RESUMEN
Granzyme B-producing B cells have been reportedly reported to be an important regulatory B cell subset in the pathogenesis of many diseases. However, its role in liver transplant recipients with acute rejection has never been well elucidated. Seventeen liver transplant recipients with acute rejection and 25 controls with stable liver function were enrolled in this study to determine the function of granzyme B-producing B cells via flow cytometry. Liver transplant recipients with acute rejection had higher expression of granzyme B in CD19+B cells when compared with controls. Following rejection, the granzyme B production was even elevated although comparison failed to reach significance. The percentages of CD27+granzyme B+CD19+B cells and CD138+granzyme B+CD19+B cells were lower than those of CD27-granzyme B+CD19+B cells and CD138-granzyme B+CD19+B cells in patients with acute rejection, respectively. While the production of CD27 and CD138 was not different between liver transplant recipients with and without acute rejection but increased expression of the two surface markers was observed following rejection. Furthermore, the frequency of granzyme B+CD19+B cells correlated with the level of alanine aminotransferase instead of tacrolimus. CD19+B cells could produce granzyme B when stimulated with IgG + IgM and CpG in the presence of the supernatant of activated CD4+CD25-T cells. In return, granzyme B+CD19+B cells could suppress the proliferation of CD4+CD25-T cells in a granzyme B- and contact-dependent manner. The increased expression of granzyme B in CD19+B cells from liver transplant recipients with acute rejection might function as a feedback loop for the activation of the CD4+CD25-T cells.
Asunto(s)
Linfocitos B/enzimología , Comunicación Celular , Rechazo de Injerto/enzimología , Granzimas/metabolismo , Trasplante de Hígado/efectos adversos , Activación de Linfocitos , Linfocitos T Colaboradores-Inductores/enzimología , Enfermedad Aguda , Adulto , Anciano , Antígenos CD19/metabolismo , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Estudios de Casos y Controles , Proliferación Celular , Células Cultivadas , Técnicas de Cocultivo , Femenino , Glucocorticoides/uso terapéutico , Rechazo de Injerto/diagnóstico , Rechazo de Injerto/tratamiento farmacológico , Rechazo de Injerto/inmunología , Humanos , Inmunosupresores/uso terapéutico , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Activación de Linfocitos/efectos de los fármacos , Masculino , Persona de Mediana Edad , Linfocitos T Colaboradores-Inductores/efectos de los fármacos , Linfocitos T Colaboradores-Inductores/inmunología , Regulación hacia ArribaRESUMEN
ABSTRACT: Acute immune rejection is one of the most serious complications of heart transplantation, and its mechanism has always been a hot spot. Th17 cells and cytokine interleukin-17 (IL-17) have been proved to be involved in acute immune rejection, and the signaling pathway mechanism has attracted our interest. It has been confirmed that the Janus kinase 2-signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway is involved in the differentiation of CD4+ T cells, so we focus on whether the JAK2/STAT3 signaling pathway is involved in the occurrence of acute immune rejection by regulating the Th17/IL-17 axis. In this study, we used Bagg's Albino c mice and C57BL/6 mice to construct heterotopic heart transplantation models, which were divided into the acute rejection group and AG490-treated group (n = 5), and donor tissue and serum were collected in 3 experimental days from the recipient mice for H&E staining analysis of paraffin sections and ELISA, Western blot, flow cytometry, and real time-polymerase chain reaction. The results showed that the acute rejection rating of the heart decreased, and the expression of related factors decreased significantly after using the inhibitor AG490, suggesting that the JAK2/STAT3 signaling pathway regulates expression of the Th17/IL-17 axis in cardiac allograft rejection.
