Search

Background: Extracellular adenosine triphosphate (ATP) binds to purinergic receptors and promotes inflammatory responses. We tested whether oxidized ATP (oATP), P2X7 receptor antagonist can attenuate acute kidney allograft rejection. Methods: Brown Norway kidney allografts were transplanted into Lewis recipients. Three groups were defined: oATP (n=8), Cyclosporine A (n=6), and no treatment (n=8). On day seven, we assessed kidney allograft survival, function, and rejection characteristics. We further determined T-cell, B-cell, and macrophage response to oATP in vivo and in vitro and examined intragraft inflammatory gene transcripts. Results: Kaplan-Meier survival analyses demonstrated significantly better graft survival rates in oATP and CsA groups compared to no treatment (p

Background: Complement activation in kidney transplantation is implicated in the pathogenesis of delayed graft function (DGF). This study evaluated the therapeutic efficacy of high-dose recombinant human C1 esterase inhibitor (rhC1INH) to prevent DGF in a nonhuman primate model of kidney transplantation after brain death and prolonged cold ischemia. Methods: Brain death donors underwent 20 h of conventional management. Procured kidneys were stored on ice for 44-48 h, then transplanted into ABO-compatible major histocompatibility complex-mismatched recipients. Recipients were treated with vehicle (n = 5) or rhC1INH 500 U/kg plus heparin 40 U/kg (n = 8) before reperfusion, 12 h, and 24 h posttransplant. Recipients were followed up for 120 d. Results: Of vehicle-treated recipients, 80% (4 of 5) developed DGF versus 12.5% (1 of 8) rhC1INH-treated recipients (P = 0.015). rhC1INH-treated recipients had faster creatinine recovery, superior urinary output, and reduced urinary neutrophil gelatinase-associated lipocalin and tissue inhibitor of metalloproteinases 2-insulin-like growth factor-binding protein 7 throughout the first week, indicating reduced allograft injury. Treated recipients presented lower postreperfusion plasma interleukin (IL)-6, IL-8, tumor necrosis factor-alpha, and IL-18, lower day 4 monocyte chemoattractant protein 1, and trended toward lower C5. Treated recipients exhibited less C3b/C5b-9 deposition on day 7 biopsies. rhC1INH-treated animals also trended toward prolonged mediated rejection-free survival. Conclusions: Our results recommend high-dose C1INH complement blockade in transplant recipients as an effective strategy to reduce kidney injury and inflammation, prevent DGF, delay antibody-mediated rejection development, and improve transplant outcomes.

Exposure to non-inherited maternal antigens (NIMA) during the fetal period induces lifelong split tolerance to grafts expressing these allo-antigens. In adult mice, the production of extracellular vesicles (EVs) from maternal microchimeric cells causes cross-decoration (XD) of offspring dendritic cells (DC) with NIMA and upregulation of PD-L1, contributing to NIMA tolerance. To see how this may apply to humans, we tested NIMA acquisition by fetal DCS in human cord blood. The average percentage of NIMA-XD among total DCs was 2.6% for myeloid and 4.5% for Plasmacytoid DC. These cells showed higher PD-L1 expression than their non-XD counterparts (mDC: p = .0016; pDC: p = .024). We detected CD9

Summary: Contractile to synthetic phenotypic switching of smooth muscle cells (SMCs) contributes to stenosis in vascular disease and vascular transplants. To generate more contractile SMCs, we performed a high-throughput differentiation screen using a MYH11-NLuc-tdTomato human embryonic stem cell reporter cell line. We identified RepSox as a factor that promotes differentiation of MYH11-positive cells by promoting NOTCH signaling. RepSox induces SMCs to exhibit a more contractile phenotype than SMCs generated using PDGF-BB and TGF-β1, two factors previously used for SMC differentiation but which also cause intimal hyperplasia. In addition, RepSox inhibited intimal hyperplasia caused by contractile to synthetic phenotypic switching of SMCs in a rat balloon injury model. Thus, in addition to providing more contractile SMCs that could prove useful for constructing artificial blood vessels, this study suggests a strategy for identifying drugs for inhibiting intimal hyperplasia that act by driving contractile differentiation rather than inhibiting proliferation non-specifically. : Thomson, Zhang, and colleagues report a high-throughput screen that can be used for optimization of the fully defined differentiation of contractile smooth muscle cells and identification of intimal hyperplasia inhibitors. Both in vitro and in vivo evidence revealed that RepSox is better than PDGF-BB and TGF-β1 in inducing contractile phenotype of smooth muscle cells and reducing the risk of intimal hyperplasia. Keywords: pluripotent stem cells, contractile smooth muscle cells, differentiation, maturation, RepSox, NOTCH, intima hyperplasia, MYH11-NLuc-tdTomato reporter cell line, high-throughput screen, restenosis

Normothermic ex vivo liver perfusion (NEVLP) is a novel system for organ preservation that may improve over static cold storage clinically and offers the chance for graft modification prior to transplantation. Although recent studies have shown the presence of inflammatory molecules during perfusion, none have yet shown the effects of NEVLP on liver-resident immune cell activation. We investigated the effects of NEVLP on liver-resident immune cell activation and assessed the ability of anti-inflammatory cytokines interleukin 10 (IL10) and transforming growth factor β (TGF-β) to improve organ function and reduce immune activation during perfusion. Rat livers were perfused for 4 hours at 37°C with or without the addition of 20 ng/mL of each IL10 and TGF-β (n = 7). Naive and cold storage (4 hours at 4°C) livers served as controls (n = 4). Following preservation, gene expression profiles were assessed through single-cell RNA sequencing; dendritic cell and macrophage activation was measured by flow cytometry; and cytokine production was assessed by enzyme-linked immunosorbent assay. NEVLP induced a global inflammatory gene expression signature, most notably in liver-resident macrophages and dendritic cells, which was accompanied by an increase in cell-surface levels of major histocompatibility complex (MHC) II, CD40, and CD86. Immune activation was partially ameliorated by IL10 and TGF-β treatment, but no changes were observed in inflammatory cytokine production. Overall levels of liver damage and cellular apoptosis from perfusion were low, and liver function was improved with IL10 and TGF-β treatment. This is the first study to demonstrate that liver-resident immune cells gain an activated phenotype during NEVLP on both the gene and protein level and that this activation can be reduced through therapeutic intervention with IL10 and TGF-β.

