Pathways of major histocompatibility complex allorecognition

PURPOSE OF REVIEW: Here, we review the pathways of allorecognition and their potential relevance to the balance between regulatory and effector responses following transplantation. RECENT FINDINGS: Transplantation between nonidentical members of the same species elicits an immune response that manifests as graft rejection or persistence. Presentation of foreign antigen to recipient T cells can occur via three nonmutually exclusive routes, the direct, indirect and semi-direct pathways. Allospecific T cells can have effector or regulatory functions, and the relative proportions of the two populations activated following alloantigen presentation are two of the factors that determine the clinical outcome. Regulatory T cells have been the subject of significant research, and there is now greater understanding of their recruitment and function in the context of allorecognition. SUMMARY: A greater understanding of the mechanisms underlying allorecognition may be fundamental to appreciating how these different populations are recruited and could in turn inform novel strategies for immunomodulation

Dendritic cells in transplantation - Friend or foe?

The existence of two sets of dendritic cells in transplantation (of donor and recipient origin) poses unique problems in alloimmunity and tolerance. Donor DCs are potent stimulators of the direct pathway, in which recipient T cells respond to peptides presented on donor MHC products or to the donor MHC molecules themselves. Reduced DC function in the direct pathway is used to explain the acceptance of certain allografts that have been depleted of passenger leukocytes. Reciprocally, purified donor DCs powerfully stimulate the rejection of these grafts as well as the mixed leukocyte reaction by purified allogeneic T cells in culture. Donor DCs also can act as specialized antigen transport vehicles in cross-priming for the indirect pathway. Here, recipient DCs process MHC molecules from the donor. The indirect pathway of rejection is actually the rule for passenger leukocyte-depleted grafts. Furthermore, the indirect pathway seems to be the dominant alloresponse detected in long-term graft recipients, both in experimental models and clinical transplantation, particularly in those with chronic rejection. The stimulatory function of DCs in both direct and indirect pathways is regulated by their maturation in response to inflammatory stimuli, especially those delivered via IL-1, TNF, and Toll receptor families. Since the normal function of DCs is to generate immunity against invading pathogens, the indirect response to peptides continually derived from the allogeneic MHC molecules in a tissue allograft (a surgically introduced pathogen because of the associated inflammation and necrosis) may be more difficult to silence. However, in addition to their allostimulatory role, immature or in vitro-manipulated DCs also have the potential to downregulate direct and indirect antidonor responses through a variety of mechanisms. Intriguing new evidence shows that immature DCs can actively induce regulatory T cells. Given the probable role of the indirect pathway in driving chronic rejection, the induction of T cell tolerance (deletion, anergy, and regulation), especially in those T cells with indirect antidonor allospecificity, by the injection of immature DCs pretransplant, could serve as a critical therapeutic strategy

Detection of donor-specific hyporesponsiveness following late failure of human renal allografts

Detection of donor-specific hyporesponsiveness following late failure of human renal allografts. Limiting dilution assays to measure the frequency of interleukin-2-secreting peripheral blood T cells were carried out in patients, whose renal allografts had failed due to acute rejection (9 patients) and in patients whose grafts failed more than two years after transplantation without any recent evidence of acute rejection. Using a modified form of the assay we demonstrate that nearly half of 18 patients whose renal transplants had failed after more than two years have low or undetectable HTLp frequencies against donor, but not third-party DR antigens. No such difference was observed in any of the nine patients studied whose transplants were lost from early acute rejection. These results provide the first indication that, as in rodent models of transplantation, T cell unresponsiveness towards donor MHC antigens can occur following prolonged residence of an allograft in humans. Furthermore, the results suggest that chronic rejection may be driven by mechanisms other than direct allorecognition. The assay may be a valuable tool to study the evolution of donor-specific direct T cell alloresponsiveness in patients with well-functioning grafts

HLA-DP Allele-Specific T Cell Responses to Beryllium Account for DP-Associated Susceptibility to Chronic Beryllium Disease