Asunto(s)
Rechazo de Injerto/prevención & control , Trasplante de Corazón , Inmunosupresores/farmacología , Interleucina-17/metabolismo , Janus Quinasa 2/antagonistas & inhibidores , Inhibidores de las Cinasas Janus/farmacología , Miocardio/enzimología , Factor de Transcripción STAT3/metabolismo , Células Th17/metabolismo , Tirfostinos/farmacología , Animales , Modelos Animales de Enfermedad , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Supervivencia de Injerto/efectos de los fármacos , Janus Quinasa 2/metabolismo , Masculino , Ratones Endogámicos C57BL , Miocardio/inmunología , Miocardio/patología , Transducción de Señal , Células Th17/inmunología , Trasplante HeterotópicoRESUMEN
Chronic allograft dysfunction is a major cause of late graft failure after kidney transplantation. One of the histological changes is interstitial fibrosis, which is associated with epithelial-mesenchymal transition. Bortezomib has been reported to prevent the progression of fibrosis in organs. We used rat renal transplantation model and human kidney 2 cell line treated with tumor necrosis factor-α (TNF-α) to examine their response to bortezomib. To explore the mechanism behind it, we assessed the previously studied TNF-α/protein kinase B (Akt)/Smad ubiquitin regulatory factor 2 (Smurf2) signaling and performed RNA sequencing. Our results suggested that bortezomib could attenuate the TNF-α-induced epithelial-mesenchymal transition and renal allograft interstitial fibrosis in vitro and in vivo. In addition to blocking Akt/mammalian target of rapamycin (mTOR)/p70S6 kinase/Smurf2 signaling, bortezomib's effect on the epithelial-mesenchymal transition was associated with inhibition of nuclear factor kappa B (NF-κB) pathway by stabilizing inhibitor of NF-κB. The study highlighted the therapeutic potential of bortezomib on renal allograft interstitial fibrosis. Such an effect may result from inhibition of NF-κB/TNF-α/Akt/mTOR/p70S6 kinase/Smurf2 signaling via stabilizing protein of inhibitor of NF-κB.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Bortezomib/farmacología , Rechazo de Injerto/prevención & control , Enfermedades Renales/prevención & control , Trasplante de Riñón/efectos adversos , Túbulos Renales Proximales/efectos de los fármacos , Inhibidores de Proteasoma/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Línea Celular , Transición Epitelial-Mesenquimal/efectos de los fármacos , Fibrosis , Rechazo de Injerto/enzimología , Rechazo de Injerto/etiología , Rechazo de Injerto/patología , Supervivencia de Injerto/efectos de los fármacos , Humanos , Enfermedades Renales/enzimología , Enfermedades Renales/etiología , Enfermedades Renales/patología , Túbulos Renales Proximales/enzimología , Túbulos Renales Proximales/patología , Masculino , Inhibidor NF-kappaB alfa/metabolismo , FN-kappa B/metabolismo , Estabilidad Proteica , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas Endogámicas F344 , Ratas Endogámicas Lew , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Factores de Tiempo , Factor de Necrosis Tumoral alfa/metabolismo , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
OBJECTIVES: Antibody-mediated injury in chronic active antibody-mediated rejection, possibly with other effector T cells, may play a role in graft injury. The role of inflammatory cells in the inflammation and fibrosis and tubular atrophy region has been recently advocated in the progression of injury. Cytotoxic T cells play a prominent role in T-cell-mediated rejection; however, the possible role of cytotoxic T cells in circulation and the intragraft compartment in chronic active antibody-mediated rejection, a common immunological cause of long-term graft failure, has not been well-studied. MATERIALS AND METHODS: We measured the frequency of circulating cytotoxic T cells with flow cytometry, serum granzyme B level by enzyme-linked immunosorbent assay and intragraft granzyme B+ cell, and mRNA by immunohistochemistry and real-time polymerase chain reaction in biopsy tissue from living donor renal allograft recipients with stable graft function and chronic active antibody-mediated rejection. RESULTS: The frequency of CD3+ and CD3+CD8+ T cells was similar in both stable graft function patients and chronic active antibody-mediated rejection patients. The frequency of CD3+CD8+granzyme B+ cytotoxic T cells was significantly lower in peripheral blood. Serum granzyme B level and intragraft number of granzyme B+ cells (counts/mm²) were also significantly higher in the chronic active antibody-mediated rejection group compared with that of patients with stable graft function. The intragraft granzyme B+ T cell count was positively correlated with serum creatinine and 24-hour urine proteinuria but negatively correlated with estimated glomerular filtration rate. CONCLUSIONS: Granzyme B mediates covert graft injury in patients with chronic active antibody-mediated rejection in addition to antibody-mediated injury.
Asunto(s)
Citotoxicidad Celular Dependiente de Anticuerpos , Rechazo de Injerto/enzimología , Granzimas/sangre , Trasplante de Riñón/efectos adversos , Riñón/enzimología , Linfocitos T Citotóxicos/enzimología , Adulto , Biomarcadores/sangre , Enfermedad Crónica , Femenino , Rechazo de Injerto/sangre , Rechazo de Injerto/genética , Rechazo de Injerto/inmunología , Granzimas/genética , Humanos , Riñón/inmunología , Riñón/patología , Masculino , Persona de Mediana Edad , Fenotipo , Linfocitos T Citotóxicos/inmunología , Resultado del TratamientoRESUMEN
BACKGROUND: Allograft vasculopathy (AV) is the primary limiting factor for long-term graft survival. An increased activity of matrix metalloproteinases (MMPs) contributes to neointima formation in AV and represents a potential therapeutic target. Adeno-associated virus (AAV)-mediated gene therapy comprises a potentially benign vector model for the long-term expression of MMP antagonists. METHODS: Aortic allografts from DBA/2 mice were incubated with control buffer, AAV-enhanced green fluorescence protein (EGFP), or tissue inhibitor of metalloproteinases 1 (TIMP-1)-loaded AAV (AAV-TIMP-1) and transplanted into the infrarenal aorta of C57BL/6 mice. Cyclosporine A (10 mg/kg body weight) was administered daily. Explantation as well as histomorphometric and immunohistochemical evaluation was performed after 30 days. Matrix metalloproteinase (MMP) activity was visualized by gelatin in situ zymography. RESULTS: Intima-to-media area ratio and neointima formation were significantly reduced in the AAV-TIMP-1 treatment group compared with those in the control group (by 40%; p < 0.001) and the AAV-EGFP group (by 38.2%; p < 0.001). TIMP-1 overexpression positively affected several pathomechanisms for the development of AV both in vitro and in vivo as compared to that in the control groups: endothelium integrity was preserved as shown by zona occludens 1 and occludin staining; MMP9 expression and activity were significantly reduced (pâ¯=â¯0.01); and smooth muscle cell migration was significantly reduced as smooth muscle actin positive cells predominantly remained in the aortic media in the treatment group (pâ¯=â¯0.001). Moreover, macrophage infiltration was markedly reduced by 49% in the AAV-TIMP-1 group (p < 0.001). CONCLUSION: Immediate post-harvesting allograft incubation with AAV-TIMP-1 reduces neointima formation and macrophage infiltration, constituting a possible adjunct therapeutic strategy to preserve graft function after transplantation.