Development of a new methodology to induce immunological chimerism after allogeneic hematopoietic cell (HC) transplantation in a rhesus macaque model is described. The chimeric state was achieved using a non-myeloablative, helical tomotherapy-based total lymphoid irradiation (TomoTLI) conditioning regimen followed by donor HC infusions between 1-haplotype matched donor/recipient pairs. The technique was tested as a feasibility study in an experimental group of seven rhesus macaques that received the novel TomoTLI tolerance protocol and HC allo-transplants. Two tomotherapy protocols were compared: TomoTLI (n = 5) and TomoTLI/total-body irradiation (TBI) (n = 2). Five of seven animals developed mixed chimerism. Three of five animals given the TomoTLI protocol generated transient mixed chimerism with no graft-versus-host disease (GVHD) with survival of 33, 152 and >180 days. However, the inclusion of belatacept in addition to a single fraction of TBI resulted in total chimerism and fatal GVHD in both animals, indicating an unacceptable conditioning regimen.

Chronic hypoxia (CH)-induced pulmonary hypertension (PH) results, in part, from T helper-17 (TH17) cell-mediated perivascular inflammation. However, the antigen(s) involved is unknown. Cellular immunity to collagen type V (col V) develops after ischemia-reperfusion injury during lung transplant and is mediated by naturally occurring (n)TH17 cells. Col5a1 gene codifies for the α1-helix of col V, which is normally hidden from the immune system within type I collagen in the extracellular matrix. COL5A1 promoter analysis revealed nuclear factor of activated T cells, cytoplasmic 3 (NFATc3) binding sites. Therefore, we hypothesized that smooth muscle NFATc3 upregulates col V expression, leading to nTH17 cell-mediated autoimmunity to col V in response to CH, representing an upstream mechanism in PH development. To test our hypothesis, we measured indexes of PH in inducible smooth muscle cell (SMC)-specific NFATc3 knockout (KO) mice exposed to either CH (380 mmHg) or normoxia and compared them with wild-type (WT) mice. KO mice did not develop PH. In addition, COL5A1 was one of the 1,792 genes differentially affected by both CH and SMC NFATc3 in isolated intrapulmonary arteries, which was confirmed by RT-PCR and immunostaining. Cellular immunity to col V was determined using a trans vivo delayed-type hypersensitivity assay (Tv-DTH). Tv-DTH response was evident only when splenocytes were used from control mice exposed to CH but not from KO mice, and mediated by nTH17 cells. Our results suggest that SMC NFATc3 is important for CH-induced PH in adult mice, in part, by regulating the expression of the lung self-antigen COL5A1 protein contributing to col V-reactive nTH17-mediated inflammation and hypertension.

Background. Individuals harbor preexisting HLA−DR/DQ−restricted responses to collagen type V (ColV) mediated by Th17 cells under Treg control, both specific to peptides that bind to inherited HLA class II antigens. Yet after transplant, the donor−DR type somehow influences graft outcome. We hypothesized that, long after a lung or heart allograft, the particular HLA−DR type of the mismatched transplant donor transforms the specificity of the “anti−self” response. This could explain why, over long term, certain donor DRs could be more immunogenic than others. Methods. We analyzed 7 HLA−DR15neg patients who had received a lung allograft from a DR15+ donor. To determine the mechanism of acquired specificity in self−reactivity, we analyzed the kinetics of DR1 (host) and DR15 (donor) peptide restriction in a heart transplant model using DR−transgenic mice. Results. Beyond 1.5 years post-lung transplant, all patients tested had acquired DR15−restricted immune responses to ColV peptides. These responses were either unrestrained Th17 type (n = 4) or Th17 controlled by Treg arising early (7 y) after transplant (n = 4). Treg suppression via conventional (transforming growth factor−β [TGF−β]) and extracellular vesicle−associated (IL−35) cytokines correlated with superior outcomes. Naïve DR1 and DR15 transgenic mice had preexisting DR−restricted responses, exclusively to ColV fragments containing DR1− or DR15−binding peptides. When HLA−DR1 transgenic recipients of a HLA−DR15 heart developed ColV reactivity post-transplant, mice that acutely rejected (20–25 d) responded only to the DR1−restricted ColV peptide epitope. In animals whose grafts survived long term, we could detect acquisition of DR from the transplant donor onto the surface of recipient dendritic cells, and immune responses against a donor DR15–restricted ColV peptide. Conclusions. These results might explain how certain donor HLA−DR types redirect host immune responses to novel peptides of critical self−antigens. Unless regulated, such responses may predispose the allograft to chronic rejection.

Originally identified as lymphocyte regulation of fellow lymphocytes, our understanding of infectious tolerance has undergone significant evolutions in understanding since being proposed in the early 1970s by Gershon and Kondo and expanded upon by Herman Waldman two decades later. The evolution of our understanding of infectious tolerance has coincided with significant cellular and humoral discoveries. The early studies leading to the isolation and identification of Regulatory T cells (Tregs) and cytokines including TGFβ and IL-10 in the control of peripheral tolerance was a paradigm shift in our understanding of infectious tolerance. More recently, another potential, paradigm shift in our understanding of the “infectious” aspect of infectious tolerance was proposed, identifying extracellular vesicles (EVs) as a mechanism for propagating infectious tolerance. In this review, we will outline the history of infectious tolerance, focusing on a potential EV mechanism for infectious tolerance and a novel, EV-associated form for the cytokine IL-35, ideally suited to the task of propagating tolerance by “infecting” other lymphocytes.

Delayed graft function (DGF) in renal transplant is associated with reduced graft survival and increased immunogenicity. The complement-driven inflammatory response after brain death (BD) and posttransplant reperfusion injury play significant roles in the pathogenesis of DGF. In a nonhuman primate model, we tested complement-blockade in BD donors to prevent DGF and improve graft survival. BD donors were maintained for 20 hours; kidneys were procured and stored at 4 degrees C for 43-48 hours prior to implantation into ABO-compatible, nonsensitized, MHC-mismatched recipients. Animals were divided into 3 donor-treatment groups: G1 - vehicle, G2 - rhC1INH+heparin, and G3 - heparin. G2 donors showed significant reduction in classical complement pathway activation and decreased levels of tumor necrosis factor alpha and monocyte chemoattractant protein 1. DGF was diagnosed in 4/6 (67%) G1 recipients, 3/3 (100%) G3 recipients, and 0/6 (0%) G2 recipients (P = .008). In addition, G2 recipients showed superior renal function, reduced sC5b-9, and reduced urinary neutrophil gelatinase-associated lipocalin in the first week posttransplant. We observed no differences in incidence or severity of graft rejection between groups. Collectively, the data indicate that donor-management targeting complement activation prevents the development of DGF. Our results suggest a pivotal role for complement activation in BD-induced renal injury and postulate complement blockade as a promising strategy for the prevention of DGF after transplantation.

