RESUMO
Multiple species within the order Hemiptera cause severe agricultural losses on a global scale. Aphids and whiteflies are of particular importance due to their role as vectors for hundreds of plant viruses, many of which enter the insect via the gut. To facilitate the identification of novel targets for disruption of plant virus transmission, we compared the relative abundance and composition of the gut plasma membrane proteomes of adult Bemisia tabaci (Hemiptera: Aleyrodidae) and Myzus persicae (Hemiptera: Aphididae), representing the first study comparing the gut plasma membrane proteomes of two different insect species. Brush border membrane vesicles were prepared from dissected guts, and proteins extracted, identified and quantified from triplicate samples via timsTOF mass spectrometry. A total of 1699 B. tabaci and 1175 M. persicae proteins were identified. Following bioinformatics analysis and manual curation, 151 B. tabaci and 115 M. persicae proteins were predicted to localize to the plasma membrane of the gut microvilli. These proteins were further categorized based on molecular function and biological process according to Gene Ontology terms. The most abundant gut plasma membrane proteins were identified. The ten plasma membrane proteins that differed in abundance between the two insect species were associated with the terms "protein binding" and "viral processes." In addition to providing insight into the gut physiology of hemipteran insects, these gut plasma membrane proteomes provide context for appropriate identification of plant virus receptors based on a combination of bioinformatic prediction and protein localization on the surface of the insect gut.
Assuntos
Afídeos , Trato Gastrointestinal , Proteínas de Insetos , Insetos Vetores , Vírus de Plantas , Animais , Proteínas de Insetos/metabolismo , Insetos Vetores/virologia , Insetos Vetores/metabolismo , Afídeos/virologia , Afídeos/metabolismo , Trato Gastrointestinal/virologia , Trato Gastrointestinal/metabolismo , Proteínas de Membrana/metabolismo , Hemípteros/virologia , Hemípteros/metabolismo , Proteoma , Membrana Celular/metabolismoRESUMO
Early immunological biomarkers that predict rejection and chronic allograft loss are needed to inform preemptive therapy and improve long-term outcomes. Here, we prospectively examined the ratio of interleukin-10 (IL-10) to tumor necrosis factor-α (TNFα) produced by transitional-1 B cells (T1B) 3 months after transplantation as a predictive biomarker for clinical and subclinical renal allograft rejection and subsequent clinical course. In both Training (n = 162) and Internal Validation (n = 82) Sets, the T1B IL-10/TNFα ratio 3 months after transplantation predicted both clinical and subclinical rejection anytime in the first year. The biomarker also predicted subsequent late rejection with a lead time averaging 8 months. Among biomarker high-risk patients, 60% had early rejection, of which 48% recurred later in the first posttransplant year. Among high-risk patients without early rejection, 74% developed rejection later in the first year. In contrast, only 5% of low-risk patients had early and 5% late rejection. The biomarker also predicted rejection in an External Validation Set (n = 95) and in key patient subgroups, confirming generalizability. Biomarker high-risk patients exhibited progressively worse renal function and decreased 5-year graft survival compared to low-risk patients. Treatment of B cells with anti-TNFα in vitro augmented the IL-10/TNFα ratio, restored regulatory activity, and inhibited plasmablast differentiation. To conclude, the T1B IL-10/TNFα ratio was validated as a strong predictive biomarker of renal allograft outcomes and provides a rationale for preemptive therapeutic intervention with TNF blockade.