Asunto(s)
Aorta Torácica/trasplante , Dependovirus/enzimología , Regulación de la Expresión Génica , Rechazo de Injerto/genética , Inhibidor Tisular de Metaloproteinasa-1/genética , Túnica Íntima/metabolismo , Aloinjertos , Animales , Aorta Torácica/metabolismo , Aorta Torácica/patología , Western Blotting , Células Cultivadas , Modelos Animales de Enfermedad , Rechazo de Injerto/enzimología , Rechazo de Injerto/patología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , ARN/genética , Inhibidor Tisular de Metaloproteinasa-1/biosíntesis , Túnica Íntima/patologíaRESUMEN
The identification of risk factors for acute rejection (AR) may lead to strategies to improve success of kidney transplantation. Ectonucleotidases are ectoenzymes that hydrolyze extracellular nucleotides into nucleosides, modulating the purinergic signaling. Some members of the Ectonucleotidase family have been linked to transplant rejection processes. However, the association of Ectonucleotide Pyrophosphatase / Phosphodiesterase 1 (ENPP1) with AR has not yet been evaluated. The aim of this study was to evaluate the association between the K121Q polymorphism of ENPP1 gene and AR in kidney transplant patients. We analyzed 449 subjects without AR and 98 with AR from a retrospective cohort of kidney transplant patients from Southern Brazil. K121Q polymorphism was genotyped using allelic discrimination-real-time PCR. Cox regression analysis was used to evaluate freedom of AR in kidney transplant patients according to genotypes. Q allele frequency was 17.6% in recipients without AR and 21.9% in those with AR (P = 0.209). Genotype frequencies of the K121Q polymorphism were in Hardy-Weinberg equilibrium in non-AR patients (P = 0.70). The Q/Q genotype (recessive model) was associated with AR (HR = 2.83, 95% CI 1.08-7.45; P = 0.034) after adjusting for confounders factors. Our findings suggest a novel association between the ENPP1 121Q/Q genotype and AR in kidney transplant recipients.
Asunto(s)
Rechazo de Injerto/enzimología , Rechazo de Injerto/genética , Trasplante de Riñón/efectos adversos , Hidrolasas Diéster Fosfóricas/genética , Pirofosfatasas/genética , Enfermedad Aguda , Sustitución de Aminoácidos , Brasil , Estudios de Casos y Controles , Estudios de Cohortes , Frecuencia de los Genes , Genes Recesivos , Estudios de Asociación Genética , Humanos , Modelos Genéticos , Polimorfismo de Nucleótido Simple , Estudios RetrospectivosRESUMEN
The immunoproteasome is expressed in cells of hematopoietic origin and is induced during inflammation by IFN-γ. Targeting the immunoproteasome with selective inhibitors has been shown to be therapeutically effective in pre-clinical models for autoimmune diseases, colitis-associated cancer formation, and transplantation. Immunoproteasome inhibition prevents activation and proliferation of lymphocytes, lowers MHC class I cell surface expression, reduces the expression of cytokines of activated immune cells, and curtails T helper 1 and 17 cell differentiation. This might explain the in vivo efficacy of immunoproteasome inhibition in different pre-clinical disease models for autoimmunity, cancer, and transplantation. In this review, we summarize the effect of immunoproteasome inhibition in different animal models for transplantation.