Background:Use of cell-based medicinal products (CBMPs) represents a state-of-the-art approach for reducing general immunosuppression in organ transplantation. We tested multiple regulatory CBMPs in kidney transplant trials to establish the safety of regulatory CBMPs when combined with reduced immunosuppressive treatment. Methods:The ONE Study consisted of seven investigator-led, single-arm trials done internationally at eight hospitals in France, Germany, Italy, the UK, and the USA (60 week follow-up). Included patients were living-donor kidney transplant recipients aged 18 years and older. The reference group trial (RGT) was a standard-of-care group given basiliximab, tapered steroids, mycophenolate mofetil, and tacrolimus. Six non-randomised phase 1/2A cell therapy group (CTG) trials were pooled and analysed, in which patients received one of six CBMPs containing regulatory T cells, dendritic cells, or macrophages; patient selection and immunosuppression mirrored the RGT, except basiliximab induction was substituted with CBMPs and mycophenolate mofetil tapering was allowed. None of the trials were randomised and none of the individuals involved were masked. The primary endpoint was biopsy-confirmed acute rejection (BCAR) within 60 weeks after transplantation; adverse event coding was centralised. The RTG and CTG trials are registered withClinicalTrials.gov,NCT01656135,NCT02252055,NCT02085629,NCT02244801,NCT02371434,NCT02129881, andNCT02091232. Findings:The seven trials took place between Dec 11, 2012, and Nov 14, 2018. Of 782 patients assessed for eligibility, 130 (17%) patients were enrolled and 104 were treated and included in the analysis. The 66 patients who were treated in the RGT were 73% male and had a median age of 47 years. The 38 patients who were treated across six CTG trials were 71% male and had a median age of 45 years. Standard-of-care immunosuppression in the recipients in the RGT resulted in a 12% BCAR rate (expected range 3·2–18·0). The overall BCAR rate for the six parallel CTG trials was 16%. 15 (40%) patients given CBMPs were successfully weaned from mycophenolate mofetil and maintained on tacrolimus monotherapy. Combined adverse event data and BCAR episodes from all six CTG trials revealed no safety concerns when compared with the RGT. Fewer episodes of infections were registered in CTG trials versus the RGT. Interpretation:Regulatory cell therapy is achievable and safe in living-donor kidney transplant recipients, and is associated with fewer infectious complications, but similar rejection rates in the first year. Therefore, immune cell therapy is a potentially useful therapeutic approach in recipients of kidney transplant to minimise the burden of general immunosuppression.

Leukocyte-associated Ig-like receptor 1 (LAIR1) is an ITIM-bearing collagen receptor expressed by leukocytes and is implicated in immune suppression. However, using a divalent soluble LAIR1/Fc recombinant protein to block interaction of cell surface LAIR1 with matrix collagen, we found that whereas Th1 responses were enhanced as predicted, Th17 responses were strongly inhibited. Indeed, LAIR1 on both T cells and monocytes was required for optimal Th17 responses to collagen type (Col)V. For pre-existing “natural” Th17 response to ColV, the LAIR1 requirement was absolute, whereas adaptive Th17 and Th1/17 immune responses in both mice and humans were profoundly reduced in the absence of LAIR1. Furthermore, the addition of C1q, a natural LAIR1 ligand, decreased Th1 responses in a dose-dependent manner, but it had no effect on Th17 responses. In IL-17–dependent murine organ transplant models of chronic rejection, LAIR1+/+ but not LAIR1−/− littermates mounted strong fibroproliferative responses. Surface LAIR1 expression was higher on human Th17 cells as compared with Th1 cells, ruling out a receptor deficiency that could account for the differences. We conclude that LAIR1 ligation by its natural ligands favors Th17 cell development, allowing for preferential activity of these cells in collagen-rich environments. The emergence of cryptic self-antigens such as the LAIR1 ligand ColV during ischemia/reperfusion injury and early acute rejection, as well as the tendency of macrophages/monocytes to accumulate in the allograft during chronic rejection, favors Th17 over Th1 development, posing a risk to long-term graft survival.

Extracellular matrix (ECM) plays an important developmental role by regulating cell behaviour through structural and biochemical stimulation. Tissue-specific ECM, attained through decellularization, has been proposed in several strategies for tissue and organ replacement. Decellularization of animal pancreata has been reported, but the same methods applied to human pancreas are less effective due to higher lipid content. Moreover, ECM-derived hydrogels can be obtained from many decellularized tissues, but methods have not been reported to obtain human pancreas-derived hydrogel. Using novel decellularization methods with human pancreas we produced an acellular, 3D biological scaffold (hP-ECM) and hydrogel (hP-HG) amenable to tissue culture, transplantation and proteomic applications. The inclusion of a homogenization step in the decellularization protocol significantly improved lipid removal and gelation capability of the resulting ECM, which was capable of gelation at 37 °C in vitro and in vivo, and is cytocompatible with a variety of cell types and islet-like tissues in vitro. Overall, this study demonstrates the characterisation of a novel protocol for the decellularization and delipidization of human pancreatic tissue for the production of acellular ECM and ECM hydrogel suitable for cell culture and transplantation applications. We also report a list of 120 proteins present within the human pancreatic matrisome.

BACKGROUND: The greatest challenge to long-term graft survival is the development of chronic lung allograft dysfunction (CLAD). Th17 responses to collagen type V (colV) predispose lung transplant patients to the severe obstructive form of CLAD, known as bronchiolitis obliterans syndrome (BOS). In a previous study cohort (n=54), pre-transplant colV responses were increased in recipients expressing HLA-DR15, consistent with the high binding avidity of colV(α1) peptides for HLA-DR15, while BOS incidence, which was known to be strongly associated with post-transplant autoimmunity to colV, was higher in patients who themselves lacked HLA-DR15, but whose lung donor expressed it. METHODS: To determine if this DR-restricted effect on BOS incidence could be validated in a larger cohort, we performed a retrospective analysis of outcomes for 351 lung transplant recipients transplanted between 1988 and 2008 at the University of Wisconsin. All subjects were followed until graft loss, death, loss to follow-up, or through 2014, with an average follow-up of 7 years. Comparisons were made between recipients who did or did not develop BOS. Grading of BOS followed the recommendations of the international society for heart and lung transplantation. RESULTS: Donor HLA-DR15 was indeed associated with increased susceptibility to severe BOS in this population. We also discovered that HLA-DR7 expression by the donor or HLA-DR17 expression by the recipient decreased susceptibility. CONCLUSION: We show in this retrospective study that specific donor HLA class II types are important in lung transplantation, as they are associated with either protection from or susceptibility to development of severe BOS.

