MHC Class II Research Articles

Thymic Epithelial Cell Alterations and Defective Thymopoiesis Lead to Central and Peripheral Tolerance Perturbation in MHCII Deficiency.

Major Histocompatibility Complex (MHC) class II (MHCII) deficiency (MHCII-D), also known as Bare Lymphocyte Syndrome (BLS), is a rare combined immunodeficiency due to mutations in genes regulating expression of MHCII molecules. MHCII deficiency results in impaired cellular and humoral immune responses, leading to severe infections and autoimmunity. Abnormal cross-talk with developing T cells due to the absence of MHCII expression likely leads to defects in thymic epithelial cells (TEC). However, the contribution of TEC alterations to the pathogenesis of this primary immunodeficiency has not been well characterized to date, in particular in regard to immune dysregulation. To this aim, we have performed an in-depth cellular and molecular characterization of TEC in this disease. We observed an overall perturbation of thymic structure and function in both MHCII−/− mice and patients. Transcriptomic and proteomic profiling of murine TEC revealed several alterations. In particular, we demonstrated that impairment of lymphostromal cross-talk in the thymus of MHCII−/− mice affects mTEC maturation and promiscuous gene expression and causes defects of central tolerance. Furthermore, we observed peripheral tolerance impairment, likely due to defective Treg cell generation and/or function and B cell tolerance breakdown. Overall, our findings reveal disease-specific TEC defects resulting in perturbation of central tolerance and limiting the potential benefits of hematopoietic stem cell transplantation in MHCII deficiency.

Induction of antigen-specific tolerance by nanobody-antigen adducts that target class-II major histocompatibility complexes.

The association of autoimmune diseases with particular allellic products of the class-II major histocompatibility complex (MHCII) region implicates the presentation of the offending self-antigens to T cells. Because antigen-presenting cells are tolerogenic when they encounter an antigen under non-inflammatory conditions, the manipulation of antigen presentation may induce antigen-specific tolerance. Here, we show that, in mouse models of experimental autoimmune encephalomyelitis, type 1 diabetes and rheumatoid arthritis, the systemic administration of a single dose of nanobodies that recognize MHCII molecules and conjugated to the relevant self-antigen under non-inflammatory conditions confers long-lasting protection against these diseases. Moreover, co-administration of a nanobody-antigen adduct and the glucocorticoid dexamethasone, conjugated to the nanobody via a cleavable linker, halted the progression of established experimental autoimmune encephalomyelitis in symptomatic mice and alleviated their symptoms. This approach may represent a means of treating autoimmune conditions.

Genetic and commensal induction of IL-18 drive intestinal epithelial MHCII via IFNγ.

Major Histocompatibility Complex Class II (MHCII) is dynamically expressed on intestinal epithelial cells (IECs) throughout the intestine, but its regulation remains poorly understood. We observed that spontaneous upregulation of IEC MHCII in locally-bred Rag1−/− mice correlated with serum Interleukin (IL)-18, was transferrable via cohousing to commercially-bred immunodeficient mice and could be inhibited by both IL-12 and IL-18 blockade. Overproduction of intestinal IL-18 due to an activating Nlrc4 mutation upregulated IEC MHCII via classical inflammasome machinery independently of immunodeficiency or dysbiosis. Immunodeficient dysbiosis increased Il18 transcription, which synergized with NLRC4 inflammasome activity to drive elevations in serum IL-18. This IL-18-MHCII axis was confirmed in several other models of intestinal and systemic inflammation. Elevated IL-18 reliably preceded MHCII upregulation, suggesting an indirect effect on IECs, and mice with IL-18 overproduction showed activation or expansion of type 1 lymphocytes. Interferon Gamma (IFNg) was uniquely able to upregulate IEC MHCII in enteroid cultures and was required for MHCII upregulation in several in vivo systems. Thus, we have linked intestinal dysbiosis, systemic inflammation, and inflammasome activity to IEC MHCII upregulation via an intestinal IL-18-IFNg axis. Understanding this process may be crucial for determining the contribution of IEC MHCII to intestinal homeostasis, host defense, and tolerance.