Asunto(s)
Enfermedades Autoinmunes/terapia , Rechazo de Injerto/etiología , Antígenos de Histocompatibilidad Clase I/inmunología , Trasplante de Órganos/efectos adversos , Complejo de la Endopetidasa Proteasomal/inmunología , Inhibidores de Proteasoma/uso terapéutico , Animales , Rechazo de Injerto/tratamiento farmacológico , Rechazo de Injerto/enzimología , Antígenos de Histocompatibilidad Clase I/efectos de los fármacos , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/metabolismo , Transducción de SeñalRESUMEN
OBJECTIVES: The authors sought to evaluate cardiac activity of angiotensin-converting enzyme (ACE) and ACE2 after heart transplantation (HT) and its relation with acute rejection (AR) and chronic allograft vasculopathy (CAV). BACKGROUND: The renin-angiotensin system is altered in heart failure and HT. However, ACE and ACE2 activities in post-HT acute and chronic rejection have not been previously studied. METHODS: HT patients (nâ¯=â¯45) were included when appropriate serial endomyocardial biopsies (EMB) and coronary angiography were available for analysis. In 21 patients, three post-HT time points were selected for CAV study in EMB tissue: basal (0-3â¯wks), second (2-3â¯months) and third (4-5â¯months). At 10â¯years post-HT, CAV was evaluated by coronary angiography (CA) and patients were grouped by degree of CAV: 0-1, non-CAV (nâ¯=â¯15) and 2-3, CAV (nâ¯=â¯6). For the AR study, 28 HT patients with evidence of one EMB rejection at grade 3 and two EMB grade 1A and/or 1B rejections were selected. RESULTS: Post-HT, ACE2 activity was increased in the CAV group, compared to non-CAV. Patients with AR showed increased ACE, but not ACE2, activity. CONCLUSIONS: Our results suggest that early post-HT cardiac ACE2 activity may have an important role in CAV development. In contrast, ACE activity was increased in AR. The renin-angiotensin system seems to be altered after HT and strategies to balance the system may be useful.
Asunto(s)
Rechazo de Injerto/enzimología , Trasplante de Corazón/efectos adversos , Miocardio/enzimología , Peptidil-Dipeptidasa A/metabolismo , Enfermedad Aguda , Adulto , Enzima Convertidora de Angiotensina 2 , Biomarcadores/metabolismo , Biopsia , Enfermedad Crónica , Angiografía Coronaria , Femenino , Estudios de Seguimiento , Rechazo de Injerto/diagnóstico , Insuficiencia Cardíaca/cirugía , Humanos , Masculino , Persona de Mediana Edad , Miocardio/patología , Proyectos Piloto , Pronóstico , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Factores de Tiempo , Trasplante HomólogoRESUMEN
Early damage to transplanted organs initiates excess inflammation that can cause ongoing injury, a leading cause for late graft loss. The endothelial glycocalyx modulates immune reactions and chemokine-mediated haptotaxis, potentially driving graft loss. In prior work, conditional deficiency of the glycocalyx-modifying enzyme N-deacetylase-N-sulfotransferase-1 (Ndst1f/f TekCre+) reduced aortic allograft inflammation. Here we investigated modification of heparan sulfate (HS) and chemokine interactions in whole-organ renal allografts. Conditional donor allograft Ndst1 deficiency (Ndst1-/-; C57Bl/6 background) was compared to systemic treatment with M-T7, a broad-spectrum chemokine-glycosaminoglycan (GAG) inhibitor. Early rejection was significantly reduced in Ndst1-/- kidneys engrafted into wildtype BALB/c mice (Ndst1+/+) and comparable to M-T7 treatment in C57Bl/6 allografts (P < 0.0081). M-T7 lost activity in Ndst1-/- allografts, while M-T7 point mutants with modified GAG-chemokine binding displayed a range of anti-rejection activity. CD3+ T cells (P < 0.0001), HS (P < 0.005) and CXC chemokine staining (P < 0.012), gene expression in NFκB and JAK/STAT pathways, and HS and CS disaccharide content were significantly altered with reduced rejection. Transplant of donor allografts with conditional Ndst1 deficiency exhibit significantly reduced acute rejection, comparable to systemic chemokine-GAG inhibition. Modified disaccharides in engrafted organs correlate with reduced rejection. Altered disaccharides in engrafted organs provide markers for rejection with potential to guide new therapeutic approaches in allograft rejection.
Asunto(s)
Células Alogénicas/enzimología , Aorta/trasplante , Células Progenitoras Endoteliales/enzimología , Rechazo de Injerto/enzimología , Células Progenitoras Mieloides/enzimología , Sulfotransferasas , Células Alogénicas/patología , Animales , Aorta/patología , Células Progenitoras Endoteliales/patología , Eliminación de Gen , Rechazo de Injerto/genética , Rechazo de Injerto/patología , Rechazo de Injerto/prevención & control , Ratones , Ratones Endogámicos BALB C , Células Progenitoras Mieloides/patología , Sulfotransferasas/genética , Sulfotransferasas/metabolismoRESUMEN
BACKGROUND: Hepatitis E virus (HEV) infection is a potential reason for elevated liver enzymes after liver transplantation (LT). Our aim was to analyze a real-world cohort of LT patients, who underwent liver biopsy for elevated transaminases and suspected acute rejection, to evaluate frequency of post-transplant HEV infection. PATIENTS: Data from 160 liver biopsies were analyzed. Seventy-one patients were biopsied on schedule after LT without elevated liver enzymes. A subgroup of 25 patients with elevated liver enzymes and suspected rejection was chosen for further analysis. Patient demographics and data were retrieved from a clinical database, patients' charts, and reports. RESULTS: Hepatitis E virus infection was diagnosed in five of 25 patients with suspected acute rejection (20%). HEV genotype 3 was detected in three of the five HEV-infected patients. Patients with HEV infection showed higher ALT levels (P = 0.014), lower De Ritis ratio (P = 0.021), and more frequent glucocorticoid therapy (P = 0.012) compared to HEV-negative patients. CONCLUSION: We found a rate of 20% HEV infections in LT patients undergoing liver biopsy for elevated liver enzymes and suspected acute rejection. These data indicate the necessity for HEV testing in all LT patients with elevated liver enzymes and suspected acute rejection.