T helper 17 (Th17)-dependent autoimmune responses can develop after heart or lung transplantation and are associated with fibro-obliterative forms of chronic rejection; however, the specific self-antigens involved are typically different from those associated with autoimmune disease. To investigate the basis of these responses, we investigated whether removal of regulatory T cells or blockade of function reveals a similar autoantigen bias. We found that Th17 cells specific for collagen type V (Col V), kα1-tubulin, and vimentin were present in healthy adult peripheral blood mononuclear cells, cord blood, and fetal thymus. Using synthetic peptides and recombinant fragments of the Col V triple helical region (α1[V]), we compared Th17 cells from healthy donors with Th17 cells from Col V-reactive heart and lung patients. Although the latter responded well to α1(V) fragments and peptides in an HLA-DR-restricted fashion, Th17 cells from healthy persons responded in an HLA-DR-restricted fashion to fragments but not to peptides. Col V, kα1-tubulin, and vimentin are preferred targets of a highly conserved, hitherto unknown, preexisting Th17 response that is MHC class II restricted. These data suggest that autoimmunity after heart and lung transplantation may result from dysregulation of an intrinsic mechanism controlling airway and vascular homeostasis.

Chronic hypoxia (CH)-induced pulmonary hypertension (PH) is a progressive and often fatal consequence of chronic lung diseases, chronic exposure to high altitude and sleep apnea. PH results from arterial remodeling, enhanced vasoreactivity, and Th17 cell-mediated perivascular inflammation. Naturally occurring “nTH17” cells, can be detected in fetal life and are in homeostatic equilibrium with natural T regulatory cells. nTH17 cells are specific for a limited range of self-antigens, including type V collagen (colV). ColV is normally hidden from the immune system within type I col in the extracellular matrix of the lungs. We have shown that type V collagen is a self-antigen uncovered by CH. On the other hand, we have shown that mice lacking nuclear factor of activated T cells isoform c3 (NFATc3) globally and specifically in smooth muscle (SMC-NFATc3 KO) are protected from CH-induced PH. No differences in cardiac function were detected between SMC-NFATc3 KO and control mice using a Vevo LAZR-X ultrasound system before CH. In silico analysis of Col5A1 promoter shows NFATc3 binding sites. Therefore, we hypothesized that smooth muscle NFATc3 upregulates Col5A1 expression contributing to the development of acute autoreactivity to collagen V in response to CH, representing an upstream mechanism in PH development. Col5A1 was one of the 1792 genes differentially affected by both CH and smooth muscle NFATc3 in isolated pulmonary arteries (RNA Illumina Sequencing). Col5A1 mRNA increased significantly after 5 days of CH in control mice (fold change from normoxia±SD=2.2±0.8, n=5) vs. SMC-NFATc3 KO (0.8±0.3, n=3, p-4 ”] ± SD of naïve recipient mice injected with cells from CH control mice 25.6±4.2, n=8 vs. CH SMC-NFATc3 KO mice 5.3±0.5, n=7; p Our results suggest that smooth muscle NFATc3 is important for CH-induced PH in adult mice, in part, by regulating the expression of the lung self-antigen ColV contributing to pulmonary vascular inflammation.

Exosomes are a potent means for intercellular communication. However, exosomes have received intensive research focus in immunobiology only relatively recently. Because they transport proteins, lipids and genetic material between cells, they are especially suited to amplify their parental cell's message and overcome the physical constraints of cell-to-cell contact, that is exosome release gives cells the ability to alter distant, non-contiguous cells. As progress is made in this field, it has become increasingly obvious that exosomes are involved in most biological processes. In the immune system, exosomes are fundamental tools used by every immune cell type to fulfil its function and promote inflammation or tolerance. In this review, we first summarize key aspects of immune cell-specific exosomes and their functions. Then, we describe how exosomes have been shown to be indispensable orchestrators of the immune response in two immunological scenarios, namely transplant rejection or tolerance, and tumour evasion or initiation of anti-tumour immune responses.

Background. The ideal minimizing strategy for maintenance immunosuppression in HLA-matched kidney transplant recipients (KTR) is unknown. We hypothesized that mycophenolate (MPA) monotherapy is a safe and effective approach for maintenance therapy in this group of KTR. Methods. Data were abstracted for 6-antigen HLA-matched KTR between 1994 and 2013. Twenty recipients receiving MPA monotherapy secondary to infection, cancer, calcineurin inhibitor (CNI) side effects, or immunosuppression minimization strategies were evaluated in this case series. Results. MPA monotherapy had a low incidence of death-censored graft failure (3.19/100 person-y), rejection (0/100 person-y), hospitalization (1.62/100 person-y), malignancy (3.61/100 person-y), and infection (1.75/100 person-y). Further, 12-month mean or median serum creatinine (1.29 mg/dL), estimated glomerular filtration rate (64.3 mL/min/1.73 m2), urine protein creatinine ratio (143.2 mg/g), hemoglobin (13.9 g/dL), platelets (237.8 K/uL), and white blood cell count (9.04 K/uL) were favorable. There was a successful conversion rate of 90% (18 of 20) with 2 patients converting back to CNI-based regimens secondary to recurrence of membranous nephropathy and post-transplant lymphoproliferative disorder. Conclusions. Our findings indicate that MPA monotherapy may be a promising immunosuppression minimization strategy for HLA-matched KTR.

Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) exhibit a fetal phenotype that limits in vitro and therapeutic applications. Strategies to promote cardiomyocyte maturation have focused interventions on differentiated hPSC-CMs, but this study tests priming of early cardiac progenitor cells (CPCs) with polyinosinic-polycytidylic acid (pIC) to accelerate cardiomyocyte maturation. CPCs were differentiated from hPSCs using a monolayer differentiation protocol with defined small molecule Wnt temporal modulation, and pIC was added during the formation of early CPCs. pIC priming did not alter the expression of cell surface markers for CPCs (>80% KDR+/PDGFRα+), expression of common cardiac transcription factors, or final purity of differentiated hPSC-CMs (∼90%). However, CPC differentiation in basal medium revealed that pIC priming resulted in hPSC-CMs with enhanced maturity manifested by increased cell size, greater contractility, faster electrical upstrokes, increased oxidative metabolism, and more mature sarcomeric structure and composition. To investigate the mechanisms of CPC priming, RNAseq revealed that cardiac progenitor-stage pIC modulated early Notch signaling and cardiomyogenic transcriptional programs. Chromatin immunoprecipitation of CPCs showed that pIC treatment increased deposition of the H3K9ac activating epigenetic mark at core promoters of cardiac myofilament genes and the Notch ligand, JAG1. Inhibition of Notch signaling blocked the effects of pIC on differentiation and cardiomyocyte maturation. Furthermore, primed CPCs showed more robust formation of hPSC-CMs grafts when transplanted to the NSGW mouse kidney capsule. Overall, epigenetic modulation of CPCs with pIC accelerates cardiomyocyte maturation enabling basic research applications and potential therapeutic uses. Stem Cells 2019;37:910–923

ObjectivesEmerging evidence has shown a role for tumor antigen-specific regulation in cancer. Identifying individuals with pre-existing regulatory responses may be key to understand those who are more likely to respond to Programmed Death-1 (PD-1) or PD-1 Ligand 1 (PD-L1) checkpoint blockade. We hypothesized that a functional assay could identify the role of PD-1/PD-L1 interactions on tumor-specific immune cells in the peripheral blood in patients with advanced non-small-cell lung cancer (NSCLC).MethodsWe performed the trans vivo delayed-type hypersensitivity assay to identify the role of PD-1/PD-L1-mediated tumor-specific immune regulation in ten patients with advanced NSCLC.ResultsThe majority of patients had PD-1-mediated anergic immune responses towards their tumor antigens. Eight out of nine of these patients did not respond to their own tumor antigens but responded in the presence of anti-PD-1 antibody (‘PD-1 anergy’ phenotype). A minority (3/9) also had ‘active’ PD-1-mediated immune suppressive regulatory responses. Our results suggest that PD-1-anergy is a common feature of NSCLC immune responses, whereas PD-1-mediated immune suppression is present only in a minority of patients. The latter was associated with poor clinical outcomes in our sample.ConclusionsOverall, our results indicate that bystander suppression or the ‘anergy-only’ phenomenon may be novel biomarkers in NSCLC and suggest prediction value based on these phenotypes.

Hypoxic pulmonary hypertension (PH) is a progressive and often fatal consequence of chronic hypoxia (CH) exposure (chronic lung diseases, high altitude and sleep apnea). We recently demonstrated that T helper 17 (T H 17) cells are localized in the perivascular region of pulmonary arteries and contribute to CH-induced PH, at least in part, by causing pulmonary arterial (PA) remodeling. Naturally occurring “nT H 17” cells, can be detected as early as week 17 of fetal life in humans, and are in homeostatic equilibrium with natural T regulatory (nTregs) cells. nT H 17 cells are specific for a limited range of self-antigens, including type V collagen (colV). ColV is normally sequestered within type I col in the extracellular matrix of the lungs hidden from the immune system. Activation of matrix metalloproteinase-2, which is induced by CH, can expose colV. Therefore, we tested the hypothesis that ColV is a lung-associated self-antigen uncovered by CH. We found colV immunoreactivity only in lungs from mice exposed to hypobaric CH for 5 days vs. controls and colV mRNA expression is significantly increased in isolated PA (log fold change mean±SEM, normoxia= 0.22±0.08 vs. CH=0.48±0.02, n=3, p=0.02) suggesting that CH uncovers hidden colV. We then, determined cellular autoimmunity to colV using a trans-vivo delayed-type hypersensitivity assay (TV-DTH). Splenocytes obtained from 5 days CH-exposed mice displayed a TV-DTH colV response significantly higher than that from normoxic mice (footpad swelling [10 -4 ”] normoxia=11.8±1.1 vs. CH= 35.6±3.0, n=14, p=0.0001), while there was minimal response to col I. To determine that reactivity to colV contributes to CH-induced PH, we induced peripheral tolerance to colV by administering 25 μl of 0.08 μg/μl of bovine colV (Sigma) in PBS in each nare or PBS every two days for two weeks prior and during CH (21 days). Right ventricular systolic pressure (RVSP) was higher in vehicle-treated compared to colV-treated mice (mmHg CH vehicle= 32.4±0.3 vs. CH colV= 27.8±0.9, n=4, p=0.0085) suggesting that oral tolerance to col V attenuated PH. These results are ground-breaking suggesting that acute autoreactivity to colV develops after exposure to hypoxia. Next, we will determine whether a disrupted balance of nTh17/nTreg mediates this response.

Leukocyte-associated Ig-like receptor 1 (LAIR1) is an ITIM-bearing collagen receptor expressed by leukocytes and is implicated in immune suppression. However, using a divalent soluble LAIR1/Fc recombinant protein to block interaction of cell surface LAIR1 with matrix collagen, we found that whereas T(h)1 responses were enhanced as predicted, T(h)17 responses were strongly inhibited. Indeed, LAIR1 on both T cells and monocytes was required for optimal T(h)17 responses to collagen type (Col)V. For pre-existing “natural” T(h)17 response to ColV, the LAIR1 requirement was absolute, whereas adaptive T(h)17 and T(h)1/17 immune responses in both mice and humans were profoundly reduced in the absence of LAIR1. Furthermore, the addition of C1q, a natural LAIR1 ligand, decreased T(h)1 responses in a dose-dependent manner, but it had no effect on T(h)17 responses. In IL-17-dependent murine organ transplant models of chronic rejection, LAIR1(+/+) but not LAIR1(−/−) littermates mounted strong fibroproliferative responses. Surface LAIR1 expression was higher on human T(h)17 cells as compared with T(h)1 cells, ruling out a receptor deficiency that could account for the differences. We conclude that LAIR1 ligation by its natural ligands favors T(h)17 cell development, allowing for preferential activity of these cells in collagen-rich environments. The emergence of cryptic self-antigens such as the LAIR1 ligand ColV during ischemia/reperfusion injury and early acute rejection, as well as the tendency of macrophages/monocytes to accumulate in the allograft during chronic rejection, favors T(h)17 over T(h)1 development, posing a risk to long-term graft survival.