Assuntos
Rejeição de Enxerto , Transplante de Rim , Aloenxertos , Citocinas , Humanos , Rim/fisiologia , Células Precursoras de Linfócitos BRESUMO
BACKGROUND: Transplantation is the treatment of choice for many patients with end-stage organ disease. Despite advances in immunosuppression, long-term outcomes remain suboptimal, hampered by drug toxicity and immune-mediated injury, the leading cause of late graft loss. The development of therapies that promote regulation while suppressing effector immunity is imperative to improve graft survival and minimize conventional immunosuppression. Notch signaling is a highly conserved pathway pivotal to T-cell differentiation and function, rendering it a target of interest in efforts to manipulate T cell-mediated immunity. METHODS: We investigated the pattern of Notch-1 expression in effector and regulatory T cells (Tregs) in both murine and human recipients of a solid-organ transplant. Using a selective human anti-Notch-1 antibody (aNotch-1), we examined the effect of Notch-1 receptor inhibition in full major histocompatibility complex-mismatch murine cardiac and lung transplant models, and in a humanized skin transplant model. On the basis of our findings, we further used a genetic approach to investigate the effect of selective Notch-1 inhibition in Tregs. RESULTS: We observed an increased proportion of Tregs expressing surface and intracellular (activated) Notch-1 in comparison with conventional T cells, both in mice with transplants and in the peripheral blood of patients with transplants. In the murine cardiac transplant model, peritransplant administration of aNotch-1 (days 0, 2, 4, 6, 8, and 10) significantly prolonged allograft survival in comparison with immunoglobulin G-treated controls. Similarly, aNotch-1 treatment improved both histological and functional outcomes in the murine lung transplant model. The use of aNotch-1 resulted in a reduced proportion of both splenic and intragraft conventional T cells, while increasing the proportion of Tregs. Furthermore, Tregs isolated from aNotch-1-treated mice showed enhanced suppressive function on a per-cell basis, confirmed with selective Notch-1 deletion in Tregs (Foxp3EGFPCreNotch1fl/fl). Notch-1 blockade inhibited the mammalian target of rapamycin pathway and increased the phosphorylation of STAT5 (signal transducer and activator of transcription 5) in murine Tregs. Notch-1low Tregs isolated from human peripheral blood exhibited more potent suppressive capacity than Notch-1high Tregs. Last, the combination of aNotch-1 with costimulation blockade induced long-term tolerance in a cardiac transplant model, and this tolerance was dependent on CTLA-4 (cytotoxic T-lymphocyte-associated antigen-4) signaling. CONCLUSIONS: Our data reveal a promising, clinically relevant approach for immune modulation in transplantation by selectively targeting Notch-1.
Assuntos
Rejeição de Enxerto/metabolismo , Receptor Notch1/antagonistas & inibidores , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Animais , Anticorpos Bloqueadores/farmacologia , Células Cultivadas , Regulação da Expressão Gênica , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/mortalidade , Humanos , Tolerância Imunológica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transplante de Órgãos , Receptor Notch1/genética , Receptor Notch1/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Análise de SobrevidaRESUMO
BACKGROUND: In kidney transplant recipients, surveillance biopsies can reveal, despite stable graft function, histologic features of acute rejection and borderline changes that are associated with undesirable graft outcomes. Noninvasive biomarkers of subclinical acute rejection are needed to avoid the risks and costs associated with repeated biopsies. METHODS: We examined subclinical histologic and functional changes in kidney transplant recipients from the prospective Genomics of Chronic Allograft Rejection (GoCAR) study who underwent surveillance biopsies over 2 years, identifying those with subclinical or borderline acute cellular rejection (ACR) at 3 months (ACR-3) post-transplant. We performed RNA sequencing on whole blood collected from 88 individuals at the time of 3-month surveillance biopsy to identify transcripts associated with ACR-3, developed a novel sequencing-based targeted expression assay, and validated this gene signature in an independent cohort. RESULTS: Study participants with ACR-3 had significantly higher risk than those without ACR-3 of subsequent clinical acute rejection at 12 and 24 months, faster decline in graft function, and decreased graft survival in adjusted Cox analysis. We identified a 17-gene signature in peripheral blood that accurately diagnosed ACR-3, and validated it using microarray expression profiles of blood samples from 65 transplant recipients in the GoCAR cohort and three public microarray datasets. In an independent cohort of 110 transplant recipients, tests of the targeted expression assay on the basis of the 17-gene set showed that it identified individuals at higher risk of ongoing acute rejection and future graft loss. CONCLUSIONS: Our targeted expression assay enabled noninvasive diagnosis of subclinical acute rejection and inflammation in the graft and may represent a useful tool to risk-stratify kidney transplant recipients.
Assuntos
Perfilação da Expressão Gênica , Rejeição de Enxerto/sangue , Rejeição de Enxerto/diagnóstico , Falência Renal Crônica/cirurgia , Transplante de Rim/efeitos adversos , Adulto , Idoso , Biomarcadores/metabolismo , Biópsia , Feminino , Genômica , Sobrevivência de Enxerto , Humanos , Imunossupressores/uso terapêutico , Inflamação , Estimativa de Kaplan-Meier , Falência Renal Crônica/sangue , Falência Renal Crônica/mortalidade , Transplante de Rim/mortalidade , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Estudos Prospectivos , Fatores de Risco , Análise de Sequência de RNARESUMO
Commonly available clinical parameters fail to predict early acute cellular rejection (EAR, occurring within 6 months after transplant), a major risk factor for graft loss after kidney transplantation. We performed whole-blood RNA sequencing at the time of transplant in 235 kidney transplant recipients enrolled in a prospective cohort study (Genomics of Chronic Allograft Rejection [GoCAR]) and evaluated the relationship of pretransplant transcriptomic profiles with EAR. EAR was associated with downregulation of NK and CD8+ T cell gene signatures in pretransplant blood. We identified a 23-gene set that predicted EAR in the discovery (n = 81, and AUC = 0.80) and validation (n = 74, and AUC = 0.74) sets. Exclusion of recipients with 5 or 6 HLA donor mismatches increased the AUC to 0.89. The risk score derived from the gene set was also significantly associated with acute cellular rejection after 6 months, antibody-mediated rejection and/or de novo donor-specific antibodies, and graft loss in a cohort of 154 patients, combining the validation set and additional GoCAR patients with surveillance biopsies between 6 and 24 months (n = 80) posttransplant. This 23-gene set is a potentially important new tool for determination of the recipient's immunological risk before kidney transplantation, and facilitation of an individualized approach to immunosuppressive therapy.