Asunto(s)
Biomarcadores/sangre , Rechazo de Injerto/diagnóstico , Virus de la Hepatitis E/genética , Hepatitis E/diagnóstico , Hepatopatías/cirugía , Trasplante de Hígado/efectos adversos , Complicaciones Posoperatorias , Adulto , Anciano , Alanina Transaminasa/sangre , Aspartato Aminotransferasas/sangre , Biopsia , Femenino , Estudios de Seguimiento , Genotipo , Rechazo de Injerto/sangre , Rechazo de Injerto/enzimología , Rechazo de Injerto/etiología , Supervivencia de Injerto , Hepatitis E/sangre , Hepatitis E/enzimología , Hepatitis E/etiología , Virus de la Hepatitis E/aislamiento & purificación , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Factores de RiesgoRESUMEN
In kidney transplantation acute allograft rejection is the most common cause of late allograft loss. Changes in indoleamine 2,3 dioxygenase (IDO) activity, which catabolizes the degradation of tryptophan to kynurenine, may predict rejection. However, exogenous IDO is immunosuppressive in rodent kidney transplantation. Thus, the increase in IDO activity observed in acute allograft rejection is insufficient to prevent rejection. To address this question, we assessed the regulation of IDO and its role in acute rejection in a porcine model of kidney transplant. In tissue samples from rejecting kidney allografts, we showed a 13-fold increase in IDO gene transcription and 20-fold increase in IDO enzyme activity when compared with autotransplanted kidneys. Allografts also demonstrated an over fourfold increase in tissue interferon (IFN)-γ, with marked increases in tumor necrosis factor (TNF)-α, TNF-ß and interleukin 1ß. Gene transcription and protein levels of kynurenine 3-monooxygenase (KMO) were decreased. KMO generates the immunosuppressive kynurenine, 3-hydroxykynurenine. The results of these studies demonstrate a clear association between rejection and increased allograft IDO expression, likely driven in part by IFN-γ and facilitated by other cytokines of the allogeneic response. Moreover, the loss of downstream enzymatic activity in the IDO metabolic pathway may suggest novel mechanisms for the perpetuation of rejection.
Asunto(s)
Aloinjertos/trasplante , Rechazo de Injerto/enzimología , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Trasplante de Riñón/efectos adversos , Animales , Creatinina/sangre , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Regulación Enzimológica de la Expresión Génica , Rechazo de Injerto/sangre , Indolamina-Pirrol 2,3,-Dioxigenasa/genética , Riñón/patología , Quinurenina/metabolismo , Porcinos , Transcripción Genética , Trasplante HomólogoRESUMEN
OBJECTIVES: We aim to identify the key biomarker of acute rejection (AR) after kidney transplantation via bioinformatics methods. METHODS: The gene expression data GSE75693 of 30 samples with stable kidney transplantation recipients and 15 AR samples were downloaded and analyzed by the limma package to identify differentially expressed genes (DEGs). Then, Gene Ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were done to explore the biological functions and potential important pathways of DEGs. Finally, protein-protein interactions (PPIs) and literature mining were applied to construct the cocitation network and to select the hub protein. RESULTS: A total of 437 upregulated genes and 353 downregulated genes were selected according to P < 0.01 and |log2(fold change)| > 1.0. DEGs of AR are mainly located on membranes and impact the activation of receptors in immune responses. In the PPI network, Src kinase, lymphocyte kinase (LCK), CD3G, B2M, interferon-γ, CD3D, tumor necrosis factor, VAV1, and CD3E in the T cell receptor signaling pathway were selected as important factors, and LCK was identified as the hub protein. CONCLUSION: LCK, via acting on T-cell receptor, might be a potential therapeutic target for AR after kidney transplantation.
