Publications by authors named "Béatrice Charreau"

61 Publications

Cellular and Molecular Crosstalk of Graft Endothelial Cells During AMR: Effector Functions and Mechanisms.

Transplantation 2021 Mar 8. Epub 2021 Mar 8.

CHU Nantes, Université de Nantes, Inserm, Centre de Recherche en Transplantation et en Immunologie, UMR 1064, ITUN, F-44000 Nantes, France.

Graft endothelial cell (EC) injury is central to the pathogenesis of antibody-mediated rejection (AMR). The ability of donor-specific antibodies (DSA) to bind C1q and activate the classical complement pathway is an efficient predictor of graft rejection highlighting complement-dependent cytotoxicity (CDC) as a key process operating during AMR. In the past 5 years, clinical studies further established the cellular and molecular signatures of AMR revealing the key contribution of other, IgG-dependent and -independent, effector mechanisms mediated by infiltrating NK cells and macrophages. Beyond binding to alloantigens, DSA IgG can activate NK cells and mediate antibody-dependent cell cytotoxicity (ADCC) through interacting with Fcγ receptors (FcγRs) such as FcγRIIIa (CD16a). FcRn, a nonconventional FcγR that allows IgG recycling, is highly expressed on ECs and may contribute to the long-term persistence of DSA in blood. Activation of NK cells and macrophages results in the production of pro-inflammatory cytokines such as TNF and IFNγ that induce transient and reversible changes in the EC phenotype and functions promoting coagulation, inflammation, vascular permeability, leukocyte trafficking. MHC class I mismatch between transplant donor and recipient can create a situation of "missing self" allowing NK cells to kill graft ECs. Depending on the microenvironment, cellular proximity with ECs may participate in macrophage polarization toward an M1 proinflammatory or an M2 phenotype favoring inflammation or vascular repair. Monocytes/macrophages participate in the loss of endothelial specificity in the process of endothelial-to-mesenchymal transition (EndMT) involved in renal and cardiac fibrosis and AMR and may differentiate into ECs enabling vessel and graft (re)-endothelialization.
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http://dx.doi.org/10.1097/TP.0000000000003741DOI Listing
March 2021

Missing Self-Induced Activation of NK Cells Combines with Non-Complement-Fixing Donor-Specific Antibodies to Accelerate Kidney Transplant Loss in Chronic Antibody-Mediated Rejection.

J Am Soc Nephrol 2021 Feb 25;32(2):479-494. Epub 2020 Nov 25.

International Center of Infectiology research (CIRI), French Institute of Health and Medical Research (INSERM) Unit 1111, Claude Bernard University Lyon I, National Center for Scientific Research (CNRS) Mixed University Unit (UMR) 5308, Ecole Normale Supérieure de Lyon, University of Lyon, Lyon, France

Background: Binding of donor-specific antibodies (DSAs) to kidney allograft endothelial cells that does not activate the classic complement cascade can trigger the recruitment of innate immune effectors, including NK cells. Activated NK cells contribute to microvascular inflammation leading to chronic antibody-mediated rejection (AMR). Recipient NK cells can also trigger antibody-independent microvascular inflammation by sensing the absence of self HLA class I molecules ("missing self") on allograft endothelial cells. This translational study investigated whether the condition of missing self amplifies DSA-dependent NK cell activation to worsen chronic AMR.

Methods And Results: Among 1682 kidney transplant recipients who underwent an allograft biopsy at Lyon University Hospital between 2004 and 2017, 135 fulfilled the diagnostic criteria for AMR and were enrolled in the study. Patients with complement-fixing DSAs identified by a positive C3d binding assay (=73, 54%) had a higher risk of transplant failure (=0.002). Among the remaining patients with complement-independent chronic AMR (=62, 46%), those in whom missing self was identified through donor and recipient genotyping exhibited worse allograft survival (=0.02). In multivariable analysis, only proteinuria (HR: 7.24; =0.01) and the presence of missing self (HR: 3.57; =0.04) were independent predictors for transplant failure following diagnosis of chronic AMR. Cocultures of human NK cells and endothelial cells confirmed that addition of missing self to DSA-induced NK cell activation increased endothelial damage.

Conclusions: The assessment of missing self at the time of diagnosis of chronic AMR identifies patients at higher risk for kidney transplant failure.
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http://dx.doi.org/10.1681/ASN.2020040433DOI Listing
February 2021

Efficient Expansion of Human Granzyme B-Expressing B Cells with Potent Regulatory Properties.

J Immunol 2020 11 18;205(9):2391-2401. Epub 2020 Sep 18.

CHU Nantes, Université de Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, F-44000 Nantes, France;

Granzyme B-expressing B cells have been shown to be an important regulatory B cell subset in humans. However, it is unclear which subpopulations of B cells express GZMB under normal conditions and which protocols effectively induce ex vivo expansion of GZMB B cells. We found that in the peripheral blood of normal individuals, plasmablasts were the major B cell subpopulation that expressed GZMB. However, when using an in vitro plasmablast differentiation protocol, we obtained only 2% GZMB B cells. Nevertheless, using an expansion mixture containing IL-21, anti-BCR, CpG oligodeoxynucleotide, CD40L, and IL-2, we were able to obtain more than 90% GZMB B cells after 3 d culture. GZMB B cells obtained through this protocol suppressed the proliferation of autologous and allogenic CD4CD25 effector T cells. The suppressive effect of GZMB B cells was partially GZMB dependent and totally contact dependent but was not associated with an increase in effector T cell apoptosis or uptake of GZMB by effector T cells. Interestingly, we showed that GZMB produced by B cells promoted GZMB B cell proliferation in ERK1/2-dependent manner, facilitating GZMB B cell expansion. However, GZMB B cells tended to undergo apoptosis after prolonged stimulation, which may be considered a negative feedback mechanism to limit their uncontrolled expansion. Finally, we found that expanded GZMB B cells exhibited a regulatory phenotype and were enriched in CD307b, CD258CD72, and CD21loPD-1 B cell subpopulations. Our study, to our knowledge, provides new insight into biology of GZMB B cells and an efficient method to expand GZMB B cells for future cell therapy applications.
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http://dx.doi.org/10.4049/jimmunol.2000335DOI Listing
November 2020

Targeting MHC Regulation Using Polycyclic Polyprenylated Acylphloroglucinols Isolated from .

Biomolecules 2020 09 2;10(9). Epub 2020 Sep 2.

Université de Nantes, CHU Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, F-44000 Nantes, France.