Assuntos
Rejeição de Enxerto , Transplante de Rim/efeitos adversos , Transcriptoma/genética , Adulto , Biomarcadores/sangue , Biomarcadores/metabolismo , Feminino , Rejeição de Enxerto/diagnóstico , Rejeição de Enxerto/epidemiologia , Rejeição de Enxerto/genética , Rejeição de Enxerto/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Estudos Prospectivos , Medição de RiscoRESUMO
BK polyomavirus (BKV) is an important cause of graft loss in renal transplant recipients that continues to pose a significant challenge to clinicians due to its frequently unpredictable onset, persistence, and the lack of effective antiviral agents or prevention strategies. This review covers our current understanding of epidemiology, viral transmission and disease progression, and treatment and prevention strategies that have been used to manage this disease.
Assuntos
Vírus BK/patogenicidade , Hospedeiro Imunocomprometido , Imunossupressores/efeitos adversos , Transplante de Rim , Infecções por Polyomavirus/patologia , Vírus BK/crescimento & desenvolvimento , Progressão da Doença , Transmissão de Doença Infecciosa , Humanos , Imunossupressores/uso terapêutico , Infecções por Polyomavirus/epidemiologia , Infecções por Polyomavirus/transmissão , Infecções por Polyomavirus/virologiaRESUMO
PURPOSE OF REVIEW: Improved long-term kidney allograft survival remains a critical goal in transplantation; the achievement of this, however, is highly dependent on the identification of biomarkers that can either predict or allow advance detection of patients at risk of allograft injury. The present review outlines the commonly used biomarkers in kidney transplantation, while also highlighting those currently under investigation, discussing their advantages and limitations. RECENT FINDINGS: Most of the approved biomarkers currently used in kidney transplantation capture antigen recognition or alloantibody production. However, tremendous progress has recently been made in the development of markers of other signaling pathways pertinent to the alloimmune response. Microarray gene sets that predict rejection or poor prognostic phenotypes have been identified in kidney biopsies (the 'molecular microscope diagnostic system' and the 'genomics of chronic allograft rejection' scores), peripheral blood (the 'kidney solid organ response test'), and urine (the '3-genes signature'). Strategies targeting serial measurements of urinary chemokines such as CXCL9 and CXCL10 also appear promising. SUMMARY: Although the range of biomarkers in current use is limited, there are many assays in the development and validation pipeline that appear promising but that have yet to reach mainstream clinical transplantation. The 'ideal biomarker' may eventually transpire to be the combination of several assays.
Assuntos
Biomarcadores/metabolismo , Rejeição de Enxerto/diagnóstico , Rejeição de Enxerto/metabolismo , Isoanticorpos/sangue , Transplante de Rim , Quimiocina CXCL10/urina , Quimiocina CXCL9/urina , Expressão Gênica , Genômica , Rejeição de Enxerto/genética , Humanos , Transplante de Rim/efeitos adversosRESUMO
The development of de novo anti-HLA donor-specific antibodies (dnDSA) is associated with poorer outcomes in kidney transplant recipients. Despite this, antibody screening post-transplant is not widespread, largely because the optimal management of patients with dnDSA remains undetermined. We hypothesized that in this population, calcineurin inhibitor blood levels would be an independent predictor of graft loss. We analyzed a cohort of unsensitized patients for whom anti-HLA antibody screening was performed prospectively post-transplant. During the screening period between January 2005 and April 2016, 42 patients developed dnDSA. There was no difference in the clinical characteristics or the histological scores of patients biopsied for clinical indication versus those biopsied solely due to detection of dnDSA. Cox modeling revealed a strong relationship between mean tacrolimus levels following dnDSA detection and graft loss, with a hazard ratio of 0.49 (95% CI, 0.33-0.75), which persisted following adjustment for established independent predictors (HR, 0.52, 95% CI, 0.30-0.89). Kaplan-Meier analysis by tertiles of tacrolimus levels and receiver operating curve analysis concurred to show that a threshold of 5.3 ng/ml could be predictive of graft loss. These data suggest that anti-HLA antibody monitoring post-transplant could guide maintenance immunosuppression and improve graft outcomes.