Lactobacillus johnsonii activates porcine monocyte derived dendritic cells maturation to modulate Th cellular immune response

Lactobacilli are abundant in the intestinal tract where they constantly regulate immune system via interacting with a great diversity of immune cells, such as dendritic cells (DCs). Notably, DCs are powerful antigen-presenting cells and they are capable of initiating primary immune responses. In this study, we studied the effects of Lactobacillus johnsonii (L. johnsonii) and Lactobacillus johnsonii cell-free supernatant (L. johnsonii-CFS) on the activation of porcine monocyte-derived dendritic cells (MoDCs) and their regulation of Th cellular immune responses in vitro. The MoDCs generated from porcine peripheral blood monocytes were stimulated by L. johnsonii and L. johnsonii-CFS, respectively. Pre-incubation with L. johnsonii increased expression of CD172a, CD80, major histocompatibility complex class II (MHCII) in MoDCs, and enhanced the ability of MoDCs to induce the proliferation of CD4+ T cell, while pre-incubation with L. johnsonii-CFS merely upregulated the expression of MHCII. Analysis of the cytokines showed that L. johnsonii stimulated up-regulation of Th1-type cytokines (IL-12p40, IFN-γ, TNF-α), pro-inflammatory cytokine IL-1β, chemokine CCL20, and Treg-type / anti-inflammatory cytokines IL-10 in MoDCs. Notably, a high production of IL-10 was observed in the MoDCs treated with L. johnsonii-CFS, indicating L. johnsonii-CFS exerted anti-inflammatory effects. Furthermore, L. johnsonii induced up-regulation of TLR2 and TLR6, but L. johnsonii-CFS not. Moreover, MoDCs stimulated by L. johnsonii mainly promoted T cell differentiate into Th1/Th2/Treg cells and plays an important role in improving the balance between Th1/Th2/Treg-type cells, whereas MoDCs stimulated by L. johnsonii-CFS mainly directed T cell to Th2/Treg subset polarization. In conclusion, L. johnsonii and L. johnsonii-CFS exhibited the ability of modulating innate immunity by regulating immunological functions of MoDCs in vitro, suggesting their potential ability to use as microecological preparations and medicines.

Fasciola gigantica tegumental calcium-binding EF-hand protein 4 exerts immunomodulatory effects on goat monocytes

BackgroundThe liver fluke Fasciola gigantica secretes excretory-secretory proteins during infection to mediate its interaction with the host. In this study, we investigated the immunomodulatory effects of a recombinant tegumental calcium-binding EF-hand protein 4 of F. gigantica (rFg-CaBP4) on goat monocytes.MethodsThe rFg-CaBP4 protein was induced and purified by affinity chromatography. The immunogenic reaction of rFg-CaBP4 against specific antibodies was detected through western blot analysis. The binding of rFg-CaBP4 on surface of goat monocytes was visualized by immunofluorescence assay. The localization of CaBP4 within adult fluke structure was detected by immunohistochemical analysis. The cytokine transcription levels in response to rFg-CaBP4 were examined using ABI 7500 real-time PCR system. The expression of the major histocompatibility complex (MHC) class-II molecule (MHC-II) in response to rFg-CaBP4 protein was analyzed using Flow cytometry.ResultsThe isopropyl-ß-D-thiogalactopyranoside-induced rFg-CaBP4 protein reacted with rat sera containing anti-rFg-CaBP4 polyclonal antibodies in a western blot analysis. The adhesion of rFg-CaBP4 to monocytes was visualized by immunofluorescence and laser scanning confocal microscopy. Immunohistochemical analysis localized native CaBP4 to the oral sucker, pharynx, genital pore, acetabulum and tegument of adult F. gigantica. Co-incubation of rFg-CaBP4 with concanavalin A-stimulated monocytes increased the transcription levels of interleukin (IL)-2, IL-4, interferon gamma and transforming growth factor-β. However, a reduction in the expression of IL-10 and no change in the expression of tumor necrosis factor-α were detected. Additionally, rFg-CaBP4-treated monocytes exhibited a marked increase in the expression of the major histocompatibility complex (MHC) class-II molecule (MHC-II) and a decrease in MHC-I expression, in a dose-dependent manner.ConclusionsThese findings provide additional evidence that calcium-binding EF-hand proteins play roles in host-parasite interaction. Further characterization of the immunomodulatory role of rFg-CaBP4 should expand our understanding of the strategies used by F. gigantica to evade the host immune responses.Graphical abstract

Comparison of immune microenvironment between primary lung tumors and paired brain metastatic tumors.