Asunto(s)
Rechazo de Injerto/metabolismo , Trasplante de Riñón , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/metabolismo , Análisis por Conglomerados , Biología Computacional , Minería de Datos , Perfilación de la Expresión Génica , Ontología de Genes , Redes Reguladoras de Genes , Rechazo de Injerto/enzimología , Rechazo de Injerto/genética , Rechazo de Injerto/terapia , Humanos , Riñón/enzimología , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/antagonistas & inhibidores , Linfocitos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Mapeo de Interacción de Proteínas , Mapas de Interacción de Proteínas , Transducción de SeñalRESUMEN
OBJECTIVE: The objective of this study was to investigate the effect of nucleoside triphosphate diphosphohydrolase-1 (NTPDase1) during acute antibody-mediated rejection (AMR). METHODS: NTPDase1 overexpression, NTPDase1 knockout, and wild-type nude mice skin graft models were used to induce acute AMR. NTPDase1 expression in B cells, NTPDase1 messenger RNA expression in skin grafts, extracellular adenosine diphosphate (ADP) concentration, B-cell volume and surface antigens expression, average platelet transport rate, and ultrastructure and apoptosis of skin graft cells were investigated. RESULTS: During acute AMR in nude mice, higher NTPDase1 expression caused lower extracellular ADP concentration, smaller increase in B-cell volume, and major histocompatibility complex II surface antigen expression, suggesting a negative correlation between them; higher NTPDase1 expression also caused slower average platelet transport rate and less severe skin graft injury, suggesting a negative correlation between them. Pretreatment with high-dose exogenous NTPDase1 inhibited platelet activation and protected skin grafts, but it resulted in prolonged bleeding time (by 51.4%) and prolonged coagulation time (by 44.1%). CONCLUSION: An NTPDase1-associated imbalance in extracellular ADP degradation may contribute to B-cell activation, platelet activation, and more severe skin graft injury in nude mice. Pretreatment with high-dose exogenous NTPDase1 effectively protected skin grafts in nude mice at 1 week, but it increased the risk of bleeding.
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Adenosina Difosfato/metabolismo , Adenosina Trifosfato/fisiología , Antígenos CD/metabolismo , Apirasa/metabolismo , Linfocitos B/fisiología , Rechazo de Injerto/enzimología , Activación Plaquetaria/fisiología , Trasplante de Piel , Animales , Antígenos CD/genética , Antígenos CD/farmacología , Apirasa/genética , Apirasa/farmacología , Tamaño de la Célula , Rechazo de Injerto/inmunología , Activación de Linfocitos/fisiología , Masculino , Ratones , Ratones Desnudos , ARN Mensajero/metabolismoRESUMEN
Background: Although immunosuppressive therapies have made organ transplantation a common medical procedure worldwide, chronic toxicity has a major issue for long-term treatment. One method to improve therapies and methods is the application of immunomodulatory agents from parasites such as Hypoderma lineatum. Hypodermin A (HA) is a serine esterase secreted by the larvae of Hypoderma lineatum, several studies demonstrated its immunosuppressive mechanism in vitro, and recently we discovered that HA inhibits the expression of interferon (IFN)-γ and interleukin (IL)-2 and activates IL-10 expression. Therefore, we hypothesized that it might be a potential agent used to block allograft rejections. However, most studies of the immunosuppressive mechanisms associated with HA were undertaken at the cellular level. In order to augment these studies, we evaluated the immunosuppressive effects of HA in vivo using an HA transgenic mouse model. Result: Our results revealed similar findings to those reported by in vitro studies, specifically that HA induced prostaglandin E2 expression, downregulated IFN-γ and IL-2 expression, and promoted IL-10 secretion via E-type prostanoid receptor 4. Additionally, we observed that HA overexpression inhibited lipopolysaccharide-induced TLR4 activation. These findings provide insight into a new potential agent capable of blocking graft rejection. Conclusion: Our founding suggested that HA-related treatment could be a promising option to improve the viability of grafts in human.
Asunto(s)
Animales , Ratones , Serina Endopeptidasas/inmunología , Dípteros/enzimología , Dípteros/inmunología , Rechazo de Injerto/inmunología , Ensayo de Inmunoadsorción Enzimática , Serina Endopeptidasas/metabolismo , Western Blotting , Citocinas , Terapia de Inmunosupresión , Interleucinas/antagonistas & inhibidores , Interferones/antagonistas & inhibidores , Interleucina-10/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Receptor Toll-Like 4 , Reacción en Cadena en Tiempo Real de la Polimerasa , Rechazo de Injerto/enzimología , Rechazo de Injerto/prevención & controlRESUMEN
Regulatory T cells were recently proposed as the central actor in operational tolerance after renal transplantation. Tolerant patients harbor increased FoxP3hi memory Treg frequency and increased demethylation in the Foxp3 Treg-specific demethylated region when compared to stable kidney recipients and exhibit greater memory Treg suppressive capacities and higher expression of the ectonucleotidase CD39. However, in this particular and unique situation the mechanisms of action of Tregs were not identified. Thus, we analyzed the ability of memory Tregs to degrade extracellular ATP in tolerant patients, healthy volunteers, and patients with stable graft function under immunosuppression and determined the role of immunosuppressive drugs on this process. The conserved proportion of memory Tregs leads to the establishment of a pro-tolerogenic balance in operationally tolerant patients. Memory Tregs in tolerant patients display normal capacity to degrade extracellular ATP/ADP. In contrast, memory Tregs from patients with stable graft function do not have this ability. Finally, in vitro, immunosuppressive drugs may favor the lower proportion of memory Tregs in stable patients, but they have no effect on CD39-dependent ATP degradation and do not explain memory Treg lack of extracellular ATP/ADP degradation ability. Thus, intrinsic active regulatory mechanisms may act long after immunosuppressive drug arrest in operationally tolerant patients and may contribute to kidney allograft tolerance via the maintenance of CD39 Treg function.