Assuntos
Inibidores de Calcineurina/sangue , Sobrevivência de Enxerto/imunologia , Antígenos HLA/imunologia , Transplante de Rim , Tacrolimo/sangue , Adulto , Idoso , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
PURPOSE OF REVIEW: The Notch signaling pathway is known to play a pivotal role in T- and B-cell development and fate, presenting it as an attractive therapeutic target in alloimmunity. This review provides an overview of the mechanisms of Notch signaling, focusing on new insights into its diverse functions in T-cell activation, differentiation and memory subset formation, and the consequences thereof in transplantation. RECENT FINDINGS: Recent evidence has shown that while not critical for early antigen-specific CD4 T-cell activation, Notch signaling regulates the survival of memory CD4 T cells via control of glycolytic metabolism; in contrast, Notch signaling is critical for the generation of short-lived CD8 effector T cells, but not memory CD8 cells. Transient, selective inhibition of various Notch receptors and ligands in models of solid organ transplantation has been shown to successfully modulate the alloimmune response, affecting the balance between effector and regulatory cells, with particular influence on the natural regulatory T-cell population. SUMMARY: These studies reveal diverse roles for individual Notch receptors and ligands in peripheral immunity and indicate that selective targeting of the Notch pathway is a promising, novel approach for immune modulation in transplantation; the advent of therapeutic human antibodies to neutralize both the Notch ligands and the individual Notch receptors suggests that this approach could be efficiently developed.
Assuntos
Receptores Notch/imunologia , Transdução de Sinais , Animais , Diferenciação Celular/imunologia , Humanos , Ligantes , Ativação Linfocitária , Linfócitos T/citologia , Linfócitos T/imunologiaRESUMO
A high-salt diet (HSD) in humans is linked to a number of complications, including hypertension and cardiovascular events. Whether a HSD affects the immune response in transplantation is unknown. Using a murine transplantation model, we investigated the effect of NaCl on the alloimmune response in vitro and in vivo. Incremental NaCl concentrations in vitro augmented T cell proliferation in the settings of both polyclonal and allospecific stimulation. Feeding a HSD to C57BL/6 wild-type recipients of bm12 allografts led to accelerated cardiac allograft rejection, despite similar mean BP and serum sodium levels in HSD and normal salt diet (NSD) groups. The accelerated rejection was associated with a reduction in the proportion of CD4(+)Foxp3(+) regulatory T cells (Tregs) and a significant decrease in Treg proliferation, leading to an increased ratio of antigen-experienced CD4(+) T cells to Tregs in mice recipients of a HSD compared with mice recipients of a NSD. Because serum- and glucocorticoid-regulated kinase-1 (SGK1) has been proposed as a potential target of salt in immune cells, we fed a HSD to CD4(Cre)SGK1(fl/fl) B6-transplanted recipients and observed abrogation of the deleterious effect of a HSD in the absence of SGK1 on CD4(+) cells. In summary, we show that NaCl negatively affects the regulatory balance of T cells in transplantation and precipitates rejection in an SGK1-dependent manner.
Assuntos
Rejeição de Enxerto/induzido quimicamente , Proteínas Imediatamente Precoces/efeitos dos fármacos , Proteínas Imediatamente Precoces/fisiologia , Proteínas Serina-Treonina Quinases/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/fisiologia , Cloreto de Sódio na Dieta/efeitos adversos , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/fisiologia , Animais , Camundongos , Camundongos Endogâmicos C57BL , Fatores de TempoRESUMO
Regulatory T cells (Tregs) are amongst the most widely studied cells in a variety of immune-mediated conditions, including transplantation and Graft Versus Host Disease (GVHD), cancer and autoimmunity; indeed, there is great interest in the tolerogenic potential of Treg-based therapy. Consequently, the need to establish the mechanisms that determine Treg survival and longevity, in addition to developing new tools to monitor these parameters, is paramount. Using both a mouse model of GVHD and a mouse model of Type 1 Diabetes (T1D), we describe herein a dual reporter system based on Gluc and multiplexed with SEAP and non-secreted Firefly luciferase (Fluc), which permits simultaneous imaging and noninvasive tracking of two different T-cell populations (CD4(+)CD25(+) Tregs and CD4(+)CD25(-) Tcon cells) in vivo by transducing the cells with different lentiviruses bearing distinct color signatures. This new technology promises to overcome the limitations of the conventional methods currently available to study lymphocyte survival in vivo. Furthermore, this novel technique has applications not only in autoimmunity and alloimmunity, but also in the wider field of immunology.
