MHC Class II Research Articles

Expansion of MHC-IIB has constrained the evolution of MHC-IIA in passerines.

The major histocompatibility complex (MHC) is central in adaptive immunity, with the highly polymorphic MHC genes encoding antigen-presenting molecules. Two MHC class II (MHC-II) loci, DA1 and DA2, pre-date the radiation of extant birds and persist throughout much of the avian phylogeny. Within each locus, the MHC-II molecules are encoded by A- (DAA) and B- (DAB) genes, which are arranged in A-B dyads. However, in passerines (order Passeriformes), the DA2 locus has been lost and the ancestral A-B dyad at the DA1 locus has been replaced by a putatively single A-gene (DAA1) and an array of highly polymorphic B-genes (DAB1). In this study, we genotyped the DAA1 gene of 15 passerine species and confirmed that passerines possess just one copy of DAA1. We then compared selection patterns in DAA1 between passerines and non-passerines and found that exon 2, which encodes the antigen-presenting domain, has been subject to weaker positive selection and stronger negative selection in passerines compared with non-passerines. Additional comparisons showed that the patterns of selection in the passerine DAA1 gene are unlikely to be related to the loss of the DA2 locus. Instead, our findings suggest that the expansion of DAB1 (MHC-IIB) has imposed an evolutionary constraint on the passerine DAA1 (MHC-IIA) gene. We speculate that this constraint may be the result of each DAA1 chain having to form heterodimers with many different DAB1 chains.

New dietary trends and alternative proteins: the emergence of novel food allergens.

New dietary trends driven by environmental and health considerations will undoubtedly lead to the emergence of novel food allergens. Assessment of the allergenic risk of new or modified protein-containing food sources and ingredients, as well as surveillance of emerging food allergies, is then required. Developments of in silico and in vitro models apprehending protein capacity to cross-react with other homologous proteins and to induce a de novo allergic sensitization are ongoing to better integrate multiple parameters such as 3D structural information or major histocompatibility complex class II (MHC-II) presentation propensity. However, the effects of food matrices and food processing still need to be addressed in these models. Consequently, clinical and postmarket surveillance remain of critical importance to alert on emergent food allergies, which are modulated by regional dietary practices. Monitoring of the emergence of food allergens requires close collaborations between allergologists, consumers, patient associations and food safety authorities. We also need to get a consensus on an acceptable level of allergenic risk that offers the possibility to develop and market innovative and sustainable food products.

Palaeognaths Reveal Evolutionary Ancestry of the Avian Major Histocompatibility Complex Class II.

The multigene family of the major histocompatibility complex (MHC) codes for the key antigen-presenting molecules of the vertebrate immune system. In birds, duplicated MHC class II (MHC-II) genes are highly homogenized by concerted evolution, and thus, identification of their orthologous relationships across long evolutionary timescales remains challenging. Relatively low evolutionary rate of avian MHC class IIA genes has been expected to provide a promising avenue to allow such inferences, but availability of MHC-IIA sequences in nonmodel bird species has been limited until recently. Here, taking advantage from accumulating genomic resources, we identified and analyzed MHC-IIA sequences from the most basal lineage of extant birds (Palaeognathae). Conserved region of the MHC-IIA membrane-proximal domain was used to search for orthologous relationships between palaeognath birds and nonavian reptiles. First, analyses of palaeognath sequences revealed the presence of a separate MHC-IIA gene lineage (DAA3) in kiwis, which did not cluster with previously described avian MHC-IIA lineages (DAA1 and DAA2). Next, phylogenetic reconstruction showed that kiwi DAA3 sequences form a single well-supported cluster with turtle MHC-IIA. High similarity of these sequences most likely reflects their remarkable evolutionary conservation and retention of ancient orthologous relationships, which can be traced back to basal archosauromorphs ca. 250 million years ago. Our analyses offer novel insights into macroevolutionary history of the MHC and reinforce the view that rapid accumulation of high-quality genome assemblies across divergent nonmodel species can substantially advance our understanding of gene evolution.