Biomarkers of transplant tolerance would enhance the safety and feasibility of clinical tolerance trials and potentially facilitate management of patients receiving immunosuppression. To this end, we examined blood from spontaneously tolerant renal transplant recipients and patients enrolled in two interventional tolerance trials using flow cytometry and gene expression profiling. Using a previously reported tolerant cohort as well as newly identified tolerant patients, we confirmed our previous finding that tolerance was associated with increased expression of B cell-associated genes relative to immunosuppressed patients. This was not accounted for merely by an increase in total B cell numbers, but was associated with the increased frequencies of transitional and naïve B cells. Moreover, serial measurements of gene expression demonstrated that this pattern persisted over several years, although patients receiving immunosuppression also displayed an increase in the two most dominant tolerance-related B cell genes, IGKV1D-13 and IGLL-1, over time. Importantly, patients rendered tolerant via induction of transient mixed chimerism, and those weaned to minimal immunosuppression, showed similar increases in IGKV1D-13 as did spontaneously tolerant individuals. Collectively, these findings support the notion that alterations in B cells may be a common theme for tolerant kidney transplant recipients, and that it is a useful monitoring tool in prospective trials.

Epstein-Barr virus (EBV) infection transforms B cells in vitro and is associated with human B cell lymphomas. The major EBV oncoprotein, latent membrane protein 1 (LMP1), mimics constitutively active CD40 and is essential for outgrowth of EBV-transformed B cells in vitro; however, EBV-positive diffuse large B cell lymphomas and Burkitt lymphomas often express little or no LMP1. Thus, EBV may contribute to the development and maintenance of human lymphomas even in the absence of LMP1. Here, we found that i.p. injection of human cord blood mononuclear cells infected with a LMP1-deficient EBV into immunodeficient mice induces B cell lymphomas. In this model, lymphoma development required the presence of CD4+ T cells in cord blood and was inhibited by CD40-blocking Abs. In contrast, LMP1-deficient EBV established persistent latency but did not induce lymphomas when directly injected into mice engrafted with human fetal CD34+ cells and human thymus. WT EBV induced lymphomas in both mouse models and did not require coinjected T cells in the cord blood model. Together, these results demonstrate that LMP1 is not essential for EBV-induced lymphomas in vivo and suggest that T cells supply signals that substitute for LMP1 in EBV-positive B cell lymphomagenesis.

Organ transplantation is the only viable treatment for several end-stage organ failures. However, chronic rejection prevents long-term graft survival. Traditionally, this rejection was attributed to the development of alloimmunity in transplant patients. However, recent evidence suggests that autoimmunity plays a larger role in chronic rejection of certain organ transplants, than alloimmunity. In this review, we will focus on the history of autoimmunity in solid organ transplantation and look at the collagen type V, Kα1 tubulin, vimentin, cardiac myosin, and heat shock proteins as classical examples of autoantigens in organ transplantation. We will also look at some of the recent reports looking at the mechanisms of autoimmunity and try to provide answers to some of the age-old questions in autoimmunity.

Summary Here, we describe the NeoThy humanized mouse model created using non-fetal human tissue sources, cryopreserved neonatal thymus and umbilical cord blood hematopoietic stem cells (HSCs). Conventional humanized mouse models are made by engrafting human fetal thymus and HSCs into immunocompromised mice. These mice harbor functional human T cells that have matured in the presence of human self-peptides and human leukocyte antigen molecules. Neonatal thymus tissue is more abundant and developmentally mature and allows for creation of up to ∼50-fold more mice per donor compared with fetal tissue models. The NeoThy has equivalent frequencies of engrafted human immune cells compared with fetal tissue humanized mice and exhibits T cell function in assays of ex vivo cell proliferation, interferon γ secretion, and in vivo graft infiltration. The NeoThy model may provide significant advantages for induced pluripotent stem cell immunogenicity studies, while bypassing the requirement for fetal tissue., Highlights • Neonatal tissue is a viable alternative to fetal for mouse humanization • Over 1,000 NeoThy mice can be made from one neonatal thymus donor • The NeoThy enables robust pre-clinical immunogenicity studies of iPSC therapies, Corresponding author William Burlingham and colleagues created a humanized mouse model called the NeoThy. The NeoThy uses human neonatal, rather than fetal, tissue sources for generating a human immune system within immunocompromised mouse hosts. NeoThy mice are an attractive alternative to conventional humanized mouse models, as they enable robust and reproducible iPSC immunogenicity experiments in vivo.

Significance Generating fully functional arterial endothelial cells is a critical problem for vascular development and disease research. Currently, the arterial endothelial cells derived from human pluripotent stem cells lack the range of arterial-specific functions in vitro and the protective function for ischemic tissues in vivo. Here, we combine single-cell RNA sequencing and CRISPR-Cas9 technology to identify pathways for regulating arterial endothelial cell differentiation. We then manipulate these pathways and generate arterial endothelial cells that demonstrate unprecedented arterial-specific functions as well as improve survival of myocardial infarction. These findings facilitate the understanding of vascular development and disease and provide a source of cells that have broad applications for vascular disease modeling and regenerative medicine.

PURPOSE OF REVIEW The passenger leukocyte hypothesis predicts that after transplantation, donor antigen-presenting cells (APCs) from the graft present donor MHC molecules to directly alloreactive T cells in lymphoid organs. However, in certain transplantation models, recent evidence contradicts this long-standing concept. New findings demonstrate that host, instead of donor, APCs play a prominent role in allosensitization against donor MHC molecules via the semidirect pathway. A similar mechanism operates in development of T-cell split tolerance to noninherited maternal antigens. RECENT FINDINGS Following fully mismatch skin or heart transplantation in mice, no or extremely few donor migrating APCs (i.e. conventional dendritic cells) are detected in the draining lymphoid organs. Instead, recipient dendritic cells that have captured donor extracellular vesicles (i.e. exosomes) carrying donor MHC molecules and APC costimulatory signals present donor MHC molecules to directly alloreactive T cells. This semidirect pathway can also give rise to a form of 'split' tolerance during chronic alloantigen exposure, as indirectly alloreactive T helper cells and directly alloreactive T-cell effectors are differentially impacted by host dendritic cells 'cross-dressed' with extracellular vesicles/exosomes derived from maternal microchimerism. SUMMARY Acquisition by recipient APCs of donor exosomes (and likely other extracellular vesicles) released by passenger leukocytes or the graft explains the potent T-cell allosensitization against donor MHC molecules, in the absence or presence of few passenger leukocytes in lymphoid organs. It also provides the basic mechanism and in-vivo relevance of the elusive semidirect pathway. Its degree of coordination with the allopeptide - specific, indirect pathway of T-cell help may determine whether semidirect allopresentation results in a sustained, effective, acute rejection response, or rather, in abortive acute rejection and 'split' tolerance.