Assuntos
Fosfatase Alcalina/metabolismo , Copépodes/enzimologia , Luciferases de Vaga-Lume/metabolismo , Medições Luminescentes , Imagem Molecular/métodos , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Reguladores/citologia , Animais , Células da Medula Óssea/citologia , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica/imunologia , Células HEK293 , Humanos , Lentivirus/genética , Luciferases de Vaga-Lume/genética , Camundongos , Células Precursoras de Linfócitos T/citologia , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Reguladores/metabolismo , Fatores de TempoRESUMO
Dendritic cells (DCs) are the central architects of the immune response, inducing inflammatory or tolerogenic immunity, dependent on their activation status. As such, DCs are highly attractive therapeutic targets and may hold the potential to control detrimental immune responses. TIM-4, expressed on APCs, has complex functions in vivo, acting both as a costimulatory molecule and a phosphatidylserine receptor. The effect of TIM-4 costimulation on T cell activation remains unclear. In this study, we demonstrate that Ab blockade of DC-expressed TIM-4 leads to increased induction of induced regulatory T cells (iTregs) from naive CD4(+) T cells, both in vitro and in vivo. iTreg induction occurs through suppression of IL-4/STAT6/Gata3-induced Th2 differentiation. In addition, blockade of TIM-4 on previously activated DCs still leads to increased iTreg induction. iTregs induced under TIM-4 blockade have equivalent potency to control and, upon adoptive transfer, significantly prolong skin allograft survival in vivo. In RAG(-/-) recipients of skin allografts adoptively transferred with CD4(+) T cells, we show that TIM-4 blockade in vivo is associated with a 3-fold prolongation in allograft survival. Furthermore, in this mouse model of skin transplantation, increased induction of allospecific iTregs and a reduction in T effector responses were observed, with decreased Th1 and Th2 responses. This enhanced allograft survival and protolerogenic skewing of the alloresponse is critically dependent on conversion of naive CD4(+) to Tregs in vivo. Collectively, these studies identify blockade of DC-expressed TIM-4 as a novel strategy that holds the capacity to induce regulatory immunity in vivo.
Assuntos
Células Dendríticas/imunologia , Sobrevivência de Enxerto , Proteínas de Membrana/metabolismo , Transplante de Pele , Linfócitos T Reguladores/imunologia , Transferência Adotiva , Aloenxertos/imunologia , Animais , Anticorpos Monoclonais/imunologia , Diferenciação Celular/imunologia , Células Cultivadas , Fator de Transcrição GATA3/imunologia , Proteínas de Homeodomínio/genética , Interleucina-4/imunologia , Ativação Linfocitária , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Transcrição STAT6/imunologia , Transdução de Sinais , Linfócitos T Reguladores/metabolismo , Células Th1/imunologia , Células Th2/imunologiaRESUMO
Regulatory T cells (Tregs) play a pivotal role in the maintenance of immune tolerance and hold great promise as cell therapy for a variety of immune-mediated diseases. However, the cellular mechanisms that regulate Treg maintenance and homeostasis have yet to be fully explored. Although Tregs express granzyme-B (GrB) to suppress effector T cells via direct killing, the mechanisms by which they protect themselves from GrB-mediated self-inflicted damage are unknown. To our knowledge, we show for the first time that both induced Tregs and natural Tregs (nTregs) increase their intracellular expression of GrB and its endogenous inhibitor, serine protease inhibitor 6 (Spi6) upon activation. Subcellular fractionation and measurement of GrB activity in the cytoplasm of Tregs show that activated Spi6(-/-) Tregs had significantly higher cytoplasmic GrB activity. We observed an increase in GrB-mediated apoptosis in Spi6(-/-) nTregs and impaired suppression of alloreactive T cells in vitro. Spi6(-/-) Tregs were rescued from apoptosis by the addition of a GrB inhibitor (Z-AAD-CMK) in vitro. Furthermore, adoptive transfer experiments showed that Spi6(-/-) nTregs were less effective than wild type nTregs in suppressing graft-versus-host disease because of their impaired survival, as shown in our in vivo bioluminescence imaging. Finally, Spi6-deficient recipients rejected MHC class II-mismatch heart allografts at a much faster rate and showed a higher rate of apoptosis among Tregs, as compared with wild type recipients. To our knowledge, our data demonstrate, for the first time, a novel role for Spi6 in Treg homeostasis by protecting activated Tregs from GrB-mediated injury. These data could have significant clinical implications for Treg-based therapy in immune-mediated diseases.