Modulation of major histocompatibility complex (MHC) expression using drugs has been proposed to control immunity. Phytochemical investigations on species have allowed the isolation of bioactive compounds such as polycyclic polyprenylated acylphloroglucinols (PPAPs). PPAPs such as guttiferone J (), display anti-inflammatory and immunoregulatory activities while garcinol () is a histone acetyltransferases (HAT) p300 inhibitor. This study reports on the isolation, identification and biological characterization of two other PPAPs, i.e., xanthochymol () and guttiferone F () from , sharing structural analogy with guttiferone J () and garcinol (). We show that PPAPs - efficiently downregulated the expression of several MHC molecules (HLA-class I, -class II, MICA/B and HLA-E) at the surface of human primary endothelial cells upon inflammation. Mechanistically, PPAPs - reduce MHC proteins by decreasing the expression and phosphorylation of the transcription factor STAT1 involved in MHC upregulation mediated by IFN-γ. Loss of STAT1 activity results from inhibition of HAT CBP/p300 activity reflected by a hypoacetylation state. The binding interactions to p300 were confirmed through molecular docking. Loss of STAT1 impairs the expression of CIITA and GATA2 but also TAP1 and Tapasin required for peptide loading and transport of MHC. Overall, we identified new PPAPs issued from with potential immunoregulatory properties.
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http://dx.doi.org/10.3390/biom10091266DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7563419PMC
September 2020

Missing self triggers NK cell-mediated chronic vascular rejection of solid organ transplants.

Nat Commun 2019 11 25;10(1):5350. Epub 2019 Nov 25.

CIRI, INSERM U1111, Université Claude Bernard Lyon I, CNRS UMR5308, Ecole Normale Supérieure de Lyon, Univ. Lyon, 21, avenue Tony Garnier, 69007, Lyon, France.

Current doctrine is that microvascular inflammation (MVI) triggered by a transplant -recipient antibody response against alloantigens (antibody-mediated rejection) is the main cause of graft failure. Here, we show that histological lesions are not mediated by antibodies in approximately half the participants in a cohort of 129 renal recipients with MVI on graft biopsy. Genetic analysis of these patients shows a higher prevalence of mismatches between donor HLA I and recipient inhibitory killer cell immunoglobulin-like receptors (KIRs). Human in vitro models and transplantation of β2-microglobulin-deficient hearts into wild-type mice demonstrates that the inability of graft endothelial cells to provide HLA I-mediated inhibitory signals to recipient circulating NK cells triggers their activation, which in turn promotes endothelial damage. Missing self-induced NK cell activation is mTORC1-dependent and the mTOR inhibitor rapamycin can prevent the development of this type of chronic vascular rejection.
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http://dx.doi.org/10.1038/s41467-019-13113-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6877588PMC
November 2019

Human CD8+ Tregs expressing a MHC-specific CAR display enhanced suppression of human skin rejection and GVHD in NSG mice.

Blood Adv 2019 11;3(22):3522-3538

Centre de Recherche en Transplantation et Immunologie, Unité Mixte de Recherche 1064, INSERM, Université de Nantes, Nantes, France.

Polyclonal CD8+CD45RClow/- Tregs are potent regulatory cells able to control solid organ transplantation rejection and even induce tolerance. However, donor major histocompatibility complex (MHC)-specific Tregs are more potent than polyclonal Tregs in suppressing T-cell responses and preventing acute as well as chronic rejection in rodent models. The difficulty of identifying disease-relevant antigens able to stimulate Tregs has reduced the possibility of obtaining antigen-specific Tregs. To bypass this requirement and gain the advantage of antigen specificity, and thus improve the therapeutic potential of CD8+ Tregs, we stably introduced a chimeric antigen receptor (CAR) derived from a HLA-A*02 antigen-specific antibody (A2-CAR) in human CD8+ Tregs and developed a clinically compatible protocol of transduction and expansion. We demonstrated that A2-CAR CD8+ Tregs were not phenotypically altered by the process, were specifically activated, and did not exhibit cytotoxic activity toward HLA-A*02+ kidney endothelial cells (ECs). We showed that A2-CAR CD8+ Tregs were more potent suppressors of immune responses induced by HLA-A*02 mismatch than control-CAR CD8+ Tregs, both in vitro and in vivo, in models of human skin graft rejection and graft-versus-host disease (GVHD) in NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice. We showed that integrity of human skin graft was preserved with A2-CAR CD8+ Tregs at least 100 days in vivo after administration, and that interaction between the A2-CAR CD8+ Tregs and HLA-A*02+ kidney ECs resulted in a fine-tuned and protolerogenic activation of the ECs without cytotoxicity. Together, our results demonstrated the relevance of the CAR engineering approach to develop antigen-specific CAR-CD8+ Tregs for clinical trials in transplantation, and potentially in other diseases.
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http://dx.doi.org/10.1182/bloodadvances.2019000411DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6880915PMC
November 2019

Elicited and pre-existing anti-Neu5Gc antibodies differentially affect human endothelial cells transcriptome.

Xenotransplantation 2019 11 10;26(6):e12535. Epub 2019 Jul 10.

Centre de Recherche en Transplantation et Immunologie (CRTI), INSERM, Université de Nantes, Nantes, France.

Humans cannot synthesize N-glycolylneuraminic acid (Neu5Gc) but dietary Neu5Gc can be absorbed and deposited on endothelial cells (ECs) and diet-induced anti-Neu5Gc antibodies (Abs) develop early in human life. While the interaction of Neu5Gc and diet-induced anti-Neu5Gc Abs occurs in all normal individuals, endothelium activation by elicited anti-Neu5Gc Abs following a challenge with animal-derived materials, such as following xenotransplantation, had been postulated. Ten primary human EC preparations were cultured with affinity-purified anti-Neu5Gc Abs from human sera obtained before or after exposure to Neu5Gc-glycosylated rabbit IgGs (elicited Abs). RNAs of each EC preparation stimulated in various conditions by purified Abs were exhaustively sequenced. EC transcriptomic patterns induced by elicited anti-Neu5Gc Abs, compared with pre-existing ones, were analyzed. qPCR, cytokines/chemokines release, and apoptosis were tested on some EC preparations. The data showed that anti-Neu5Gc Abs induced 967 differentially expressed (DE) genes. Most DE genes are shared following EC activation by pre-existing or anti-human T-cell globulin (ATG)-elicited anti-Neu5Gc Abs. Compared with pre-existing anti-Neu5Gc Abs, which are normal component of ECs environment, elicited anti-Neu5Gc Abs down-regulated 66 genes, including master genes of EC function. Furthermore, elicited anti-Neu5Gc Abs combined with complement-containing serum down-regulated most transcripts mobilized by serum alone. Both types of anti-Neu5Gc Abs-induced a dose- and complement-dependent release of selected cytokines and chemokines. Altogether, these data show that, compared with pre-existing anti-Neu5Gc Abs, ATG-elicited anti-Neu5Gc Abs specifically modulate genes related to cytokine responses, MAPkinase cascades, chemotaxis, and integrins and do not skew the EC transcriptome toward a pro-inflammatory profile in vitro.
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http://dx.doi.org/10.1111/xen.12535DOI Listing
November 2019

Early Acute Microvascular Kidney Transplant Rejection in the Absence of Anti-HLA Antibodies Is Associated with Preformed IgG Antibodies against Diverse Glomerular Endothelial Cell Antigens.

J Am Soc Nephrol 2019 04 8;30(4):692-709. Epub 2019 Mar 8.

Paris Descartes, Sorbonne Paris Cité University, Paris, France;

Background: Although anti-HLA antibodies (Abs) cause most antibody-mediated rejections of renal allografts, non-anti-HLA Abs have also been postulated to contribute. A better understanding of such Abs in rejection is needed.