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Adenosina Trifosfato/metabolismo , Apirasa/metabolismo , Metabolismo Energético , Rechazo de Injerto/prevención & control , Supervivencia de Injerto , Memoria Inmunológica , Trasplante de Riñón , Linfocitos T Reguladores/enzimología , Tolerancia al Trasplante , Adenosina Difosfato/metabolismo , Adulto , Anciano , Estudios de Casos y Controles , Células Cultivadas , Metabolismo Energético/efectos de los fármacos , Femenino , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Supervivencia de Injerto/efectos de los fármacos , Humanos , Hidrólisis , Memoria Inmunológica/efectos de los fármacos , Inmunosupresores/uso terapéutico , Trasplante de Riñón/efectos adversos , Masculino , Persona de Mediana Edad , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Tolerancia al Trasplante/efectos de los fármacos , Adulto JovenAsunto(s)
Caspasa 8/inmunología , Inhibidores de Caspasas/farmacología , Trasplante de Córnea , Rechazo de Injerto/enzimología , Linfangiogénesis/efectos de los fármacos , Animales , Rechazo de Injerto/inmunología , Inflamación/enzimología , Inflamación/inmunología , Linfangiogénesis/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacos , Transducción de Señal/inmunologíaRESUMEN
Chronic antibody-mediated rejection is the major cause of fading allograft function and loss after renal transplantation. Currently, pharmacological agents for the suppression of chronic antibody-mediated rejection are lacking. Non-selective proteasome inhibitors suppress antibody-mediated allograft rejection. However, extensive adverse side effects of these inhibitors severely limit their application. In contrast, immunoproteasome inhibition is effective in preclinical models of autoimmune diseases and was applied over weeks without obvious adverse side effects. ONX 0914, an immunoproteasome subunit LMP7 (ß5i)-selective inhibitor, impeded the chronic rejection of kidneys transplanted from Fischer to allogeneic Lewis rats. ONX 0914 inhibited immunoproteasome induction both in immune organs and renal allografts. Selective immunoproteasome inhibition reduced the numbers of B and plasma cells, and suppressed donor-specific alloantibody production. The infiltration of T cells, B cells and macrophages as well as interferon-γ, interleukin-17, IgG and complement deposition were reduced in renal allografts of ONX 0914-treated recipients. Chronic nephropathy was ameliorated and renal allograft function preserved, enabling long-term survival of recipients. Thus, our studies define a critical role of the immunoproteasome in chronic kidney allograft rejection and suggest immunoproteasome inhibition as a promising therapeutic approach to suppress chronic antibody-mediated rejection.
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Rechazo de Injerto/prevención & control , Supervivencia de Injerto/efectos de los fármacos , Inmunosupresores/farmacología , Isoanticuerpos/inmunología , Trasplante de Riñón/efectos adversos , Riñón/efectos de los fármacos , Oligopéptidos/farmacología , Complejo de la Endopetidasa Proteasomal/efectos de los fármacos , Inhibidores de Proteasoma/farmacología , Aloinjertos , Animales , Enfermedad Crónica , Modelos Animales de Enfermedad , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Riñón/enzimología , Riñón/inmunología , Riñón/patología , Masculino , Complejo de la Endopetidasa Proteasomal/inmunología , Complejo de la Endopetidasa Proteasomal/metabolismo , Ratas Endogámicas F344 , Ratas Endogámicas Lew , Factores de TiempoRESUMEN
AIM: To investigate whether bone marrow mesenchymal stem cells (BMMSCs) modified with the HO-1 and CXCR3 genes can augment the inhibitory effect of BMMSCs on small bowel transplant rejection. METHODS: Lewis rat BMMSCs were cultured in vitro. Third-passage BMMSCs were transduced with the CXCR3/HO-1 genes or the HO-1 gene alone. The rats were divided into six groups and rats in the experimental group were pretreated with BMMSCs 7 d prior to small bowel transplant. Six time points (instant, 1 d, 3 d, 7 d, 10 d, and 14 d) (n = 6) were chosen for each group. Hematoxylin-eosin staining was used to observe pathologic rejection, while immunohistochemistry and Western blot were used to detect protein expression. Flow cytometry was used to detect T lymphocytes and enzyme linked immunosorbent assay was used to detect cytokines. RESULTS: The median survival time of BMMSCs from the CXCR3/HO-1 modified group (53 d) was significantly longer than that of the HO-1 modified BMMSCs group (39 d), the BMMSCs group (26 d), and the NS group (control group) (16 d) (P < 0.05). Compared with BMMSCs from the HO-1 modified BMMSCs, BMMSCs, and NS groups, rejection of the small bowel in the CXCR3/HO-1 modified group was significantly reduced, while the weight of transplant recipients was also significantly decreased (P < 0.05). Furthermore, IL-2, IL-6, IL-17, IFN-γ, and TNF-α levels were significantly decreased and the levels of IL-10 and TGF-ß were significantly increased (P < 0.05). CONCLUSION: BMMSCs modified with the CXCR3 and HO-1 genes can abrogate the rejection of transplanted small bowel more effectively and significantly increase the survival time of rats that receive a small bowel transplant.