Assuntos
Granzimas/imunologia , Homeostase/imunologia , Proteínas de Membrana/imunologia , Serina Endopeptidases/imunologia , Serpinas/imunologia , Linfócitos T Reguladores/imunologia , Animais , Apoptose/imunologia , Citometria de Fluxo , Doença Enxerto-Hospedeiro/imunologia , Granzimas/metabolismo , Transplante de Coração/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Confocal , Serina Endopeptidases/metabolismo , Serpinas/metabolismo , Linfócitos T Reguladores/metabolismoRESUMO
The Notch pathway is an important intercellular signaling pathway that plays a major role in controlling cell fate. Accumulating evidence indicates that Notch and its ligands present on antigen-presenting cells might be important mediators of T helper cell differentiation. In this study, we investigated the role of Jagged2 in murine cardiac transplantation by using a signaling Jagged2 mAb (Jag2) that activates recombinant signal-binding protein-Jκ. While administration of Jag2 mAb had little effect on graft survival in the fully allogeneic mismatched model BALB/câB6, it hastened rejection in CD28-deficient recipients. Similarly, Jag2 precipitated rejection in the bm12âB6 model. In this MHC class II-mismatched model, allografts spontaneously survive for >56 days due to the emergence of Treg cells that inhibit the expansion of alloreactive T cells. The accelerated rejection was associated with upregulation of Th2 cytokines and proinflammatory cytokine IL-6, despite expansion of Treg cells. Incubation of Treg cells with recombinant IL-6 abrogated their inhibitory effects in vitro. Furthermore, neutralization of IL-6 in vivo protected Jag2-treated recipients from rejection and Jagged2 signaling was unable to further accelerate rejection in the absence of Treg cells. Our findings therefore suggest that Jagged2 signaling can affect graft acceptance by upregulation of IL-6 and consequent resistance to Treg-cell suppression.
Assuntos
Rejeição de Enxerto/imunologia , Transplante de Coração/imunologia , Interleucina-6/imunologia , Proteínas de Membrana/metabolismo , Linfócitos T Reguladores/efeitos dos fármacos , Células Th2/efeitos dos fármacos , Animais , Anticorpos Bloqueadores/administração & dosagem , Anticorpos Monoclonais/administração & dosagem , Antígenos CD28/genética , Células Cultivadas , Antígenos de Histocompatibilidade/imunologia , Proteína Jagged-2 , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Linfócitos T Reguladores/imunologia , Células Th2/imunologiaRESUMO
BACKGROUND: Despite significant nephrotoxicity, calcineurin inhibitors (CNIs) remain the cornerstone of immunosuppression in solid organ transplantation. We, along with others, have reported tolerogenic properties of anti-thymocyte globulin (ATG, Thymoglobulin®), evinced by its ability both to spare Tregs from depletion in vivo and, when administered at low, non-depleting doses, to expand Tregs ex vivo. Clinical trials investigating B7/CD28 blockade (LEA29Y, Belatacept) in kidney transplant recipients have proven that the replacement of toxic CNI use is feasible in selected populations. METHODS: Rabbit polyclonal anti-murine thymocyte globulin (mATG) was administered as induction and/or prolonged, low-dose therapy, in combination with CTLA4-Ig, in a stringent, fully MHC-mismatched murine skin transplant model to assess graft survival and mechanisms of action. RESULTS: Prolonged, low-dose mATG, combined with CTLA4-Ig, effectively promotes engraftment in a stringent transplant model. Our data demonstrate that mATG achieves graft acceptance primarily by promoting Tregs, while CTLA4-Ig enhances mATG function by limiting activation of the effector T cell pool in the early stages of treatment, and by inhibiting production of anti-rabbit antibodies in the maintenance phase, thereby promoting regulation of alloreactivity. CONCLUSION: These data provide the rationale for development of novel, CNI-free clinical protocols in human transplant recipients.