Methods: We conducted a nationwide study to identify kidney transplant recipients without anti-HLA donor-specific Abs who experienced acute graft dysfunction within 3 months after transplantation and showed evidence of microvascular injury, called acute microvascular rejection (AMVR). We developed a crossmatch assay to assess serum reactivity to human microvascular endothelial cells, and used a combination of transcriptomic and proteomic approaches to identify non-HLA Abs.

Results: We identified a highly selected cohort of 38 patients with early acute AMVR. Biopsy specimens revealed intense microvascular inflammation and the presence of vasculitis (in 60.5%), interstitial hemorrhages (31.6%), or thrombotic microangiopathy (15.8%). Serum samples collected at the time of transplant showed that previously proposed anti-endothelial cell Abs-angiotensin type 1 receptor (AT1R), endothelin-1 type A and natural polyreactive Abs-did not increase significantly among patients with AMVR compared with a control group of stable kidney transplant recipients. However, 26% of the tested AMVR samples were positive for AT1R Abs when a threshold of 10 IU/ml was used. The crossmatch assay identified a common IgG response that was specifically directed against constitutively expressed antigens of microvascular glomerular cells in patients with AMVR. Transcriptomic and proteomic analyses identified new targets of non-HLA Abs, with little redundancy among individuals.

Conclusions: Our findings indicate that preformed IgG Abs targeting non-HLA antigens expressed on glomerular endothelial cells are associated with early AMVR, and that cell-based assays are needed to improve risk assessments before transplant.
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http://dx.doi.org/10.1681/ASN.2018080868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6442343PMC
April 2019

An early increase in endothelial protein C receptor is associated with excess mortality in pneumococcal pneumonia with septic shock in the ICU.

Crit Care 2018 10 5;22(1):251. Epub 2018 Oct 5.

Medical Intensive Care Unit, Nantes University Hospital, Nantes, France.

Background: This study investigated changes in plasma level of soluble endothelial protein C receptor (sEPCR) in association with outcome in patients with septic shock. We explored sEPCR for early sepsis prognosis assessment and constructed a scoring system based on clinical and biological data, in order to discriminate between surviving at hospital discharge and non-surviving patients.

Methods: Clinical data and samples were extracted from the prospective "STREPTOGENE" cohort. We enrolled 278 patients, from 50 intensive care units (ICUs), with septic shock caused by pneumococcal pneumonia. Patients were divided into survivors (n = 194) and non-survivors (n = 84) based on in-hospital mortality. Soluble EPCR plasma levels were quantified at day 1 (D1) and day 2 (D2) by ELISA. The EPCR gene A3 haplotype was determined. Patients were followed up until hospital discharge. Univariate and multivariate analyses were performed. A scoring system was constructed using least absolute shrinkage and selection operator (lasso) logistic regression for selecting predictive variables.

Results: In-hospital mortality was 30.2% (n = 84). Plasma sEPCR level was significantly higher at D1 and D2 in non-surviving patients compared to patients surviving to hospital discharge (p = 0.0447 and 0.0047, respectively). Early increase in sEPCR at D2 was found in non-survivors while a decrease was observed in the survival group (p = 0.0268). EPCR A3 polymorphism was not associated with mortality. Baseline sEPCR level and its variation from D1 to D2 were independent predictors of in-hospital mortality. The scoring system including sEPCR predicted mortality with an AUC of 0.75.

Conclusions: Our findings confirm that high plasma sEPCR and its increase at D2 are associated with poor outcome in sepsis and thus we propose sEPCR as a key player in the pathogenesis of sepsis and as a potential biomarker of sepsis outcome.
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http://dx.doi.org/10.1186/s13054-018-2179-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6173894PMC
October 2018

Additional Insights into Hypericum perforatum Content: Isolation, Total Synthesis, and Absolute Configuration of Hyperbiphenyls A and B from Immunomodulatory Root Extracts.

J Nat Prod 2018 08 19;81(8):1850-1859. Epub 2018 Jul 19.

EA921 SONAS, SFR4207 QUASAV, UNIV Angers , Université Bretagne Loire , 49035 Angers , France.

Phytochemical investigation of the root extracts of Hypericum perforatum led to the isolation of two biphenyl derivatives named hyperbiphenyls A and B (1 and 2) and four known xanthones (3-6). These structures were elucidated by spectroscopic and spectrometric methods including UV, NMR, and HRMS. The absolute configuration of the biphenyl derivatives was defined by two different approaches: biomimetic total synthesis of racemic hyperbiphenyl A followed by H and F NMR Mosher's esters analysis and stereoselective total synthesis of hyperbiphenyl B, permitting assignment of the S absolute configuration for both compounds. The bioactivity of compounds 1-6 toward a set of biomolecules, including major histocompatibility complex (MHC) molecules expressed on vascular endothelial cells, was measured. The results showed that the major xanthone, i.e., 5- O-methyl-2-deprenylrheediaxanthone B (3), is a potent inhibitor of MHC that efficiently reduces HLA-E, MHC-II, and MICA biomolecules on cell surfaces.
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http://dx.doi.org/10.1021/acs.jnatprod.8b00325DOI Listing
August 2018

HCMV triggers frequent and persistent UL40-specific unconventional HLA-E-restricted CD8 T-cell responses with potential autologous and allogeneic peptide recognition.

PLoS Pathog 2018 04 30;14(4):e1007041. Epub 2018 Apr 30.

Centre de Recherche en Transplantation et Immunologie (CRTI), UMR1064, INSERM, Université de Nantes, Nantes, France.

Immune response against human cytomegalovirus (HCMV) includes a set of persistent cytotoxic NK and CD8 T cells devoted to eliminate infected cells and to prevent reactivation. CD8 T cells against HCMV antigens (pp65, IE1) presented by HLA class-I molecules are well characterized and they associate with efficient virus control. HLA-E-restricted CD8 T cells targeting HCMV UL40 signal peptides (HLA-EUL40) have recently emerged as a non-conventional T-cell response also observed in some hosts. The occurrence, specificity and features of HLA-EUL40 CD8 T-cell responses remain mostly unknown. Here, we detected and quantified these responses in blood samples from healthy blood donors (n = 25) and kidney transplant recipients (n = 121) and we investigated the biological determinants involved in their occurrence. Longitudinal and phenotype ex vivo analyses were performed in comparison to HLA-A*02/pp65-specific CD8 T cells. Using a set of 11 HLA-E/UL40 peptide tetramers we demonstrated the presence of HLA-EUL40 CD8 αβT cells in up to 32% of seropositive HCMV+ hosts that may represent up to 38% of total circulating CD8 T-cells at a time point suggesting a strong expansion post-infection. Host's HLA-A*02 allele, HLA-E *01:01/*01:03 genotype and sequence of the UL40 peptide from the infecting strain are major factors affecting the incidence of HLA-EUL40 CD8 T cells. These cells are effector memory CD8 (CD45RAhighROlow, CCR7-, CD27-, CD28-) characterized by a low level of PD-1 expression. HLA-EUL40 responses appear early post-infection and display a broad, unbiased, Vβ repertoire. Although induced in HCMV strain-dependent, UL4015-23-specific manner, HLA-EUL40 CD8 T cells are reactive toward a broader set of nonapeptides varying in 1-3 residues including most HLA-I signal peptides. Thus, HCMV induces strong and life-long lasting HLA-EUL40 CD8 T cells with potential allogeneic or/and autologous reactivity that take place selectively in at least a third of infections according to virus strain and host HLA concordance.
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http://dx.doi.org/10.1371/journal.ppat.1007041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945056PMC
April 2018

Notch signaling triggered via the ligand DLL4 impedes M2 macrophage differentiation and promotes their apoptosis.

