CD4/biossíntese , Antígenos CD8/biossíntese , Transplante de Células , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase , Compostos de Amônio Quaternário/química , RNA Mensageiro/metabolismo , Receptores CXCR3 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Resultado do Tratamento , Regulação para CimaRESUMO
BACKGROUND: Inducible costimulator (ICOS) is the third member of the CD28 superfamily and has a unique role in T cell activation and function. Recent studies indicated that the ICOS-B7h pathway plays an important role in alloimmune responses. We further investigated the role of the ICOS pathway in the pathologic process of chronic rejection in vivo. METHODS: An established major histocompatibility complex class II disparate cardiac transplantation model was used. We treated mice with a blocking anti-B7h monoclonal antibody (mAb) either in the initiation phase (early blockade) or in the progression phase (delayed blockade) of disease. In addition, some mice received mAb in the entire period (whole blockade). At 6 weeks after transplantation, cardiac grafts were evaluated by histopathologic analysis in terms of vasculopathy, fibrosis, and cellular infiltration. The intragraft expressions of cytokines and chemokines were also examined by quantitative real-time polymerase chain reaction analysis. RESULTS: Early blockade of the ICOS-B7h pathway did not show any protective effect on chronic allograft rejection compared with untreated controls. In contrast, delayed blockade significantly inhibited the development of vasculopathy, fibrosis, and cellular infiltration (P=0.043, P=0.004, and P=0.03 vs. untreated control, respectively). Interestingly, whole blockade did not prevent the chronic rejection process. Furthermore, the inhibitory effect of delayed ICOS blockade on chronic rejection was associated with down-regulation of local intragraft expression of several cytokines and chemokines. CONCLUSIONS: These data suggest that the ICOS-B7h pathway is critical in the activation of effector/memory T cells that are necessary for the progression of chronic rejection and provide the rationale to develop novel and specific therapies to prevent this process.
Assuntos
Antígenos de Diferenciação de Linfócitos T/imunologia , Regulação da Expressão Gênica/imunologia , Rejeição de Enxerto/imunologia , Proteínas/imunologia , Linfócitos T/imunologia , Transplante Homólogo/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T/genética , Doença Crônica , Fibrose , Rejeição de Enxerto/patologia , Rejeição de Enxerto/fisiopatologia , Tolerância Imunológica , Memória Imunológica , Ligante Coestimulador de Linfócitos T Induzíveis , Proteína Coestimuladora de Linfócitos T Induzíveis , Camundongos , Camundongos Endogâmicos C57BL , Transplante Homólogo/patologiaRESUMO
BACKGROUND: Vascular endothelial growth factor (VEGF), a major angiogenic factor, mediates a variety of disease conditions through promotion of angiogenesis. It also plays a critical role as a potent proinflammatory cytokine in a variety of physiologic and pathologic immune responses. In the present study, we evaluated the expression of VEGF in hepatic warm ischemia-reperfusion (I/R) injury and examined the effect of recombinant human (rh)VEGF administration in an established murine model. METHOD: The expression of VEGF in the liver was assessed by quantitative real-time polymerase chain reaction and immunohistochemistry during I/R injury using 70% partial hepatic ischemia model. The effect of rhVEGF administration on I/R injury was evaluated by measuring liver function and histology. In addition, local inducible nitric oxide synthase (iNOS) and endothelial NO synthase expressions were examined to address the underlying mechanisms. RESULTS: The local expression of VEGF was significantly up-regulated at 2 hours after reperfusion after 60 minutes of ischemia compared with that in the naive liver. VEGF was expressed predominantly in CD11b+ cells infiltrating into the ischemic liver. The administration of rhVEGF had a significant protective effect on ischemic injury in the liver. This effect was associated with the up-regulation of iNOS expression in the rhVEGF-treated liver. CONCLUSION: We demonstrate a dual role of VEGF in hepatic warm I/R injury. Although endogenous VEGF is expressed and functional to initiate hepatic I/R injury, exogenous rhVEGF has a beneficial effect on the ischemic liver. These data may provide new insights into the role of VEGF as well as pathophysiology of hepatic I/R injury.
Assuntos
Circulação Hepática/fisiologia , Fígado/irrigação sanguínea , Traumatismo por Reperfusão/prevenção & controle , Fator A de Crescimento do Endotélio Vascular/genética , Animais , Regulação da Expressão Gênica/fisiologia , Humanos , Fígado/patologia , Camundongos , Camundongos Endogâmicos C57BL , Necrose , Óxido Nítrico Sintase/genética , Reação em Cadeia da Polimerase/métodos , Proteínas Recombinantes/farmacologia , Fator A de Crescimento do Endotélio Vascular/farmacologiaRESUMO
BACKGROUND: Establishment of hematopoietic chimerism is the most stable strategy for donor-specific tolerance. Safer pretreatment regimens are needed for clinical application. We evaluated the efficacy of a simple protocol using cyclophosphamide (CYP) on induction of chimerism and organ transplant tolerance across major histocompatibility complex (MHC) barriers in the rat. MATERIALS AND METHODS: Bone marrow cells from BN (RT1(n)) donors were infused to LEW (RT1(l)) recipients on day 0 after a single injection of CYP at various doses on day -1. Donor-derived hematopoietic chimerism was evaluated by flowcytometry. The recipients received BN or third party (BUF) heart allografts on day 100. RESULTS: While pretreatment with 200 mg/kg of CYP induced high levels of hematopoietic chimerism, six of eight recipients died of severe graft-versus-host-disease (GVHD). CYP at dose of 150 mg/kg induced 36.5 +/- 24.1% of donor-derived chimerism on day 10, and sustained macrochimerism was seen until day 100 without GVHD. Pretreatment with 100 mg/kg of CYP resulted in only transient chimerism (4.8 +/- 5.2%) which disappeared by day 20. In the recipients with 50 mg/kg of CYP, donor bone marrow cells were rapidly rejected and no chimerism was observed. The recipients with 150 mg/kg of CYP accepted BN heart allografts (>100 days x 5), while rejecting BUF allografts by day 12 (n = 4). BN heart allografts were rejected in the recipients with 100 (MST: 57 days, n = 5) and 50 mg/kg (MST: 7 days, n = 5) of CYP. CONCLUSIONS: A single dose of CYP can induce hematopoietic chimerism across MHC-barriers. The dose of 150 mg/kg seems to be optimal to induce organ transplant tolerance without developing GVHD.