Assuntos
Soro Antilinfocitário/administração & dosagem , Imunoconjugados/administração & dosagem , Terapia de Imunossupressão , Imunossupressores/administração & dosagem , Transplante de Órgãos , Abatacepte , Animais , Relação Dose-Resposta a Droga , Sobrevivência de Enxerto/efeitos dos fármacos , Sobrevivência de Enxerto/imunologia , Humanos , Tolerância Imunológica/efeitos dos fármacos , Imunoconjugados/metabolismo , Camundongos , Coelhos , Transplante Homólogo/imunologiaRESUMO
Ligands of the B7 family provide both positive and negative costimulatory signals to the CD28 family of receptors on T lymphocytes, the balance of which determines the immune response. B7-H3 is a member of the B7 family whose function in T-cell activation has been the subject of some controversy: in autoimmunity and tumor immunity, it has been described as both costimulatory and coinhibitory, while in transplantation, B7-H3 signaling is thought to contribute to graft rejection. However, we now demonstrate results to the contrary. Signaling through a putative B7-H3 receptor prolonged allograft survival in a fully MHC-mismatched cardiac model and promoted a shift toward a Th2 milieu; conversely, B7-H3 blockade, achieved by use of a blocking antibody, resulted in accelerated rejection, an effect associated with enhanced IFN-γ production. Finally, graft prolongation achieved by CTLA4 Ig was shortened both by B7-H3 blockade and the absence of recipient B7-H3. These findings suggest a coinhibitory role for B7-H3. However, experience with other CD28/B7 family members suggests that immune redundancy plays a crucial role in determining the functions of various pathways. Given the abundance of conflicting data, it is plausible that, under differing conditions, B7-H3 may have both positive and negative costimulatory functions.
Assuntos
Antígenos B7/imunologia , Transplante de Coração/imunologia , Transdução de Sinais/imunologia , Células Th1/imunologia , Transplante Homólogo/imunologia , Abatacepte , Animais , Antígenos B7/metabolismo , Antígenos CD28/imunologia , Antígenos CD28/metabolismo , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/metabolismo , Imunoconjugados/imunologia , Imunoconjugados/metabolismo , Interferon gama/imunologia , Interferon gama/metabolismo , Cinética , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Células Th1/metabolismo , Células Th2/imunologia , Células Th2/metabolismo , Imunologia de TransplantesRESUMO
BACKGROUND AND OBJECTIVES: Acute rejection remains a problem in renal transplantation. This study sought to determine the utility of a noninvasive cytokine assay in screening of acute rejection. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: In this observational cross-sectional study, 64 patients from two centers were recruited upon admission for allograft biopsy to investigate acute graft dysfunction. Blood was collected before biopsy and assayed for a panel of 21 cytokines secreted by PBMCs. Patients were classified as acute rejectors or nonrejectors according to a classification rule derived from an initial set of 32 patients (training cohort) and subsequently validated in the remaining patients (validation cohort). RESULTS: Although six cytokines (IL-1ß, IL-6, TNF-α, IL-4, GM-CSF, and monocyte chemoattractant protein-1) distinguished acute rejectors in the training cohort, logistic regression modeling identified a single cytokine, IL-6, as the best predictor. In the validation cohort, IL-6 was consistently the most accurate cytokine (area under the receiver-operating characteristic curve, 0.85; P=0.006), whereas the application of a prespecified cutoff level, as determined from the training cohort, resulted in a sensitivity and specificity of 92% and 63%, respectively. Secondary analyses revealed a strong association between IL-6 levels and acute rejection after multivariate adjustment for clinical characteristics (P<0.001). CONCLUSIONS: In this pilot study, the measurement of a single cytokine can exclude acute rejection with a sensitivity of 92% in renal transplant recipients presenting with acute graft dysfunction. Prospective studies are needed to determine the utility of this simple assay, particularly for low-risk or remote patients.
Assuntos
Rejeição de Enxerto/diagnóstico , Interleucina-6/sangue , Transplante de Rim/imunologia , Doença Aguda , Adulto , Idoso , Biomarcadores/sangue , Boston , Células Cultivadas , Quimiocina CCL2/sangue , Estudos Transversais , Feminino , Rejeição de Enxerto/sangue , Rejeição de Enxerto/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/sangue , Humanos , Interleucina-1beta/sangue , Interleucina-4/sangue , Transplante de Rim/efeitos adversos , Leucócitos Mononucleares/imunologia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Projetos Piloto , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Medição de Risco , Fatores de Risco , Sensibilidade e Especificidade , Fatores de Tempo , Resultado do Tratamento , Fator de Necrose Tumoral alfa/sangueRESUMO