2020 Background: Lung cancer is one of the most common causes of brain metastases (BMs) and is always associated with poor prognosis. To evaluate the characteristics of the tumor immune microenvironment in brain metastases of non-small-cell lung cancer (NSCLC), we investigated the immunophenotype of primary NSCLC and paired brain metastases. Methods: Forty-three Chinese patients with NSCLC who had BMs at presentation or during the course of their disease were admitted to the Sun Yat-Sen University Cancer Center (Guangzhou, China) from 2000 to 2019. RNA sequencing (RNA-seq) of eighty-six formalin-fixed, paraffin embedded (FFPE) samples from primary lung tumors and paired brain metastases of 43 patients was conducted to comprehensively analyze the tumor immune microenvironment. Results: Our data revealed that brain metastases compared with primary lung tumors exhibited reduced tumor infiltrating lymphocytes (TILs) (all 28 immune cell subtypes P < 0.05), lower fraction of activated CD8 T cell and effector memory CD8 T cell in total TILs (P = 0.028, P < 0.001, respectively); higher fraction of macrophage and neutrophil in total TILs (P < 0.001, P < 0.01, respectively). Comparing with the primary lung tumors, the scores of some immune related signatures, including MHC non-class signature, IFN gamma signature and T-cell-inflamed gene-expression profile (GEP) signature, were significantly lower in brain metastases (P = 0.004, P = 0.009, P = 0.004, respectively), while the score of MHC class-II signature was higher in brain metastases (P = 0.045). We found the distributions of tumor microenvironment immune types (TMIT) in brain metastases and primary lung tumors were different. Brain metastases contained significantly lower proportion of TMIT I (high PD-L1/ high CD8A) (23%) than primary lung tumors (47%) (P < 0.05). Besides, we found three immune inhibitory checkpoint molecules, namely C10orf54 (VISTA), CTLA4 and CD274 (PD-L1) were downregulated in brain metastases than in primary lung tumors (P < 0.001, P < 0.001, P = 0.034, respectively). Moreover, there was poor correlation of PD-L1 expression between paired brain metastases and primary lung tumors (R = 0.28, P = 0.068). Unsupervised hierarchic cluster analysis revealed the primary lung tumors had two distinct patterns of immune gene signatures, namely Cluster A and Cluster B, and the tumors in Cluster B were immune rich, but associated with poor prognosis (log-rank P = 0.021). Conclusions: Our work illustrates the immune landscape of brain metastases from NSCLC, and suggests that the tumor immune microenvironment in brain metastases compared with primary lung tumors is further immunosuppressed, that may help guide immunotherapeutic strategies for NSCLC brain metastases.

How can the MHC mediate social odor via the microbiota community? A deep dive into mechanisms

AbstractGenes of the major histocompatibility complex (MHC) have long been linked to odor signaling and recently researchers’ attention has focused on MHC structuring of microbial communities and how this may in turn impact odor. However, understanding of the mechanisms through which the MHC could affect the microbiota to produce a chemical signal that is both reliable and strong enough to ensure unambiguous transmission of behaviorally important information remains poor. This is largely because empirical studies are rare, predictions are unclear, and the underlying immunological mechanisms governing MHC–microbiota interactions are often neglected. Here, we review the immunological processes involving MHC class II (MHC-II) that could affect the commensal community. Focusing on immunological and medical research, we provide background knowledge for nonimmunologists by describing key players within the vertebrate immune system relating to MHC-II molecules (which present extracellular-derived peptides, and thus interact with extracellular commensal microbes). We then systematically review the literature investigating MHC–odor–microbiota interactions in animals and identify areas for future research. These insights will help to design studies that are able to explore the role of MHC-II and the microbiota in the behavior of wild populations in their natural environment and consequently propel this research area forward.

B cells modulate the expression of MHC-II on cardiac CCR2− macrophages

The uninjured murine heart contains a heterogeneous population of macrophages with disparate ontogenies and functions. These macrophages are often associated with blood vessels and can be subclassified based on the expression of CC chemokine receptor 2 (CCR2) and major histocompatibility complex class II (MHC-II). The biological cues that modulate these macrophage pool subpopulations have not been completely identified. It has been recently shown that a sub-population of circulating naïve B cells adheres to the myocardial microvasculature. We hypothesized that B cells might modulate the phenotype of myocardial macrophages. To test this hypothesis, we analyzed both the relative location of B cells and macrophages in myocardial histological section and the prevalence of myocardial macrophage subsets in hearts from B cell–deficient mice (μMT) and mice depleted of B cells through administration of an anti-CD20 antibody. We found that B cells pause in the microvasculature in proximity of macrophages and modulate the number of myocardial CCR2−MHC-IIhigh cells. Through in vitro studies we found that this is likely the result of a paracrine effect of B cells on the expression of MHC-II in CCR2− cells. These results reveal an unexpected relationship between B cells and resident macrophages and, highlighting a direct intramyocardial effect of circulating B cells, challenge the currently held belief that naïve recirculating B lymphocytes merely shuttle between lymphoid stations.