Cell Commun Signal 2018 01 10;16(1). Epub 2018 Jan 10.

Centre de Recherche en Transplantation et Immunologie UMR1064, INSERM, Université de Nantes, 30 bd J. Monnet, 44093, Nantes, France.

Background: Notch signaling controls many cellular processes, including cell fate determination, cell differentiation, proliferation and apoptosis. In mammals, four Notch receptors (Notch 1-4) can interact with five distinct ligands [Jagged1, Jagged2, Delta-like 1 (DLL1), DLL3, and DLL4]. We previously reported that Notch activation is modulated in endothelial cells and monocytes during inflammation and showed that inflammation upregulates DLL4 on endothelial cells. DLL4 promotes differentiation of blood monocytes into proinflammatory M1 macrophages. Here, we further investigated the ability of DLL4 to interfere with the polarization of blood monocytes into immunosuppressive M2 macrophages.

Methods: Human blood monocytes were differentiated in vitro into M0 macrophages and then polarized into M1 or M2 macrophages with LPS/IFNγ and IL-4, respectively. Polarization steps were performed in the presence of immobilized recombinant DLL4. Immune phenotype and apoptosis of macrophage subsets were analyzed and quantified by flow cytometry. Regulatory effects of DLL4 on gene expression, cell signaling and apoptotic pathways were investigated by QPCR and western blots.

Results: The phenotype of M2 macrophages was subject to specific alterations in the presence of recombinant DLL4. DLL4 inhibits the upregulation of IL-4 induced M2 markers such as CD11b, CD206, and CD200R. Survival of macrophages upon M2 polarization was also strongly reduced in the presence of DLL4. DLL4 induces a caspase3/7-dependent apoptosis during M2 but not M1 macrophage polarization. The Notch ligand DLL1 has no apoptotic effect. Both DLL4 signaling via Notch1 as well as DLL4-mediated apoptosis are Notch-dependent. Fully differentiated M2 macrophages became resistant to DLL4 action. Mechanistically, DLL4 selectively upregulates gene expression in macrophages upon M2 polarization, thereby affecting the Notch pattern (Notch1, 3, Jag1), activity (HES1), and transcription (IRF5, STAT1). The pro-apoptotic effectors Bax and Bak and the BH3-only proteins Bid and Bim seem to convey DLL4 apoptotic signal.

Conclusion: Interplay between the DLL4/Notch and IL-4/IL-4R signaling pathways impairs M2 differentiation. Thus, DLL4 may drive a Notch-dependent selection process not only by promoting M1 macrophage differentiation but also by preventing M2 macrophage differentiation through inhibition of M2-specific gene expression and apoptotic cell death.
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http://dx.doi.org/10.1186/s12964-017-0214-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5764024PMC
January 2018

Prenylated Polyphenols from Clusiaceae and Calophyllaceae with Immunomodulatory Activity on Endothelial Cells.

PLoS One 2016 1;11(12):e0167361. Epub 2016 Dec 1.

INSERM UMR1064, Centre de Recherche en Transplantation et Immunologie, IHU CESTI, LabEx IGO and LabEx Transplantex, Nantes, France.

Endothelial cells (ECs) are key players in inflammation and immune responses involved in numerous pathologies. Although attempts were experimentally undertaken to prevent and control EC activation, drug leads and probes still remain necessary. Natural products (NPs) from Clusiaceous and Calophyllaceous plants were previously reported as potential candidates to prevent endothelial dysfunction. The present study aimed to identify more precisely the molecular scaffolds that could limit EC activation. Here, 13 polyphenols belonging to 5 different chemical types of secondary metabolites (i.e., mammea coumarins, a biflavonoid, a pyranochromanone acid, a polyprenylated polycyclic acylphloroglucinol (PPAP) and two xanthones) were tested on resting and cytokine-activated EC cultures. Quantitative and qualitative changes in the expression of both adhesion molecules (VCAM-1, ICAM-1, E-selectin) and major histocompatibility complex (MHC) molecules have been used to measure their pharmaceutical potential. As a result, we identified 3 mammea coumarins that efficiently reduce (up to >90% at 10 μM) both basal and cytokine-regulated levels of MHC class I, class II, MICA and HLA-E on EC surface. They also prevented VCAM-1 induction upon inflammation. From a structural point of view, our results associate the loss of the free prenyl group substituting mammea coumarins with a reduced cellular cytotoxicity but also an abrogation of their anti-inflammatory potential and a reduction of their immunosuppressive effects. A PPAP, guttiferone J, also triggers a strong immunomodulation but restricted to HLA-E and MHC class II molecules. In conclusion, mammea coumarins with a free prenyl group and the PPAP guttiferone J emerge as NPs able to drastically decrease both VCAM-1 and a set of MHC molecules and to potentially reduce the immunogenicity of the endothelium.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0167361PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5131938PMC
July 2017

Alternative Splice Transcripts for MHC Class I-like MICA Encode Novel NKG2D Ligands with Agonist or Antagonist Functions.

J Immunol 2016 08 24;197(3):736-46. Epub 2016 Jun 24.

INSERM, UMR1064, LabEx Transplantex, LabEx Immunology-Graft-Oncology, and Hospital/University Institute European Center for Transplantation and Immunotherapy Sciences, Nantes, F44000 France; Centre Hospitalo-Universitaire Nantes, Institut de Transplantation et de Recherche en Transplantation-Urologie-Néphrologie, Institut de Transplantation et de Recherche en Transplantation-Urologie-Néphrologie, Nantes, F44000 France; L'Université Nantes Angers Le Mans, Université de Nantes, Faculté de Médecine, Nantes, F44000 France;

MHC class I chain-related proteins A and B (MICA and MICB) and UL16-binding proteins are ligands of the activating NKG2D receptor involved in cancer and immune surveillance of infection. Structurally, MICA/B proteins contain an α3 domain, whereas UL16-binding proteins do not. We identified novel alternative splice transcripts for MICA encoding five novel MICA isoforms: MICA-A, -B1, -B2, -C, and -D. Alternative splicing associates with MICA*015 and *017 and results from a point deletion (G) in the 5' splice donor site of MICA intron 4 leading to exon 3 and exon 4 skipping and/or deletions. These changes delete the α3 domain in all isoforms, and the α2 domain in the majority of isoforms (A, B1, C, and D). Endothelial and hematopoietic cells contained endogenous alternative splice transcripts and isoforms. MICA-B1, -B2, and -D bound NKG2D by surface plasmon resonance and were expressed at the cell surface. Functionally, MICA-B2 contains two extracellular domains (α1 and α2) and is a novel potent agonist ligand for NKG2D. We found that MICA-D is a new truncated form of MICA with weak affinity for NKG2D despite lacking α2 and α3 domains. MICA-D may functionally impair NKG2D activation by competing with full-length MICA or MICA-B2 for NKG2D engagement. Our study established NKG2D binding for recombinant MICA-B1 but found no function for this isoform. New truncated MICA isoforms exhibit a range of functions that may drive unexpected immune mechanisms and provide new tools for immunotherapy.
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http://dx.doi.org/10.4049/jimmunol.1501416DOI Listing
August 2016

Pathogenesis of non-HLA antibodies in solid organ transplantation: Where do we stand?