Graft survival of major histocompatibility complex deficient stem cell-derived retinal cells

BackgroundGene editing of immunomodulating molecules is a potential transplantation strategy to control immune rejection. As we noticed the successful transplantation of retinal pigment epithelium (RPE) derived from embryonic stem cells of a cynomolgus monkey that accidentally lacked MHC class II (MHC-II) molecules, we hypothesized immune rejection could be evaded by suppressing MHC-II.MethodsGene editing by the Crispr/Cas9 system was performed in induced pluripotent stem cells derived from a cynomolgus monkey (miPSCs) for targeted deletion of the gene coding class II MHC trans-activator (CIITA). Then the CIITA-knocked out miPSCs were differentiated into RPE cells to generate miPSC-derived MHC-II knockout RPE. The MHC-II knockout or wild-type RPEs were transplanted into the eyes of healthy cynomolgus monkeys. All monkeys used in this study were male.ResultsHere we show when MHC-II knockout RPE are transplanted into monkey eyes, they show suppressed immunogenicity with no infiltration of inflammatory cells, leading to successful engraftment.ConclusionsOur results reasonably evidence the efficacy of MHC-II knockout iPSC-RPE transplants for clinical application.

Nano-selenium ameliorates microplastics-induced injury: Histology, antioxidant capacity, immunity and intestinal microbiota of grass carp (Ctenopharyngodon idella)

Microplastics (MPs) are pollutants widely distributed in the aquatic environments and causing various degrees of aquatic toxicity to aquatic organisms, which has attracted global attention in recent years. Nano-selenium (NSe) has been shown to have the potential to mitigate the harmful impacts of toxic substances. However, there is currently no reported evidence regarding the protective influence of NSe against the adverse effects of MPs. The aim of this study is to determine whether NSe could ameliorate the polystyrene (PS)-MPs-induced injury in grass carp (Ctenopharyngodon idella). The individuals of grass carp were assigned into three groups: (1) the control group fed with basal diet, (2) the PS group fed with basal diet and exposed to PS-MPs, and (3) the NSe group fed with diet supplemented with NSe and exposed to PS-MPs. Our results indicated that NSe administration significantly alleviated the histological damage caused by the PS-MPs in the liver and intestine with lower goblet cell count and larger villus height in the intestine, and significantly lower damage score in the liver. Moreover, NSe mitigated PS-MPs-induced oxidative stress through restoring the activities of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA)) except the intestinal CAT activity. Furthermore, NSe supplementation could help fish maintain lower transcriptional level of the immune-related genes (Toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 (MyD88)), inflammation-related genes (major histocompatibility complex class II (MHC-II) and interleukin 8 (IL-8)) and antioxidant enzyme-related genes (nuclear factor (erythroid-derived 2)-like 2 (Nrf-2) and kelch-like ECH-associated protein 1 (Keap-1)) after PS-MPs exposure. Besides, NSe supplementation dramatically helped maintain the intestinal microbial composition, for example, the proportion of Proteobacteria in the grass carp intestine of the NSe group (41 %) was similar to that of the control group (34 %) while 85 % of the PS group. NSe also played a significant protective role in intestinal microbial diversity, effectively resisting the damage on intestinal microbial diversity due to PS-MPs exposure. PS-MPs reduced the beneficial bacteria and increased the pathogenic microorganism like Aeromonas, which was undeniable signs of intestinal dysbiosis. Functional analysis indicated that PS-MPs affected intestinal microbiota functions like inhibition of metabolism, while NSe could significantly alleviate the damage. Our findings suggested that NSe could ameliorate PS-MPs-induced injury, which could contribute to the better understanding of the ecotoxicological effects of MPs on fish and help develop relevant mitigation strategies.

PDCD4 deficiency in hepatocytes exacerbates nonalcoholic steatohepatitis through enhanced MHC class II transactivator expression