Postoperative abdominal sepsis induces selective and persistent changes in CTCF binding within the MHC-II region of human monocytes.

BackgroundPostoperative abdominal infections belong to the most common triggers of sepsis and septic shock in intensive care units worldwide. While monocytes play a central role in mediating the initial host response to infections, sepsis-induced immune dysregulation is characterized by a defective antigen presentation to T-cells via loss of Major Histocompatibility Complex Class II DR (HLA-DR) surface expression. Here, we hypothesized a sepsis-induced differential occupancy of the CCCTC-Binding Factor (CTCF), an architectural protein and superordinate regulator of transcription, inside the Major Histocompatibility Complex Class II (MHC-II) region in patients with postoperative sepsis, contributing to an altered monocytic transcriptional response during critical illness.ResultsCompared to a matched surgical control cohort, postoperative sepsis was associated with selective and enduring increase in CTCF binding within the MHC-II. In detail, increased CTCF binding was detected at four sites adjacent to classical HLA class II genes coding for proteins expressed on monocyte surface. Gene expression analysis revealed a sepsis-associated decreased transcription of (i) the classical HLA genes HLA-DRA, HLA-DRB1, HLA-DPA1 and HLA-DPB1 and (ii) the gene of the MHC-II master regulator, CIITA (Class II Major Histocompatibility Complex Transactivator). Increased CTCF binding persisted in all sepsis patients, while transcriptional recovery CIITA was exclusively found in long-term survivors.ConclusionOur experiments demonstrate differential and persisting alterations of CTCF occupancy within the MHC-II, accompanied by selective changes in the expression of spatially related HLA class II genes, indicating an important role of CTCF in modulating the transcriptional response of immunocompromised human monocytes during critical illness.

MHC class II tetramers engineered for enhanced binding to CD4 improve detection of antigen-specific T cells.

The ability to identify T cells that recognize specific peptide antigens bound to major histocompatibility complex (MHC) molecules has enabled enumeration and molecular characterization of the lymphocytes responsible for cell-mediated immunity. Fluorophore-labeled peptide:MHC class I (p:MHCI) tetramers are well-established reagents for identifying antigen-specific CD8+ T cells by flow cytometry, but efforts to extend the approach to CD4+ T cells have been less successful, perhaps owing to lower binding strength between CD4 and MHC class II (MHCII) molecules. Here we show that p:MHCII tetramers engineered by directed evolution for enhanced CD4 binding outperform conventional tetramers for the detection of cognate T cells. Using the engineered tetramers, we identified about twice as many antigen-specific CD4+ T cells in mice immunized against multiple peptides than when using traditional tetramers. CD4 affinity-enhanced p:MHCII tetramers, therefore, allow direct sampling of antigen-specific CD4+ T cells that cannot be accessed with conventional p:MHCII tetramer technology. These new reagents could provide a deeper understanding of the T cell repertoire.

B-cell-intrinsic MHCII signaling promotes microbiota diversity

Abstract The expression of major histocompatibility complex class II (MHCII) molecules is essential for the formation of germinal centers (GC) in lymphoid follicles, which are the primary sites for the generation of T-cell-dependent (TD) high-affinity antibody responses. Gut peyer's patches (PPs) are the dominant peripheral lymphoid tissues that give rise to TD antibody (primarily immunoglobulin A (IgA)) responses generated against the gut microbiota. While it is anticipated, a requirement of B-cell-intrinsic MHCII expression for GC formation has never been formally described. Additionally, while anti-commensal TD IgA responses have been shown to regulate microbiota composition and function, to what degree B-cell-intrinsic MHCII influences this process is also undefined. Here, we use a RAG1−/− adoptive transfer model, where RAG1−/− mice are either reconstituted with naive CD4+ T cells and MHCII+ B cells or naive CD4+ T cells and MHCII−B cells to address these gaps in our knowledge. Results from these experiments demonstrate that B-cell-intrinsic MHCII signaling is a strict requirement for GC-TFH cell development. Consequently, B-cell-intrinsic MHCII signaling promotes the generation of high-affinity anti-commensal IgA responses in the gut, which lead to increased species richness within the fecal but not small intestinal microbial community. Collectively, our data suggest that B-cell-intrinsic MHCII signaling is crucial for the generation of high-affinity anti-commensal IgA responses generated against the gut microbiota, and that this response favors a more diverse bacterial community.