Hum Immunol 2016 Nov 26;77(11):1055-1062. Epub 2016 May 26.

Department of Nephrology and Kidney Transplantation, Necker Hospital, Assistance Publique-Hôpitaux de Paris, Paris, France; Paris Descartes, Sorbonne Paris Cité University, Paris, France; Centaure Foundation and Labex Transplantex, Necker Hospital, Paris, France; Necker-Enfants Malades Institute, French National Institute of Health and Medical Research U1151, Paris, France. Electronic address:

Antibody-mediated rejection (ABMR) is associated with poor transplant outcome. Pathogenic alloantibodies are usually directed against human leukocyte antigens (HLAs). Histological findings suggestive of ABMR usually demonstrate an anti-HLA donor-specific antibody (DSA)-mediated injury, while a small subset of patients develop acute dysfunction with histological lesions suggestive of ABMR in the absence of anti-HLA DSAs. Although this non-HLA ABMR is not well recognized by current diagnostic classifications, it is associated with graft dysfunction and allograft loss. These clinical descriptions suggest a pathogenic role for non-HLA anti-endothelial cell antibodies. Diverse antigenic targets have been described during the last decade. This review discusses recent findings in the field and addresses the clinical relevance of anti-endothelial cell antibodies (AECAs).
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http://dx.doi.org/10.1016/j.humimm.2016.05.021DOI Listing
November 2016

MICA Mutant A5.1 Influences BK Polyomavirus Reactivation and Associated Nephropathy After Kidney Transplantation.

J Infect Dis 2016 Sep 29;214(5):807-16. Epub 2016 Apr 29.

INSERM, UMR1064, LabEx Transplantex, LabEx IGO and IHU-CESTI CHU Nantes, Institut de Transplantation et de Recherche en Transplantation-Urologie-Néphrologie, ITUN LUNAM, Université de Nantes, Faculté de Médecine.

Background: BK polyomavirus (BKPyV) frequently reactivates in kidney transplant recipients during immunosuppressive therapy and triggers BKPyV-associated nephropathy and graft rejection. Determining effective risk factors for BKPyV reactivation is required to achieve efficient prevention.

Methods: This study investigated the role of major histocompatibility complex (MHC) class I-related chain A (MICA) in BKPyV reactivation in a cohort of 144 transplant donor/recipient pairs, including recipients with no reactivation (controllers) and those with mild (virurics) or severe (viremics) BKPyV reactivation after graft receipt.

Results: We show that, in the kidney, MICA is predominantly expressed in tubule epithelial cells, the natural targets of BKPyV, questioning a role for MICA in the immune control of BKPyV infection. Focusing on MICA genotype, we found a lower incidence of BKPyV reactivation in recipients of a renal graft from a donor carrying the MICA A5.1 mutant, which encodes a truncated nonconventional MICA. We established that a mismatch for MICA A5.1 between transplant donor and recipient is critical for BKPyV reactivation and BKPyV-associated nephropathy. Functionally, we found that a low prevalence of BKPyV reactivation was associated with elevated anti-MICA sensitization and reduced plasma level of soluble MICA in recipients, 2 potential effector mechanisms.

Discussions: These findings identify the MHC-related MICA as an immunogenetic factor that may functionally influence anti-BKPyV immune responses and infection outcomes.
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http://dx.doi.org/10.1093/infdis/jiw168DOI Listing
September 2016

Cytomegalovirus-Infected Primary Endothelial Cells Trigger NKG2C+ Natural Killer Cells.

J Innate Immun 2016 27;8(4):374-85. Epub 2016 Apr 27.

Etablissement Franx00E7;ais du Sang, ImmunoVirologie et Polymorphisme Gx00E9;nx00E9;tique, Universitx00E9; de Nantes, Nantes, France.

Among innate cells, natural killer (NK) cells play a crucial role in the defense against cytomegalovirus (CMV). In some individuals, CMV infection induces the expansion of NKG2C+ NK cells that persist after control of the infection. We have previously shown that KIR2DL+ NK cells, in contrast to NKG2C+ NK cells, contribute to controlling CMV infection using a CMV-infected monocyte-derived dendritic cell (MDDC) model. However, the nature of CMV-infected cells contributing to the expansion of the NKG2C+ NK cell subset remains unclear. To gain more insight into this question, we investigated the contribution of NKG2C+ NK cell activation by CMV-infected primary human aortic endothelial cells (EC) isolated from kidney transplant donors, which constitutively express the human leukocyte antigen (HLA)-E molecule. Here, we show that, although classic HLA class I expression was drastically downregulated, nonclassic HLA-E expression was maintained in CMV-infected EC. By comparing HLA expression patterns in CMV-infected EC, fibroblasts and MDDC, we demonstrate a cell-dependent modulation of HLA-E expression by CMV infection. NKG2C+ NK cell degranulation was significantly triggered by CMV-infected EC regardless of the nature of the HLA-E allele product. EC, predominantly present in vessels, may constitute a privileged site for CMV infection that drives a 'memory' NKG2C+ NK cell subset.
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http://dx.doi.org/10.1159/000445320DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6738823PMC
September 2017

Expression of MHC class I-related molecules MICA, HLA-E and EPCR shape endothelial cells with unique functions in innate and adaptive immunity.

Hum Immunol 2016 Nov 23;77(11):1084-1091. Epub 2016 Feb 23.

INSERM, UMR1064, LabEx Transplantex, Nantes F44000, France; CHU Nantes, Institut de Transplantation et de Recherche en Transplantation-Urologie-Néphrologie, ITUN, Nantes F44000, France; LUNAM Université de Nantes, Faculté de Médecine, Nantes F44000, France. Electronic address:

Endothelial cells (ECs) located at the interface of blood and tissues display regulatory activities toward coagulation, inflammation and vascular homeostasis. By expressing MHC class I and II antigens, ECs also contribute to immune responses. In transplantation, graft ECs are both trigger and target of alloimmune responses. ECs express a set of MHC class I-like or structural related molecules such as HLA-E, MHC class I related chain A (MICA) and the endothelial protein C receptor (EPCR) that provide multiple and unique functions to ECs. HLA-E is a low polymorphic ligand for the CD94/NKG2A/C receptors, and triggers HLA-E-restricted CD8αβT cell responses against viral and bacterial peptides. MICA is a highly polymorphic ligand for NKG2D activating NK and costimulating CD8T cells and a ligand for tissue-resident Vδ1 γδ T subsets. More intriguing is the role of EPCR, a key regulator of coagulation, as a ligand for a circulating subset of Vδ2 γδ T cells. Coexpression of this set of MHC class I-related molecules that allow ECs to activate a subtle array of immune responses upon stress and infection may also influence transplant outcome. Here, the respective structure, expression, and functions of HLA-E, MICA and EPCR as well as the impact of their polymorphism are reviewed.
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http://dx.doi.org/10.1016/j.humimm.2016.02.007DOI Listing
November 2016

Notch signaling mediates crosstalk between endothelial cells and macrophages via Dll4 and IL6 in cardiac microvascular inflammation.