Nonalcoholic steatohepatitis (NASH) is a primary cause of liver cirrhosis and hepatocellular carcinoma, presenting a significant and unmet medical challenge. The necessity to investigate the molecular mechanisms underlying NASH is highlighted by the observed decrease in programmed cell death 4 (PDCD4) expression in NASH patients, suggesting that PDCD4 may play a protective role in maintaining liver health. In this study, we identify PDCD4 as a natural inhibitor of NASH development in mice. The absence of PDCD4 leads to the spontaneous progression of NASH. Notably, PDCD4-deficient hepatocytes display elevated major histocompatibility complex class II (MHCII) expression due to CIITA activation, indicating that PCDC4 prevents the abnormal transformation of hepatocytes into antigen-presenting cells (APCs). Cell co-culture experiments reveal that hepatocytes lacking PDCD4, which resemble APCs, can directly activate CD4+ T cells by presenting multiple peptides, resulting in the release of inflammatory factors. Additionally, both cellular and animal studies show that CIITA promotes lipid accumulation in hepatocytes and exacerbates NASH progression. In summary, our findings reveal a novel role of PDCD4 in regulating CIITA and MHCII expression during NASH development, offering new therapeutic approaches for NASH treatment.

Neoself-antigens are the primary target for autoreactive T cells in human lupus

Major histocompatibility complex class II (MHC-II) is the most significant genetic risk factor for systemic lupus erythematosus (SLE), but the nature of the self-antigens that trigger autoimmunity remains unclear. Unusual self-antigens, termed neoself-antigens, are presented on MHC-II in the absence of the invariant chain essential for peptide presentation. Here, we demonstrate that neoself-antigens are the primary target for autoreactive Tcells clonally expanded in SLE. When neoself-antigen presentation was induced by deleting the invariant chain in adult mice, neoself-reactive Tcells were clonally expanded, leading to the development of lupus-like disease. Furthermore, we found that neoself-reactive CD4+ Tcells were significantly expanded in SLE patients. A high frequency of Epstein-Barr virus reactivation is a risk factor for SLE. Neoself-reactive lupus Tcells were activated by Epstein-Barr-virus-reactivated cells through downregulation of the invariant chain. Together, our findings imply that neoself-antigen presentation by MHC-II plays a crucial role in the pathogenesis of SLE.

Structural basis for mouse LAG3 interactions with the MHC class II molecule I-Ab

The immune checkpoint protein, Lymphocyte activation gene-3 (LAG3), binds Major Histocompatibility Complex Class II (MHC-II) and suppresses T cell activation. Despite the recent FDA approval of a LAG3 inhibitor for the treatment of melanoma, how LAG3 engages MHC-II on the cell surface remains poorly understood. Here, we determine the 3.84 Å-resolution structure of mouse LAG3 bound to the MHC-II molecule I-Ab, revealing that domain 1 (D1) of LAG3 binds a conserved, membrane-proximal region of MHC-II spanning both the α2 and β2 subdomains. LAG3 dimerization restricts the intermolecular spacing of MHC-II molecules, which may attenuate T cell activation by enforcing suboptimal signaling geometry. The LAG3-MHC-II interface overlaps with the MHC-II-binding site of the T cell coreceptor CD4, implicating disruption of CD4-MHC-II interactions as a mechanism for LAG3 immunosuppressive function. Lastly, antibody epitope analysis indicates that multiple LAG3 inhibitors do not recognize the MHC-II-binding interface of LAG3, suggesting a role for functionally distinct mechanisms of LAG3 antagonism in therapeutic development.

Deciphering the cellular landscape and potential targets of nasopharyngeal and oral cancers using single-cell RNA sequencing.

Cellular heterogeneity in nasopharyngeal cancer (NPC) and oral cancer remains unclear. In the current study, using single-cell RNA sequencing techniques, we investigated the cellular landscape in NPC and oral cancers. We identified a diverse range of cell types within the tumor microenvironment (TME) and variations in cell infiltration between NPC and oral cancer. In oral cancer, we observed a predominant infiltration of epithelial cells, fibroblasts, and endothelial cells (ECs), while T cells were the main infiltrating cell population in NPCs. We further classified these infiltrating cells into subclusters. Additionally, we observed complex interactions among cells that led to distinct trajectories. In particular, a unique epithelial subcluster with high expression of major histocompatibility complex class II (MHC-II) molecules was correlated with a favorable outcome and infiltration of CD4+ T cells. In addition, MHC-II+ epithelial cells inhibited mouse tumor growth and promoted T-cell infiltration. Consequently, our findings provide a deep understanding of the TME showing a significant prognostic value and therapeutic potential.