Maternal microchimerism (MMc) has been associated with development of allospecific transplant tolerance, antitumor immunity, and cross-generational reproductive fitness, but its mode of action is unknown. We found in a murine model that MMc caused exposure to the noninherited maternal antigens in all offspring, but in some, MMc magnitude was enough to cause membrane alloantigen acquisition (mAAQ; “cross-dressing”) of host dendritic cells (DCs). Extracellular vesicle (EV)-enriched serum fractions from mAAQ+, but not from non-mAAQ, mice reproduced the DC cross-dressing phenomenon in vitro. In vivo, mAAQ was associated with increased expression of immune modulators PD-L1 (programmed death-ligand 1) and CD86 by myeloid DCs (mDCs) and decreased presentation of allopeptide+self-MHC complexes, along with increased PD-L1, on plasmacytoid DCs (pDCs). Remarkably, both serum EV-enriched fractions and membrane microdomains containing the acquired MHC alloantigens included CD86, but completely excluded PD-L1. In contrast, EV-enriched fractions and microdomains containing allopeptide+self-MHC did not exclude PD-L1. Adoptive transfer of allospecific transgenic CD4 T cells revealed a “split tolerance” status in mAAQ+ mice: T cells recognizing intact acquired MHC alloantigens proliferated, whereas those responding to allopeptide+self-MHC did not. Using isolated pDCs and mDCs for in vitro culture with allopeptide+self-MHC–specific CD4 T cells, we could replicate their normal activation in non-mAAQ mice, and PD-L1–dependent anergy in mAAQ+ hosts. We propose that EVs provide a physiologic link between microchimerism and split tolerance, with implications for tumor immunity, transplantation, autoimmunity, and reproductive success.

It has become increasingly clear that the immune system of viviparous mammals is much more in the business of acquiring tolerance to non-self antigens, than it is in rejecting cells that express them (for a recent review, highlighting the role of Treg cells, see ref. (1) ). It is also clear that both self-tolerance, and acquired tolerance to non-self is a dynamic process, with a natural ebb and flow. As has been often said of an effective team defense in sports, tolerance will "bend but does not break." How microchimerism, defined as the presence of extremely rare [1/10(4)-1/10(6)] cells of a genetically different individual, can induce either new immunogenetic pressures that push self-tolerance to the breaking point, or alternatively, provide relief from pre-existing immunogenetic risk, preventing development of autoimmune disease, remains a mystery. Indeed, the inability to directly correlate DNA-level microchimerism detected in blood samples by qPCR, with naturally occurring regulation to minor H and MHC alloantigens expressed by the rare cells themselves, has been frustrating to researchers in this field. (2) [Haynes, W.J. et al, this issue] However, recent developments in the areas of transplantation and reproductive immunology offer clues to how the effects of microchimerism can be amplified, and how a disproportionate immune impact might occur from a very limited cell source.

While transplantation research continues to focus on the development of Treg-sparing and plasma cell/B-cell-targeting strategies to improve long-term outcomes, little attention has been paid to the emerging role of the TH17 cell in mediating chronic rejection. This review aimed to summarize the recent emergence of the TH17 and the newly described TH1/17 cell as major players in transplantation and to suggest ways of improving graft outcome through TH17-targeted immunosuppression.

Studies of the effect of minor H antigen mismatching on the outcome of renal transplantation are scarce and concern mainly single center studies. The International Histocompatibility and Immunogenetics Workshops (IHIW) provide a collaborative platform to execute crucial large studies. In collaboration with 16 laboratories of the IHIW, the role of 15 autosomal, 10 Y-chromosome encoded minor H antigens and 3 CD31 polymorphisms, was investigated in relation to the incidence of renal graft rejection and graft loss in 444 human leukocyte antigens (HLA)-identical sibling renal transplantations. Recipient and donor DNA samples were genotyped for the minor H antigens HA-1, HA-2, HA-3, HA-8, HB-1, ACC-1, ACC-2, SP110, PANE1, UGT2B17, C19Orf48, LB-ECGF-1, CTSH, LRH-1, LB-ADIR and HY. The correlation between minor H antigen mismatch and the primary outcome graft rejection or graft loss was statistically analyzed. The incidence of rejection was very low and no correlation was observed between one or more minor H antigen mismatch(es) and a rejection episode (n = 36), of which only eight resulted in graft loss. In summary, in our study cohort of 444 renal transplants, mismatching for neither autosomal nor HY minor H antigens correlate with rejection episodes or with graft loss.

Chronic graft-versus-host disease (cGVHD) is a significant roadblock to long-term hematopoietic stem cell (HSC) transplantation success. Effective treatments for cGVHD have been difficult to develop, in part because of a paucity of animal models that recapitulate the multiorgan pathologies observed in clinical cGVHD. Here we present an analysis of the pathology that occurs in immunodeficient mice engrafted with human fetal HSCs and implanted with fragments of human fetal thymus and liver. Starting at time points generally later than 100 days post-transplantation, the mice developed signs of illness, including multiorgan cellular infiltrates containing human T cells, B cells, and macrophages; fibrosis in sites such as lungs and liver; and thickened skin with alopecia. Experimental manipulations that delayed or reduced the efficiency of the HSC engraftment did not affect the timing or progression of disease manifestations, suggesting that pathology in this model is driven more by factors associated with the engrafted human thymic organoid. Disease progression was typically accompanied by extensive fibrosis and degradation of the thymic organoid, and there was an inverse correlation of disease severity with the frequency of FoxP3+ thymocytes. Hence, the human thymic tissue may contribute T cells with pathogenic potential, but the generation of regulatory T cells in the thymic organoid may help to control these cells before pathology resembling cGVHD eventually develops. This model thus provides a new system to investigate disease pathophysiology relating to human thymic events and to evaluate treatment strategies to combat multiorgan fibrotic pathology produced by human immune cells.