The Murine MHC Class II Super Enhancer IA/IE-SE Contains a Functionally Redundant CTCF-Binding Component and a Novel Element Critical for Maximal Expression.

In both humans and mice, CTCF-binding elements form a series of interacting loops across the MHC class II (MHC-II) locus, and CTCF is required for maximal MHC-II gene expression. In humans, a CTCF-bound chromatin insulator termed XL9 and a super enhancer (SE) DR/DQ-SE situated in the intergenic region between HLA-DRB1 and HLA-DQA1 play critical roles in regulating MHC-II expression. In this study, we identify a similar SE, termed IA/IE-SE, located between H2-Eb1 and H2-Aa of the mouse that contains a CTCF site (C15) and a novel region of high histone H3K27 acetylation. A genetic knockout of C15 was created and its role on MHC-II expression tested on immune cells. We found that C15 deletion did not alter MHC-II expression in B cells, macrophages, and macrophages treated with IFN-γ because of functional redundancy of the remaining MHC-II CTCF sites. Surprisingly, embryonic fibroblasts derived from C15-deleted mice failed to induce MHC-II gene expression in response to IFN-γ, suggesting that at least in this developmental lineage, C15 was required. Examination of the three-dimensional interactions with C15 and the H2-Eb1 and H2-Aa promoters identified interactions within the novel region of high histone acetylation within the IA/IE-SE (termed N1) that contains a PU.1 binding site. CRISPR/Cas9 deletion of N1 altered chromatin interactions across the locus and resulted in reduced MHC-II expression. Together, these data demonstrate the functional redundancy of the MHC-II CTCF elements and identify a functionally conserved SE that is critical for maximal expression of MHC-II genes.

Elevated phytohaemagglutinin‐induced skin‐swelling response at an intermediate number of MHC class II alleles in bluethroat nestlings

Genes of the major histocompatibility complex (MHC) are crucial for adaptive immunity, and theory predicts that the optimal number of alleles is intermediate rather than maximal. Support for such an intermediate optimum in MHC allele count has mainly been demonstrated in studies on parasite load and reproductive traits. In this study we tested if an intermediate number of MHC class II (MHCII) alleles based on positively selected sites was correlated with an increased skin‐swelling response to two consecutive injections of phytohaemagglutinin (PHA) in bluethroat Luscinia svecica nestlings. We found that nestlings that were closer to the population mean in their number of alleles had a higher response to the first injection, supporting an intermediate, optimal number of alleles. After the second PHA injection, we found that the skin swelling was associated with MHCII allele count only at the among‐brood level, and with paternity (extra‐pair offspring had a larger swelling than within‐pair offspring). We therefore conclude that the PHA response is partly correlated with MHCII allele count, but that additional factors must be involved in both the PHA response and in female extra‐pair mate choice in this species.

Generation of hypoimmunogenic T cells from genetically engineered allogeneic human induced pluripotent stem cells.

Avoiding the immune rejection of transplanted T cells is central to the success of allogeneic cancer immunotherapies. One solution to protecting T-cell grafts from immune rejection involves the deletion of allogeneic factors and of factors that activate cytotoxic immune cells. Here we report the generation of hypoimmunogenic cancer-antigen-specific T cells derived from induced pluripotent stem cells (iPSCs) lacking β2-microglobulin, the class-II major histocompatibility complex (MHC) transactivator and the natural killer (NK) cell-ligand poliovirus receptor CD155, and expressing single-chain MHC class-I antigen E. In mouse models of CD20-expressing leukaemia or lymphoma, differentiated T cells expressing a CD20 chimeric antigen receptor largely escaped recognition by NKG2A+ and DNAM-1+ NK cells and by CD8 and CD4 T cells in the allogeneic recipients while maintaining anti-tumour potency. Hypoimmunogenic iPSC-derived T cells may contribute to the creation of off-the-shelf T cell immunotherapies.