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Rechazo de Injerto/prevención & control , Supervivencia de Injerto , Hemo-Oxigenasa 1/metabolismo , Intestino Delgado/trasplante , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/enzimología , Receptores CXCR3/metabolismo , Animales , Apoptosis , Supervivencia Celular , Células Cultivadas , Citocinas/sangre , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Hemo-Oxigenasa 1/genética , Intestino Delgado/enzimología , Intestino Delgado/inmunología , Intestino Delgado/patología , Masculino , Células Madre Mesenquimatosas/inmunología , Fenotipo , Ratas Endogámicas BN , Ratas Endogámicas Lew , Receptores CXCR3/genética , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Factores de Tiempo , TransfecciónRESUMEN
BACKGROUND: Organ transplantation into sensitized patients with preexisting donor-specific antibodies (DSA) is very challenging. Spleen tyrosine kinase (Syk) promotes leukocyte recruitment and activation via signaling through various cell surface receptors. We investigated whether a selective Syk inhibitor (GS-492429) could suppress antibody-mediated rejection (AMR) in a rat model of AMR in sensitized recipients. METHODS: Recipient Lewis rats (RT1) were immunized with donor (Dark Agouti, RT1) spleen cells (day -5). Recipients underwent bilateral nephrectomy and orthotopic renal transplantation (day 0). Cellular rejection was minimized by tacrolimus treatment from day -1. Groups received GS-492429 (30 mg/kg, twice a day) (n = 11) or vehicle (n = 12) from 1 hour before transplantation until being killed on day 3. RESULTS: Vehicle-treated recipients developed graft dysfunction on day 1 which rapidly worsened by day 3. Histology showed severe damage (thrombosis, acute tubular injury, capillaritis) and infiltration of many Syk leukocytes. GS-492429 did not affect graft dysfunction on day 1, but treatment reduced allograft damage and prevented the rapid deterioration of graft function on day 3. GS-492429 reduced the prominent macrophage infiltrate and reduced the M1 proinflammatory response. Neutrophil and NK cell infiltration and capillary thrombosis were also significantly reduced by GS-492429 treatment. Serum DSA levels and the deposition of IgG and C4d in the allograft were equivalent in the 2 groups. CONCLUSIONS: Treatment with a Syk inhibitor significantly reduced renal allograft injury in a model of severe antibody-mediated damage in highly sensitized recipients. Further studies are warranted to determine whether Syk inhibition is a potential adjunctive treatment in clinical AMR.
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Anticuerpos/inmunología , Rechazo de Injerto/prevención & control , Trasplante de Riñón/efectos adversos , Inhibidores de Proteínas Quinasas/farmacología , Quinasa Syk/antagonistas & inhibidores , Receptores de Trasplantes , Enfermedad Aguda , Animales , Western Blotting , Modelos Animales de Enfermedad , Rechazo de Injerto/enzimología , Rechazo de Injerto/inmunología , Ratas , Ratas Endogámicas Lew , Transducción de Señal , Quinasa Syk/metabolismo , Trasplante HomólogoRESUMEN
Immunosuppression after liver transplantation (LT) is presently based on use of calcineurin inhibitors (CNI), although they are associated with an increased incidence of renal dysfunction, cardiovascular complications, and de novo and recurrent malignancies. Over the past decade, mammalian target of rapamycin inhibitors have received considerable attention as immunosuppressants because they are associated with a more favorable renal profile versus CNI, as well as antiproliferative activity in clinical studies. Comprehensive guidelines on use of everolimus (EVR) in LT are still lacking. In Italy, a project, named Everolimus: the road to long-term functioning, was initiated to collect the experience on EVR after LT with the aim of providing guidance for transplant clinicians. Herein, recommendations by this national consensus group, based on Delphi methodology, are presented. Consensus was reached on 20 of the 23 statements proposed, and their level of evidence, grade of recommendation, and percent of agreement are reported. Statements are grouped into 4 areas: (A) renal function; (B) time of EVR introduction, CNI reduction and elimination, and risk for graft rejection; (C) antiproliferative effects of EVR; and (D) management of EVR-related adverse events. The high level of consensus shows that there is good agreement on the routine use of EVR in predefined clinical scenarios, especially in light of posttransplant nephrotoxicity and other adverse events associated with long-term administration of CNIs.