Biochem Pharmacol 2016 Mar 28;104:95-107. Epub 2016 Jan 28.

INSERM UMR1064, Centre de Recherche en Transplantation et Immunologie, LabEx IGO and LabEx Transplantex, Nantes F44000, France; CHU de Nantes, Institut de Transplantation-Urologie-Néphrologie, ITUN, Nantes F44000, France; LUNAM Université de Nantes, Faculté de Médecine, Nantes F44000, France. Electronic address:

Although short-term outcomes have improved with modern era immunosuppression, little progress has been made in long-term graft survival in cardiac transplantation. Antibody-mediated rejection (AMR) is one of the leading causes of graft failure and contributes significantly to poor long-term outcomes. Endothelial cell (EC) injury, intravascular macrophage infiltrate and microvascular inflammation are the histological features of AMR. Nevertheless, mechanisms of AMR remain unclear and treatment is still limited. Here, we investigated the mechanisms underlying vascular and inflammatory cell network involved in AMR at endothelial and macrophage levels, using endomyocardial transplant biopsies and EC/monocyte cocultures. First, we found that AMR associates with changes in Notch signaling at endothelium/monocyte interface including loss of endothelial Notch4 and the acquisition of the Notch ligand Dll4 in both cell types. We showed that endothelial Dll4 induces macrophage polarization into a pro-inflammatory fate (CD40(high)CD64(high)CD200R(low) HLA-DR(low)CD11b(low)) eliciting the production of IL-6. Dll4 and IL-6 are both Notch-dependent and are required for macrophage polarization through selective down and upregulation of M2- and M1-type markers, respectively. Overall, these findings highlight the impact of the graft's endothelium on macrophage recruitment and differentiation upon AMR via Notch signaling. We identified Dll4 and IL-6 as coregulators of vascular inflammation in cardiac transplantation and as potential targets for immunotherapy.
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http://dx.doi.org/10.1016/j.bcp.2016.01.016DOI Listing
March 2016

Rabbit antithymocyte globulin-induced serum sickness disease and human kidney graft survival.

J Clin Invest 2015 Dec 9;125(12):4655-65. Epub 2015 Nov 9.

Background: Rabbit-generated antithymocyte globulins (ATGs), which target human T cells, are widely used as immunosuppressive agents during treatment of kidney allograft recipients. However, ATGs can induce immune complex diseases, including serum sickness disease (SSD). Rabbit and human IgGs have various antigenic differences, including expression of the sialic acid Neu5Gc and α-1-3-Gal (Gal), which are not synthesized by human beings. Moreover, anti-Neu5Gc antibodies have been shown to preexist and be elicited by immunization in human subjects. This study aimed to assess the effect of SSD on long-term kidney allograft outcome and to compare the immunization status of grafted patients presenting with SSD following ATG induction treatment.

Methods: We analyzed data from a cohort of 889 first kidney graft recipients with ATG induction (86 with SSD [SSD(+)] and 803 without SSD [SSD(-)]) from the Données Informatisées et Validées en Transplantation data bank. Two subgroups of SSD(+) and SSD(-) patients that had received ATG induction treatment were then assessed for total anti-ATG, anti-Neu5Gc, and anti-Gal antibodies using ELISA assays on sera before and after transplantation.

Results: SSD was significantly associated with long-term graft loss (>10 years, P = 0.02). Moreover, SSD(+) patients exhibited significantly elevated titers of anti-ATG (P = 0.043) and anti-Neu5Gc (P = 0.007) IgGs in late post-graft samples compared with SSD(-) recipients.

Conclusion: In conclusion, our data indicate that SSD is a major contributing factor of late graft loss following ATG induction and that anti-Neu5Gc antibodies increase over time in SSD(+) patients.

Funding: This study was funded by Société d'Accélération du Transfert de Technologies Ouest Valorisation, the European FP7 "Translink" research program, the French National Agency of Research, Labex Transplantex, the Natural Science and Engineering Research Council of Canada, and the Canadian Foundation for Innovation.
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http://dx.doi.org/10.1172/JCI82267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4665787PMC
December 2015

Lepidotol A from Mesua lepidota Inhibits Inflammatory and Immune Mediators in Human Endothelial Cells.

J Nat Prod 2015 Sep 24;78(9):2187-97. Epub 2015 Aug 24.

EA921 SONAS, SFR4207 QUASAV, Université d'Angers , Angers, France.

Phytochemical investigation on the fruits of Mesua lepidota (Calophyllaceae) led to the isolation of seven new phenylcoumarin derivatives named lepidotols A-E (1-5) and lepidotins A and B (6, 7). These structures were elucidated by spectroscopic and spectrometric methods including UV, NMR, and HRMS. Lepidotol A (1), the major compound, was evaluated for its inhibitory effect on inflammation and immunity using endothelial cell-based cellular assays. At 10 μM, 1 exhibited an anti-inflammatory activity, with a significant inhibition of vascular cell adhesion molecule 1 and intercellular adhesion molecule 1 expression induced by tumor necrosis factor-α. Lepidotol A also showed a mild immunosuppressive effect, with inhibition of the major histocompatibility complex molecules, namely, human leukocyte antigen (HLA)-DR and HLA-E.
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http://dx.doi.org/10.1021/acs.jnatprod.5b00222DOI Listing
September 2015

The disintegrin and metalloproteinase ADAM10 mediates a canonical Notch-dependent regulation of IL-6 through Dll4 in human endothelial cells.

Biochem Pharmacol 2014 Oct 15;91(4):510-21. Epub 2014 Aug 15.