CD4+ Memory T-Cell Formation during Type 1 Immune Responses.

Naive CD4+ T cells become memory cells after proliferating in response to their cognate major histocompatibility complex class II (MHCII)-bound peptide and passing through an effector cell stage. The process by which CD4+ memory T cells emerge from the effector cell pool, however, is less well understood than in the case of CD8+ T cells. During certain acute infections, naive CD4+ T cells proliferate and differentiate into various forms of type 1 (Th1) and follicular helper (Tfh) effector cells. We review the evidence that about 10% of the cells in each of these subsets survive to become memory cells that resemble their effector cell precursors. The roles that asymmetric cell division, the TCF-1 transcription factor, metabolic activity, reactive oxygen species, and the IL-7 receptor play in the effector to memory cell transition are discussed. We propose a speculative model in which the metabolic activity needed for rapid clonal expansion also generates toxic products that induce apoptosis in most effector cells. Memory cells then arise from the effector cells in each subset that are at the low end of the metabolic activity spectrum.

Improvement of the immunogenicity of ESAT-6 via fusion with the dodecameric protein dodecin of Mycobacterium tuberculosis

Tuberculosis (TB) is a chronic infectious disease that creates a heavy medical burden worldwide. The only approved vaccine, Bacillus Calmette-Guérin (BCG), cannot fully protect adolescents and adults from TB. Therefore, there is an urgent need to develop an effective new vaccine. Previous studies have found that dodecin, a flavin-binding protein of Mycobacterium tuberculosis (Mtb), can form stable dodecamers and has the potential to improve the immunogenicity of Mtb antigens. In this study, we constructed the fusion protein dodecin-ESAT-6 and evaluated the immunogenicity of dodecin, ESAT-6, and dodecin-ESAT-6 separately. Our results showed that dodecin-ESAT-6 is a dodecameric protein that can withstand heat at 95 °C and under SDS-PAGE conditions. Dodecin-ESAT-6 increased the expression of the costimulatory molecules CD80, CD86, and major histocompatibility complex class II (MHC-II) on the surface of RAW264.7 macrophages. Mice immunized with dodecin-ESAT-6 exhibited higher percentages of antigen-specific CD4+ and CD8+ T lymphocytes, higher levels of spleen lymphocyte proliferation and IFN-γ and IL-2 secretion, and a lower level of IL-4 secretion than those immunized with ESAT-6. The IgG, IgG1, and IgG2a titers of the dodecin-ESAT-6 group were significantly higher than those of the ESAT-6 group. Dodecin-ESAT-6 elicited a high IgG2a/IgG1 ratio and tended to produce a predominantly Th1-like response. These results support the conclusion that the dodecin-ESAT-6 dodecameric protein induced strong Th1 immune responses and improved the immunogenicity of ESAT-6, which provides a new strategy for TB vaccine development.

640 Single-cell transcriptomics of human pressure ulcers reveals MHCII expressing keratinocytes in patients with a worse healing outcome

Pressure ulcer (PU) is a chronic non-healing wound caused by continuous pressure of the bodyweight to the skin, which is often seen in spinal cord injury patients and the bedridden elderly population. Despite high mortality, the pathophysiology of PU remains poorly understood. Here we compared single-cell transcriptomic profiles on human epidermal cells from PU wound-edges with uninjured skin and acute wounds (AW) from healthy donors. We identified significant shifts of cellular composition and gene expression pattern in PU wound-edges, which could stratify PU into two groups. Interestingly, our study identified a subset of keratinocytes expressing major histocompatibility complex class II (MHCII), and these cells are enriched in patients with worse healing outcomes. We showed that IFNg in PU-derived wound fluid could induce MHCII expression in keratinocytes, and these wound fluid-treated keratinocytes inhibit autologous T cell activation. In line with this, we found that T cells from PUs enriched with MHCII+ keratinocytes produced less inflammatory cytokines. Together, our study provides a high-resolution molecular map of human PU compared to AW and the skin, which sheds new insights into the understanding of PU pathology and the future development of tailored wound therapy.

Tregitopes Improve Asthma by Promoting Highly Suppressive and Antigen-Specific Tregs.