Abstract Occupational exposure to small molecules, such as metals, is frequently associated with hypersensitivity reactions. Chronic beryllium (Be) disease (CBD) is a multisystem granulomatous disease that primarily affects the lung, and occurs in ∼3% of individuals exposed to this element. Immunogenetic studies have demonstrated a strong association between CBD and possession of alleles of HLA-DP containing glutamic acid (Glu) at position 69 in the HLA-DPβ-chain. T cell clones were raised from three patients with CBD in whom exposure occurred 10 and 30 years previously. Of 25 Be-specific clones that were obtained, all were restricted by HLA-DP alleles with Glu at DPβ69. Furthermore, the proliferative responses of the clones were absolutely dependent upon DPβ Glu69 in that a single amino acid substitution at this position abolished the response. As befits a disease whose pathogenesis involves a delayed type hypersensitivity response, the large majority of Be-specific clones secreted IFN-γ (Th1) and little or no IL-4 (Th2) cytokines. This study provides insights into the molecular basis of DP2-associated susceptibility to CBD

Inhibition of Thrombin Receptor Signaling on alpha-Smooth Muscle Actin(+) CD34(+) Progenitors Leads to Repair After Murine Immune Vascular Injury

OBJECTIVE: The goal of this study was to use mice expressing human tissue factor pathway inhibitor (TFPI) on α-smooth muscle actin (α-SMA)(+) cells as recipients of allogeneic aortas to gain insights into the cellular mechanisms of intimal hyperplasia (IH). METHODS AND RESULTS: BALB/c aortas (H-2(d)) transplanted into α-TFPI-transgenic (Tg) mice (H-2(b)) regenerated a quiescent endothelium in contrast to progressive IH seen in C57BL/6 wild-type (WT) mice even though both developed aggressive anti-H-2(d) alloresponses, indicating similar vascular injuries. Adoptively transferred Tg CD34(+) (but not CD34(-)) cells inhibited IH in WT recipients, indicating the phenotype of α-TFPI-Tg mice was due to these cells. Compared with syngeneic controls, endogenous CD34(+) cells were mobilized in significant numbers after allogeneic transplantation, the majority showing sustained expression of tissue factor and protease-activated receptor-1 (PAR-1). In WT, most were CD45(+) myeloid progenitors coexpressing CD31, vascular endothelial growth factor receptor-2 and E-selectin; 10% of these cells coexpressed α-SMA and were recruited to the neointima. In contrast, the α-SMA(+) human TFPI(+) CD34(+) cells recruited in Tg recipients were from a CD45(-) lineage. WT CD34(+) cells incubated with a PAR-1 antagonist or taken from PAR-1-deficient mice inhibited IH as Tg cells did. CONCLUSIONS: Specific inhibition of thrombin generation or PAR-1 signaling on α-SMA(+) CD34(+) cells inhibits IH and promotes regenerative repair despite ongoing immune-mediated damage

Constitutive expression of the anti-apoptotic Bcl-2 family member A1 in murine endothelial cells leads to transplant tolerance

Anti-apoptotic genes including those of the Bcl-2 family have been shown to have dual functionality in as much as they inhibit cell death but also regulate inflammation. Several anti-apoptotic molecules have been associated with endothelial cell (EC) survival following transplantation however their exact role has yet to be elucidated in respect to controlling inflammation. In this study we created mice expressing murine A1 (Bfl-1), a Bcl-2 family member, under the control of the human ICAM-2 promoter. Constitutive expression of A1 in murine vascular ECs conferred protection from cell death induced by the pro-inflammatory cytokine TNF-α. Importantly, in a mouse model of heart allograft transplantation, expression of A1 in vascular endothelium increased survival in the absence of CD8+ T cells. Better graft outcome in mice receiving an A1 transgenic heart correlated with a reduced immune infiltration, which maybe related to increased EC survival and reduced expression of adhesion molecules on ECs. In conclusion, constitutive expression of the anti-apoptotic molecule Bfl1 (A1) in murine vascular ECs leads to prolonged allograft survival due to modifying inflammation

Monitoring of In Vivo Function of Superparamagnetic Iron Oxide Labelled Murine Dendritic Cells during Anti-Tumour Vaccination