The PDIA3-STAT3 protein complex regulates IBS formation and development via CTSS/MHC-II pathway-mediated intestinal inflammation

Irritable bowel syndrome (IBS) is a persistent functional gastrointestinal disorder characterised by abdominal pain and altered patterns of defecation. This study aims to clarify an increase in the expression and interaction of protein disulfide-isomerase A3 (PDIA3) and Signal Transducer and Activator of Transcription 3 (STAT3) within the membrane of dendritic cells (DCs) from individuals with IBS. Mechanistically, the heightened interaction between PDIA3 and STAT3 at the DC membrane results in reduced translocation of phosphorylated STAT3 (p-STAT3) into the nucleus. The reduction of p-STAT3 to nuclear transport subsequently increased the levels of cathepsin S (CTSS) and major histocompatibility complex class II (MHC-II). Consequently, activated DCs promote CD4+ T cell proliferation and cytokine secretion, including interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-9 (IL-9), and tumour necrosis factor-alpha (TNF-α), thereby contributing to the development of IBS. Importantly, the downregulation of PDIA3 and the administration of punicalagin (Pun), a crucial active compound found in pomegranate peel, alleviate IBS symptoms in rats, such as increased visceral hypersensitivity and abnormal stool characteristics. Collectively, these findings highlight the involvement of the PDIA3-STAT3 protein complex in IBS, providing a novel perspective on the modulation of immune and inflammatory responses. Additionally, this research advances our understanding of the role and mechanisms of PDIA3 inhibitors, presenting new therapeutic possibilities for managing IBS.

IFNh and IRF9 influence the transcription of MHCII mediated by IFNγ to maintain immune balance in sea perch lateolabrax japonicus

The major histocompatibility complex class II (MHCII) molecules are crucial elements of the adaptive immune system, essential for orchestrating immune responses against foreign pathogens. However, excessive expression of MHCII can disrupt normal physiological functions. Therefore, the host employs various mechanisms to regulate MHCII expression and maintain immune homeostasis. Despite this importance, limited studies have explored the negative regulation of MHCII transcription in bony fish. In this study, we found that interferon h (IFNh), a subtype of type I IFN in sea perch Lateolabrax japonicus, could inhibit the activation of IFNγ induced-MHCII expression by modulating the transcription of the class II major histocompatibility complex transactivator (CIITA). Transcriptome analysis revealed 57 up-regulated and 69 down-regulated genes in cells treated with both IFNγ and IFNh compared to those treated with IFNγ alone. To maintain cellular homeostasis, interferon regulatory factor 9 (IRF9) was up-regulated following IFNγ stimulation, thereby preventing MHCII overexpression. Mechanistically, IRF9 bound to the CIITA promoter and suppressed its expression activated by IRF1. Furthermore, IRF9 inhibited the promoter activity of both MHCII-α and MHCII-β induced by CIITA. Our findings highlight the roles of IFNh and IRF9 as suppressors regulating MHCII expression at different hierarchical levels. This study provides insights into the intricate regulation of antigen presentation and the foundation for further exploration of the interaction mechanisms between aquatic virus and fish.

Structure, function, and immunomodulation of the CD8 co-receptor.

Expressed on the surface of CD8+ T cells, the CD8 co-receptor is a key component of the T cells that contributes to antigen recognition, immune cell maturation, and immune cell signaling. While CD8 is widely recognized as a co-stimulatory molecule for conventional CD8+ αβ T cells, recent reports highlight its multifaceted role in both adaptive and innate immune responses. In this review, we discuss the utility of CD8 in relation to its immunomodulatory properties. We outline the unique structure and function of different CD8 domains (ectodomain, hinge, transmembrane, cytoplasmic tail) in the context of the distinct properties of CD8αα homodimers and CD8αβ heterodimers. We discuss CD8 features commonly used to construct chimeric antigen receptors for immunotherapy. We describe the molecular interactions of CD8 with classical MHC-I, non-classical MHCs, and Lck partners involved in T cell signaling. Engineered and naturally occurring CD8 mutations that alter immune responses are discussed. The applications of anti-CD8 monoclonal antibodies (mABs) that target CD8 are summarized. Finally, we examine the unique structure and function of several CD8/mAB complexes. Collectively, these findings reveal the promising immunomodulatory properties of CD8 and CD8 binding partners, not only to uncover basic immune system function, but to advance efforts towards translational research for targeted immunotherapy.