The incidence of developing circulating anti-human leukocyte antigen antibodies and the kinetics of T cell depletion and recovery among pediatric renal transplant recipients who receive alemtuzumab induction therapy are unknown. In a collaborative endeavor to minimize maintenance immunosuppression in pediatric renal transplant recipients, we enrolled 35 participants from four centers and treated them with alemtuzumab induction therapy and a steroid-free, calcineurin-inhibitor-withdrawal maintenance regimen. At 3 months after transplant, there was greater depletion of CD4(+) than CD8(+) T cells within the total, naive, memory, and effector memory subsets, although depletion of the central memory subset was similar for CD4(+) and CD8(+) cells. Although CD8(+) T cells recovered faster than CD4(+) subsets overall, they failed to return to pretransplant levels by 24 months after transplant. There was no evidence for greater recovery of either CD4(+) or CD8(+) memory cells than naïve cells. Alemtuzumab relatively spared CD4(+)CD25(+)FoxP3(+) regulatory T cells, resulting in a rise in their numbers relative to total CD4(+) cells and a ratio that remained at least at pretransplant levels throughout the study period. Seven participants (20%) developed anti-human leukocyte antigen antibodies without adversely affecting allograft function or histology on 2-year biopsies. Long-term follow-up is underway to assess the potential benefits of this regimen in children.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Antineoplásicos/farmacologia , Antineoplásicos/farmacologia , Antígenos HLA/imunologia , Transplante de Rim/imunologia , Linfócitos T/efeitos dos fármacos , Adolescente , Alemtuzumab , Criança , Feminino , Humanos , Terapia de Imunossupressão , Masculino , Estudos ProspectivosRESUMO
BACKGROUND: Although there is ample evidence about the role of adaptive immunity in the development of chronic allograft dysfunction, little is known about the contribution of innate immunity to this process. Herein, we studied the relationship between inflammation, chronic biopsy scores, and anti-human leukocyte antigen (HLA) circulating alloantibodies in a cohort of 57 patients recruited at our center. METHODS: Available biopsies (n=27) were graded for chronic lesion scores according to Banff criteria. The production of cytokines by peripheral blood mononuclear cells after 48 hr of culture under resting conditions was quantified by Luminex. Tumor necrosis factor (TNF)-α secretion assay and depletion studies were used to identify the source of these cytokines. RESULTS: There was a high correlation between the levels of interleukin (IL)-1ß, IL-6, and TNF-α (r>0.8, P<0.001 for all correlations). The levels of these cytokines were associated with transplant glomerulopathy (IL-1ß, P=0.019; IL-6, P=0.015; and TNF-α, P=0.006) but not with other chronic lesions or anti-HLA circulating alloantibodies. TNF-α was predominantly secreted by monocytes (percent of TNF-α secreting cells: 20.4±4.8 vs. 1.2±0.5 vs. 1.4±0.6 vs. 1.7±0.5 for CD14, CD4, CD8, and CD19 cells, respectively; all P<0.01 vs. CD14). The levels of all three proinflammatory cytokines were significantly reduced after monocyte depletion. Intriguingly, cytokine levels increased after ex vivo depletion of regulatory T cells (all P<0.001). CONCLUSIONS: Taken together, these data suggest that in vivo-activated monocytes in peripheral blood spontaneously secrete proinflammatory cytokines in renal allograft recipients with transplant glomerulopathy and seem to be under the regulation of functional regulatory T cells in this setting.
Assuntos
Glomerulonefrite/patologia , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Transplante de Rim/patologia , Monócitos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Idoso , Biópsia , Antígenos CD4/metabolismo , Células Cultivadas , Estudos de Coortes , Estudos Transversais , Feminino , Glomerulonefrite/epidemiologia , Glomerulonefrite/imunologia , Humanos , Imunidade Inata/imunologia , Incidência , Transplante de Rim/imunologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Linfócitos T Reguladores/fisiologia , Transplante HomólogoRESUMO
Rhabdomyolysis is a known complication of statin therapy and may be triggered by a pharmacokinetic interaction between a statin and a second medication. Fatal statin-induced rhabdomyolysis has an incidence of 0.15 deaths/million prescriptions. We describe 4 cases of severe rhabdomyolysis with the common feature of atorvastatin use and coadministration of fusidic acid. All cases involved long-term therapy with atorvastatin; fusidic acid was introduced for treatment of osteomyelitis or septic arthritis. Three cases occurred in the setting of diabetes mellitus, with 2 in patients with end-stage renal disease, suggesting increased susceptibility to atorvastatin-fusidic acid-induced rhabdomyolysis in these patient populations. Of the 4 patients in this series, 3 died. Fusidic acid is a unique bacteriostatic antimicrobial agent with principal antistaphylococcal activity. There have been isolated reports of rhabdomyolysis attributed to the interaction of statins and fusidic acid, the cause of which is unclear. Fusidic acid does not inhibit the cytochrome P450 3A4 isoenzyme responsible for atorvastatin metabolism; increased atorvastatin levels due to inhibition of the glucuronidation pathway may be responsible. Considering the low frequency of fusidic acid use, the appearance of 4 such cases within a short time and in a small population suggests the probability that development of this potentially fatal complication may be relatively high.