Obliterative bronchiolitis (OB), a fibrotic airway lesion, is the leading cause of death after lung transplantation. Type V collagen [col(V)] overexpression and IL-17-mediated anti-col(V) immunity are key contributors to OB pathogenesis. Here, we report a previously undefined role of IL-17 in inducing col(V) overexpression, leading to epithelial mesenchymal transition (EMT) and subsequent OB. We observed IL-17-mediated induction of col(V) α1 chains [α1 (V)] in normal airway epithelial cells in vitro and detected α1 (V)-specific antibodies in bronchoalveolar lavage fluid of lung transplant patients. Overexpression of IL-17 and col(V) was detected in OB lesions in patient lung biopsies and in a murine OB model. IL-17 is shown to induce EMT, TGF-β mRNA expression, and SMAD3 activation, whereas downregulating SMAD7 expression in vitro. Pharmacological inhibition of TGF-βRI tyrosine kinase, p38 MAPK, or focal adhesion kinase prevented col(V) overexpression and EMT. In murine orthotopic lung transplants, neutralizing IL-17 significantly decreased TGF-β mRNA and protein expression and prevented epithelial repair/OB. Our findings highlight a feed-forward loop between IL-17 and TGF-β, leading to induction of col(V) and associated epithelial repair, thus providing one possible link between autoimmunity and OB after lung transplantation.

Understanding the evolution of the direct and indirect pathways of allorecognition following tissue transplantation is essential in the design of tolerance‐promoting protocols. On the basis that donor bone marrow–derived antigen‐presenting cells are eliminated within days of transplantation, it has been argued that the indirect response represents the major threat to long‐term transplant survival, and is consequently the key target for regulation. However, the detection of MHC transfer between cells, and particularly the capture of MHC:peptide complexes by dendritic cells (DCs), led us to propose a third, semidirect, pathway of MHC allorecognition. Persistence of this pathway would lead to sustained activation of direct‐pathway T cells, arguably persisting for the life of the transplant. In this study, we focused on the contribution of acquired MHC‐class I on recipient DCs during the life span of a skin graft. We observed that MHC‐class I acquisition by recipient DCs occurs for at least 1 month following transplantation and may be the main source of alloantigen that drives CD8+ cytotoxic T cell responses. In addition, acquired MHC‐class I:peptide complexes stimulate T cell responses in vivo, further emphasizing the need to regulate both pathways to induce indefinite survival of the graft., Using a skin transplant model, Smyth et al observe that transference of MHC:peptide complexes between donor graft tissue and recipient dendritic cells occurs throughout the life span of the graft and contributes to anti‐graft CD8 T cell responses. See the editorial from Burlingham on page 5.

Steady state alveolar macrophages (AM) are long-lived lung-resident macrophages with sentinel function. Evidence suggests that AM precursors originate during embryogenesis and populate lungs without replenishment by circulating leukocytes. However, their presence and persistence are unclear following human lung transplantation (LTx). Our goal was to examine donor AM longevity and evaluate whether AM of recipient origin seeds the transplanted lungs. Origin of AM was accessed using donor-recipient HLA mismatches. We demonstrate that 94–100% of AM present in bronchoalveolar lavage (BAL) were donor derived and importantly AM of recipient origin were not detected. Further, analysis of BAL cells up to 3.5 yrs post-LTx revealed that majority of AM (>87%) was donor derived. Elicitation of de novo donor specific antibody (DSA) is a major post-LTx complication and a risk factor for development of chronic rejection. The donor AM responded to anti-HLA framework Ab with secretion of inflammatory cytokines. Further, in an experimental murine model, we demonstrate that adoptive transfer of allogeneic AM stimulated humoral and cellular immune responses to alloantigen and lung-associated self-antigens and led to bronchiolar obstruction. Therefore, donor derived AM play an essential role in the DSA induced inflammatory cascade leading to obliterative airway disease of the transplanted lungs.

Cell based therapies have been studied extensively in the context of transplantation tolerance induction. The most successful protocols have relied on transfusion of bone marrow prior to the transplantation of a renal allograft. However, it is not clear that stem cells found in bone marrow are required in order to render a transplant candidate immunologically tolerant. Accordingly, mesenchymal stem cells, regulatory myeloid cells, T regulatory cells, and other cell types, are being tested as possible routes to tolerance induction, in the absence of donor derived stem cells. Early data with each of these cell types have been encouraging. However, the induction regimen capable of achieving consistent tolerance, whilst avoiding unwanted sided effects, and which is scalable to the human patient, has yet to be identified. Here we present the status of investigations of various tolerogenic cell types and the mechanistic rationale for their use in in tolerance induction protocols.

Conventional wisdom argues against inbreeding, to maintain hybrid vigor and increase MHC diversity in response to pathogens. A recent report from the laboratory of Sing-Sing Way uses a mouse model to test a hypothesis put forward by Ray D. Owen more than 60 years ago: that a certain amount of inbreeding is a good thing. Owen proposed that antigens not inherited from the mother (noninherited maternal antigens), when replicated on the mate of the daughter, could protect the latter's developing child from fetal wastage due to immune attack during her pregnancy. Kinder et al use elegant mouse breeding models and MHC class II peptide tetramers to show that Owen's hypothesis, based only on humoral (anti-Rh IgG) data and a small sample size, was indeed correct. The mediators of this cross-generational protection turn out to be a special kind of Foxp3+ T regulatory cell, the development of which requires the persistence of maternal microchimerism into adulthood. The implications of this discovery for the role of microchimerism in tolerance to transplants are discussed.

Objective— Apoptosis of smooth muscle cells (SMCs) is a prominent pathological characteristic of abdominal aortic aneurysm (AAA). We have previously shown that SMC apoptosis stimulates proinflammatory signaling in a mouse model of AAA. Here, we test whether protein kinase C-δ (PKCδ), an apoptotic mediator, participates in the pathogenesis of AAA by regulating apoptosis and proinflammatory signals. Methods and Results— Mouse experimental AAA is induced by perivascular administration of CaCl 2 . Mice deficient in PKCδ exhibit a profound reduction in aneurysmal expansion, SMC apoptosis, and transmural inflammation as compared with wild-type littermates. Delivery of PKCδ to the aortic wall of PKCδ –/– mice restores aneurysm, whereas overexpression of a dominant negative PKCδ mutant in the aorta of wild-type mice attenuates aneurysm. In vitro, PKCδ –/– aortic SMCs exhibit significantly impaired monocyte chemoattractant protein-1 production. Ectopic administration of recombinant monocyte chemoattractant protein-1 to the arterial wall of PKCδ –/– mice restores inflammatory response and aneurysm development. Conclusion— PKCδ is an important signaling mediator for SMC apoptosis and inflammation in a mouse model of AAA. By stimulating monocyte chemoattractant protein-1 expression in aortic SMCs, upregulated PKCδ exacerbates the inflammatory process, in turn perpetuating elastin degradation and aneurysmal dilatation. Inhibition of PKCδ may serve as a potential therapeutic strategy for AAA.