INSERM UMR1064, Centre de Recherche en Transplantation et Immunologie, LabEx IGO and LabEx Transplantex, Nantes F44000, France; CHU de Nantes, Institut de Transplantation-Urologie-Néphrologie, ITUN, Nantes F44000, France; LUNAM, Université de Nantes, Faculté de Médecine, Nantes F44000, France. Electronic address:

Although the involvement of the disintegrin and metalloproteinase ADAM10 in several areas of vascular biology is now clearly established, its role in vascular inflammation and in Notch signaling at the endothelial level remains unclear. In this study, we demonstrated that ADAM10 specifically localizes in the CD31(+) endothelial cells (ECs) in normal human cardiac tissues and in cultured primary arterial ECs. In vitro, ADAM10 drives a specific regulation of the Notch pathway in vascular ECs. Using an ADAM10 gain and loss of function approach we show an ADAM10-dependent regulation of Dll1 and Dll4 expression in association with changes in Hes1 and Hey1 expression. We also identified IL-6, IL-8, MCP-1 and sVCAM-1 as novel targets of ADAM10 upon inflammation. Although Notch pathway does not seem to be required for the production of IL-8, MCP-1 and sVCAM-1, the release of IL-6 by ECs occurred through ADAM10 and a canonical Notch signaling pathway, dependent of γ-secretase activity. Moreover, sustained expression of Dll4 mediated by ADAM10 elicits an increased release of IL-6 suggesting a strong implication of the specific Dll4 signaling in this mechanism. Modulation of IL-6 mediated by ADAM10/Notch signaling required PI3K activity. Thus, our findings suggest that ADAM10/Dll4 signaling is a major signaling pathway in ECs driving inflammatory events involved in inflammation and immune cell recruitment.
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http://dx.doi.org/10.1016/j.bcp.2014.08.007DOI Listing
October 2014

Advanced glycation inhibition and protection against endothelial dysfunction induced by coumarins and procyanidins from Mammea neurophylla.

Fitoterapia 2014 Jul 13;96:65-75. Epub 2014 Apr 13.

Université d'Angers, SFR QUASAV, EA 921 SONAS, Angers, France. Electronic address:

Advanced glycation end-products (AGEs) are associated with many pathogenic disorders such as pathogenesis of diabetes or endothelial dysfunction leading to cardiovascular events. Therefore, the identification of new anti-AGE molecules or extracts aims at preventing such pathologies. Many Clusiaceae and Calophyllaceae species are used in traditional medicines to treat arterial hypertension as well as diabetes. Focusing on these plant families, an anti-AGE plant screening allowed us to select Mammea neurophylla for further phytochemical and biological studies. Indeed, both DCM and MeOH stem bark extracts demonstrated in vitro their ability to prevent inflammation in endothelial cells and to reduce vasoconstriction. A bioguided fractionation of these extracts allowed us to point out 4-phenyl- and 4-(1-acetoxypropyl)coumarins and procyanidins as potent inhibitors of AGE formation, potentially preventing endothelial dysfunction. The fractionation steps also led to the isolation of two new compounds, namely neurophyllols A and B from the DCM bark extract together with thirteen known mammea A and E coumarins (mammea A/AA, mammea A/AB, mammea A/BA, mammea A/BB, mammea A/AA cycloD, mammea A/AB cycloD, disparinol B, mammea A/AB cycloE, ochrocarpin A, mammea A/AA cycloF, mammea A/AB cycloF, mammea E/BA, mammea E/BB) as well as δ-tocotrienol, xanthones (1-hydroxy-7-methoxyxanthone, 2-hydroxyxanthone) and triterpenes (friedelin and betulinic acid). During this study, R,S-asperphenamate, previously described from fungal origin was also purified.
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http://dx.doi.org/10.1016/j.fitote.2014.04.005DOI Listing
July 2014

Long-term IgG response to porcine Neu5Gc antigens without transmission of PERV in burn patients treated with porcine skin xenografts.

J Immunol 2013 Sep 14;191(6):2907-15. Epub 2013 Aug 14.

Department of Life Sciences, Glasgow Caledonian University, Glasgow G4 0BA, United Kingdom.

Acellular materials of xenogenic origin are used worldwide as xenografts, and phase I trials of viable pig pancreatic islets are currently being performed. However, limited information is available on transmission of porcine endogenous retrovirus (PERV) after xenotransplantation and on the long-term immune response of recipients to xenoantigens. We analyzed the blood of burn patients who had received living pig-skin dressings for up to 8 wk for the presence of PERV as well as for the level and nature of their long term (maximum, 34 y) immune response against pig Ags. Although no evidence of PERV genomic material or anti-PERV Ab response was found, we observed a moderate increase in anti-αGal Abs and a high and sustained anti-non-αGal IgG response in those patients. Abs against the nonhuman sialic acid Neu5Gc constituted the anti-non-αGal response with the recognition pattern on a sialoglycan array differing from that of burn patients treated without pig skin. These data suggest that anti-Neu5Gc Abs represent a barrier for long-term acceptance of porcine xenografts. Because anti-Neu5Gc Abs can promote chronic inflammation, the long-term safety of living and acellular pig tissue implants in recipients warrants further evaluation.
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http://dx.doi.org/10.4049/jimmunol.1301195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3782708PMC
September 2013

Endothelial cell activation and proliferation modulate NKG2D activity by regulating MICA expression and shedding.

J Innate Immun 2014 16;6(1):89-104. Epub 2013 Jul 16.

Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche 1064, Centre de Recherche en Transplantation et Immunologie, Laboratoire d'Excellence Transplantex, Centre Européen des Sciences de la Transplantation et Immunothérapie, Centre Hospitalier Universitaire de Nantes, Faculté de Médecine, Université de Nantes, Nantes, France.

MICA are major histocompatibility complex class I-related molecules, expressed by endothelial cells (ECs), that may be targets for alloantibodies and NKG2D-expressing natural killer (NK) and T effector cells in organ allografts. This study shows that basal levels of MICA expressed on vascular ECs is sufficient to functionally modulate the expression and activity of the immunoreceptor NKG2D in allogeneic NK cells. We found that MICA expression is differentially regulated at the EC surface in response to cytokines. TNFα upregulates MICA while IFNγ significantly decreases MICA at the EC surface. Both cytokines induce the release of soluble MICA by ECs. Modulation of NKG2D correlates with the MICA level on the EC surface. Glycosylation and metalloproteinase activities account for major post-transcriptional mechanisms controlling MICA level and the function in ECs. Our results indicate that, in addition to the NFκB pathway, the mitogen-activated protein kinase pathways JNK, ERK1/2 and p38 are key signaling pathways in the control of MICA by the cytokines. Finally, we show that EC proliferation mediated by FGF-2 or wound healing increases the MICA level. Together, our data suggest that inflammation and proliferation regulate endothelial MICA expression and shedding, enabling ECs to modulate NKG2D activity on effector NK and T cells, and provide further evidence of a role for ECs in immunoregulation.
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http://dx.doi.org/10.1159/000351605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6784110PMC
October 2014

MICA variant promotes allosensitization after kidney transplantation.

J Am Soc Nephrol 2013 May 28;24(6):954-66. Epub 2013 Mar 28.

Centre de Recherche en Transplantation et Immunologie, LabEx Transplantex, Nantes, France.