Dendritic cells (DCs) generated in vitro to present tumour antigens have been injected in cancer patients to boost in vivo anti-tumour immune responses. This approach to cancer immunotherapy has had limited success. For anti-tumour therapy, delivery and subsequent migration of DCs to lymph nodes leading to effective stimulation of effector T cells is thought to be essential. The ability to non-invasively monitor the fate of adoptively transferred DCs in vivo using magnetic resonance imaging (MRI) is an important clinical tool to correlate their in vivo behavior with response to treatment. Previous reports of superparamagnetic iron oxides (SPIOs) labelling of different cell types, including DCs, have indicated varying detrimental effects on cell viability, migration, differentiation and immune function. Here we describe an optimised labelling procedure using a short incubation time and low concentration of clinically used SPIO Endorem to successfully track murine DC migration in vivo using MRI in a mouse tumour model. First, intracellular labelling of bone marrow derived DCs was monitored in vitro using electron microscopy and MRI relaxometry. Second, the in vitro characterisation of SPIO labelled DCs demonstrated that viability, phenotype and functions were comparable to unlabelled DCs. Third, ex vivo SPIO labelled DCs, when injected subcutaneously, allowed for the longitudinal monitoring by MR imaging of their migration in vivo. Fourth, the SPIO DCs induced the proliferation of adoptively transferred CD4+ T cells but, most importantly, they primed cytotoxic CD8+ T cell responses to protect against a B16-Ova tumour challenge. Finally, using anatomical information from the MR images, the immigration of DCs was confirmed by the increase in lymph node size post-DC injection. These results demonstrate that the SPIO labelling protocol developed in this study is not detrimental for DC function in vitro and in vivo has potential clinical application in monitoring therapeutic DCs in patients with cancer

Regulatory T Cell-Derived Exosomes: Possible Therapeutic and Diagnostic Tools in Transplantation

Exosomes are extracellular vesicles released by many cells of the body. These small vesicles play an important part in intercellular communication both in the local environment and systemically, facilitating in the transfer of proteins, cytokines as well as miRNA between cells. The observation that exosomes isolated from immune cells such as dendritic cells (DCs) modulate the immune response has paved the way for these structures to be considered as potential immunotherapeutic reagents. Indeed, clinical trials using DC derived exosomes to facilitate immune responses to specific cancer antigens are now underway. Exosomes can also have a negative effect on the immune response and exosomes isolated from regulatory T cells (Tregs) and other subsets of T cells have been shown to have immune suppressive capacities. Here, we review what is currently known about Treg derived exosomes and their contribution to immune regulation, as well as highlighting their possible therapeutic potential for preventing graft rejection, and use as diagnostic tools to assess transplant outcome

In Vivo SPECT Reporter Gene Imaging of Regulatory T Cells

Regulatory T cells (Tregs) were identified several years ago and are key in controlling autoimmune diseases and limiting immune responses to foreign antigens, including alloantigens. In vivo imaging techniques including intravital microscopy as well as whole body imaging using bioluminescence probes have contributed to the understanding of in vivo Treg function, their mechanisms of action and target cells. Imaging of the human sodium/iodide symporter via Single Photon Emission Computed Tomography (SPECT) has been used to image various cell types in vivo. It has several advantages over the aforementioned imaging techniques including high sensitivity, it allows non-invasive whole body studies of viable cell migration and localisation of cells over time and lastly it may offer the possibility to be translated to the clinic. This study addresses whether SPECT/CT imaging can be used to visualise the migratory pattern of Tregs in vivo. Treg lines derived from CD4+CD25+FoxP3+ cells were retrovirally transduced with a construct encoding for the human Sodium Iodide Symporter (NIS) and the fluorescent protein mCherry and stimulated with autologous DCs. NIS expressing self-specific Tregs were specifically radiolabelled in vitro with Technetium-99m pertechnetate (99mTcO4−) and exposure of these cells to radioactivity did not affect cell viability, phenotype or function. In addition adoptively transferred Treg-NIS cells were imaged in vivo in C57BL/6 (BL/6) mice by SPECT/CT using 99mTcO4−. After 24 hours NIS expressing Tregs were observed in the spleen and their localisation was further confirmed by organ biodistribution studies and flow cytometry analysis. The data presented here suggests that SPECT/CT imaging can be utilised in preclinical imaging studies of adoptively transferred Tregs without affecting Treg function and viability thereby allowing longitudinal studies within disease models