Tregitopes (T regulatory epitopes) are IgG-derived peptides with high affinity to major histocompatibility complex class II (MHCII), that are known to promote tolerance by activating T regulatory cell (Treg) activity. Here we characterized the effect of IgG Tregitopes in a well-established murine model of allergic asthma, demonstrating in vivo antigen-specific tolerance via adoptive transfer of Tregitope-and-allergen-activated Tregs. Asthma is a heterogeneous chronic inflammatory condition affecting the airways and impacting over 300 million individuals worldwide. Treatment is suppressive, and no current therapy addresses immune regulation in severely affected asthmatics. Although high dose intra-venous immunoglobulin (IVIg) is not commonly used in the asthma clinic setting, it has been shown to improve severe asthma in children and in adults. In our laboratory, we previously demonstrated that IVIg abrogates airway hyperresponsiveness (AHR) in a murine model of asthma and induces suppressive antigen-specific T-regulatory cells. We hypothesized that IgG-derived Tregitopes would modulate allergic airway disease by inducing highly suppressive antigen-specific Tregs capable of diminishing T effector cell responses and establishing antigen-specific tolerance. Using ovalbumin (OVA-) and ragweed-driven murine models of allergic airway disease, we characterized the immunoregulatory properties of Tregitopes and performed Treg adoptive transfer to OVA- and ragweed-allergic mice to test for allergen specificity. Treatment with Tregitopes attenuated allergen-induced airway hyperresponsiveness and lung inflammation. We demonstrated that Tregitopes induce highly suppressive allergen-specific Tregs. The tolerogenic action of IgG Tregitopes in our model is very similar to that of IVIg, so we foresee that IgG Tregitopes could potentially replace steroid-based treatment and can offer a synthetic alternative to IVIg in a range of inflammatory and allergic conditions.

Divergent Functions of Tissue-Resident and Blood-Derived Macrophages in the Hemorrhagic Brain.

Brain tissue-resident microglia and monocyte-derived macrophages (MDMs) are innate immune cells that contribute to the inflammatory response, phagocytosis of debris, and tissue repair after injury. We have previously reported that both microglia and MDMs transition from proinflammatory to reparative phenotypes over days after an intracerebral hemorrhage (ICH). However, their individual functional properties in the brain remain largely unknown. Here we characterized the differences between microglia and MDMs and further elucidate their distinct activation states and functional contributions to the pathophysiology and recovery after ICH. Autologous blood injection was used to model ICH in mice. Longitudinal transcriptomic analyses on isolated microglia and MDMs from mice at days 1, 3, 7 and 10 after ICH and naive controls identified core transcriptional programs that distinguish these cells. Imaging flow cytometry and in vivo phagocytosis assays were used to study phagocytic ability of microglia and MDMs. Antigen presentation was evaluated by ovalbumin-OTII CD4 T-cell proliferation assays with bone marrow–derived macrophages and primary microglia cultures. MDMs had higher phagocytic activity and higher erythrophagocytosis in the ICH brain. Differential gene expression revealed distinct transcriptional signatures in the MDMs and microglia after ICH. MDMs had higher expression of MHCII (major histocompatibility complex class II) genes than microglia at all time points and greater ability to induce antigen-specific T-cell proliferation. The different ontogeny of microglia and MDMs lead to divergent responses and functions in the inflamed brain as these 2 cell populations differ in phagocytic functions and antigen-presenting capabilities in the brain after ICH.

CD74 regulates cellularity and maturation of medullary thymic epithelial cells partially by activating the canonical NF-κB signaling pathway.

Thymic epithelial cells (TECs) are indispensable for T cell development, T cell receptor (TCR) repertoire selection, and specific lineage differentiation. Medullary thymic epithelial cells (mTECs), which account for the majority of TECs in adults, are critical for thymocyte selection and self-tolerance. CD74 is a nonpolymorphic transmembrane glycoprotein of major histocompatibility complex class II (MHCII) that is expressed in TECs. However, the exact role of CD74 in regulating the development of mTEC is poorly defined. In this research, we found that loss of CD74 resulted in a significant diminution in the medulla, a selective reduction in the cell number of mature mTECs expressing CD80 molecules, which eventually led to impaired thymic CD4+ T cell development. Moreover, RNA-sequence analysis showed that CD74 deficiency obviously downregulated the canonical nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway in mTECs. Our results suggest that CD74 positively controls mTEC cellularity and maturation partially by activating the canonical NF-κB signaling pathway.