MHC class I-related chain A (MICA) antigens are surface glycoproteins strongly implicated in innate immunity, and the MICA gene is highly polymorphic. Clinical observations suggest a role for donor MICA antigens expressed on transplant endothelial cells in the alloimmune response, but the effect of MICA genotype is not well understood. Here, we investigated the immunologic effect of the A5.1 mutation, related to the common MICA*008 allele. Compared with wild-type endothelial cells (ECs), homozygosity for MICA A5.1 associated with an endothelial phenotype characterized by 7- to 10-fold higher levels of MICA mRNA and MICA proteins at the cell surface, as well as exclusive release in exosomes instead of enzymatic cleavage. Mechanistically, we did not detect quantitative changes in regulatory microRNAs. Functionally, A5.1 ECs enhanced NKG2D interaction and natural killer cell activation, promoting NKG2D-dependent lysis of ECs. In kidney transplant recipients, polyreactive anti-MICA sera bound preferentially to ECs from MICA A5.1 donors, suggesting that MICA*008(A5.1) molecules are the preferential antigenic determinants on ECs of grafts. Furthermore, the incidence of MICA A5.1 mismatch revealed a statistically significant association between donor MICA A5.1 and both anti-MICA sensitization and increased proteinuria in kidney recipients. Taken together, these results identify the A5.1 mutation as an immunodominant factor and a potential risk factor for transplant survival.
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http://dx.doi.org/10.1681/ASN.2012080814DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3665393PMC
May 2013

Impact of notch signaling on inflammatory responses in cardiovascular disorders.

Int J Mol Sci 2013 Mar 26;14(4):6863-88. Epub 2013 Mar 26.

Division of Cardiovascular Medicine, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA.

Notch signaling is a major pathway in cell fate decisions. Since the first reports showing the major role of Notch in embryonic development, a considerable and still growing literature further highlights its key contributions in various pathological processes during adult life. In particular, Notch is now considered as a major player in vascular homeostasis through the control of key cellular functions. In parallel, confounding evidence emerged that inflammatory responses regulate Notch signaling in vitro in endothelial cells, smooth muscle cells or vascular infiltrating cells and in vivo in vascular and inflammatory disorders and in cardiovascular diseases. This review presents how inflammation influences Notch in vascular cells and, reciprocally, emphasizes the functional role of Notch on inflammatory processes, notably by regulating key cell functions (differentiation, proliferation, apoptosis/survival, activation). Understanding how the disparity of Notch receptors and ligands impacts on vasculature biology remains critical for the design of relevant and adequate therapeutic strategies targeting Notch in this major pathological context.
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http://dx.doi.org/10.3390/ijms14046863DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3645668PMC
March 2013

Differential roles of Hath1, MUC2 and P27Kip1 in relation with gamma-secretase inhibition in human colonic carcinomas: a translational study.

PLoS One 2013 11;8(2):e55904. Epub 2013 Feb 11.

EA Biometadys, Université de Nantes, Nantes, France.

Hath1, a bHLH transcription factor negatively regulated by the γ-secretase-dependent Notch pathway, is required for intestinal secretory cell differentiation. Our aim was fourfold: 1) determine whether Hath1 is able to alter the phenotype of colon cancer cells that are committed to a differentiated phenotype, 2) determine whether the Hath1-dependent alteration of differentiation is coupled to a restriction of anchorage-dependent growth, 3) decipher the respective roles of three putative tumor suppressor genes Hath1, MUC2 and P27kip1 in this coupling and, 4) examine how our findings translate to primary tumors. Human colon carcinoma cell lines that differentiate along a mucin secreting (MUC2/MUC5AC) and/or enterocytic (DPPIV) lineages were maintained on inserts with or without a γ-secretase inhibitor (DBZ). Then the cells were detached and their ability to survive/proliferate in the absence of substratum was assessed. γ-secretase inhibition led to a Hath1-mediated preferential induction of MUC2 over MUC5AC, without DPPIV modification, in association with a decrease in anchorage-independent growth. While P27kip1 silencing relieved the cells from the Hath1-induced decrease of anchorage-independent growth, MUC2 silencing did not modify this parameter. Hath1 ectopic expression in the Hath1 negative enterocytic Caco2 cells led to a decreased anchorage-independent growth in a P27kip1-independent manner. In cultured primary human colon carcinomas, Hath1 was up-regulated in 7 out of 10 tumors upon DBZ treatment. Parallel MUC2 up-regulation occurred in 4 (4/7) and P27kip1 in only 2 (2/7) tumors. Interestingly, the response patterns of primary tumors to DBZ fitted with the hierarchical model of divergent signalling derived from our findings on cell lines.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0055904PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3569436PMC
August 2013

HLA-E-restricted cross-recognition of allogeneic endothelial cells by CMV-associated CD8 T cells: a potential risk factor following transplantation.

PLoS One 2012 30;7(11):e50951. Epub 2012 Nov 30.

Inserm, U892, Nantes, France.

Although association between CMV infection and allograft rejection is well admitted, the precise mechanisms involved remain uncertain. Here, we report the characterization of an alloreactive HLA-E-restricted CD8 T cell population that was detected in the PBL of a kidney transplant patient after its CMV conversion. This monoclonal CD8 T cell population represents a sizable fraction in the blood (3% of PBL) and is characterized by an effector-memory phenotype and the expression of multiple NK receptors. Interestingly, these unconventional T cells display HLA-E-dependent reactivity against peptides derived from the leader sequences of both various HCMV-UL40 and allogeneic classical HLA-I molecules. Consequently, while HLA-E-restricted CD8 T cells have potential to contribute to the control of CMV infection in vivo, they may also directly mediate graft rejection through recognition of peptides derived from allogeneic HLA-I molecules on graft cells. Therefore, as HLA-E expression in nonlymphoid organs is mainly restricted to endothelial cells, we investigated the reactivity of this HLA-E-restricted T cell population towards allogeneic endothelial cells. We clearly demonstrated that CMV-associated HLA-E-restricted T cells efficiently recognized and killed allogeneic endothelial cells in vitro. Moreover, our data indicate that this alloreactivity is tightly regulated by NK receptors, especially by inhibitory KIR2DL2 that strongly prevents TCR-induced activation through recognition of HLA-C molecules. Hence, a better evaluation of the role of CMV-associated HLA-E-restricted T cells in transplantation and of the impact of HLA-genotype, especially HLA-C, on their alloreactivity may determine whether they indeed represent a risk factor following organ transplantation.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0050951PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3511380PMC
May 2013

Signaling of endothelial cytoprotection in transplantation.

Hum Immunol 2012 Dec 25;73(12):1245-52. Epub 2012 Jul 25.

INSERM, UMR1064, Nantes, France.

A better knowledge of the processes by which endothelium can resist to cell death and adapt to injury by specific intracellular signaling pathways and dedicated protein regulation is a key step to understand how vascular inflammation/injury develops and how it is regulated. This review focuses on signaling pathways and molecular effectors that trigger the balance between endothelial cell activation and dysfunction. In addition to the canonical nuclear factor-κB (NF-κB), phosphatidyl inositol 3-kinase (PI-3K) and mitogen-activated protein kinases (MAPK) that orchestrated the inflammatory response and its termination we report here additive pathways such as Notch pathway and protein C/protease activated receptor (PAR) pathway that have been also reported to play a role in the control of EC activation and apoptosis. This review also provides an update of the characteristics of some established and novel protective molecules for the endothelium, identified in transplantation.
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http://dx.doi.org/10.1016/j.humimm.2012.07.012DOI Listing
December 2012