Gluten-Dependent Activation of CD4+ T Cells by MHC Class II–Expressing Epithelium

Intestinal epithelial cell (IEC) damage is a hallmark of celiac disease (CeD); however, its role in gluten-dependent T-cell activation is unknown. We investigated IEC-gluten-T-cell interactions in organoid monolayers expressing human major histocompatibility complex class II (HLA-DQ2.5), which facilitates gluten antigen recognition by CD4+ T cells in CeD. Epithelial major histocompatibility complex class II (MHCII) was determined in active and treated CeD, and in nonimmunized and gluten-immunized DR3-DQ2.5 transgenic mice, lacking mouse MHCII molecules. Organoid monolayers from DR3-DQ2.5 mice were treated with or without interferon (IFN)-γ, and MHCII expression was evaluated by flow cytometry. Organoid monolayers and CD4+ T-cell co-cultures were incubated with gluten, predigested, or not by elastase-producing Pseudomonas aeruginosa or its lasB mutant. T-cell function was assessed based on proliferation, expression of activation markers, and cytokine release in the co-culture supernatants. Patients with active CeD and gluten-immunized DR3-DQ2.5 mice demonstrated epithelial MHCII expression. Organoid monolayers derived from gluten-immunized DR3-DQ2.5 mice expressed MHCII, which was upregulated by IFN-γ. In organoid monolayer T-cell co-cultures, gluten increased the proliferation of CD4+ T cells, expression of T-cell activation markers, and the release of interleukin-2, IFN-γ, and interleukin-15 in co-culture supernatants. Gluten metabolized by P aeruginosa, but not the lasB mutant, enhanced CD4+ T-cell proliferation and activation. Gluten antigens are efficiently presented by MHCII-expressing IECs, resulting in the activation of gluten-specific CD4+ T cells, which is enhanced by gluten predigestion with microbial elastase. Therapeutics directed at IECs may offer a novel approach for modulating both adaptive and innate immunity in patients with CeD.

Polygenic TB control and the sequence of innate/adaptive immune responses to infection: MHC-II alleles determine the size of the S100A8/9-producing neutrophil population.

Among several quantitative trait loci involved in tuberculosis (TB) control in mice, one was mapped within the chromosome 17 segment occupied by the H2 complex and another within the chromosome 3 segment comprising the S100A8/9 genes, which encode neutrophil inflammatory factor S100A8/9. Previously, we developed a panel of H2-congenic mouse strains differing by small segments of the major histocompatibility complex Class II (MHC-II) region from TB-susceptible H2j mice transferred onto the genetic background of the TB-resistant C57BL/6 (H2b) strain. Susceptible B6.I-9.3 mice differ from B6 progenitors by the alleles of their only classical MHC-II H2-Aβ gene. The goals of the present study were to: (i) comprehensively characterise the differences in TB-related phenotypes between mice of the two strains and (ii) decipher interactions between the H2-Aβ and S100A8/9 genes. Here, we describe the dynamics of TB-related phenotypes differentiating B6.I-9.3 and B6 mice (colony forming unitscounts, histopathology, lung immune cell infiltration and cytokine profiles). We show that disproportionally diminished CD4+ T-cell population, an enlarged S100A8/9-positive neutrophil population and higher S100A8/9 serum levels in B6.I-9.3 mice collectively form the 'susceptible' phenotype before infection. An increase in IL-17 and a decrease in intrferon-gamma production by CD4+ T-cells in these mice provide a mechanistic explanation of this phenotype. Using F2 segregation analysis, we show that the number of S100A8/9-producing neutrophils in lungs and spleens and the proportion of Th17 CD4+ T-cells in lungs are significantly lower in the presence of the MHC-II dominant 'resistant' b allele compared to the recessive 'susceptible' j/j genotype. This provides direct genetic evidence that MHC-II-regulated CD4+ T-cell landscapes determine neutrophil abundance before infection, an important pathogenic factor in TB immunity.

Cutting Edge: Redundant Roles for MHC Class II-, CD1d-, and MR1-restricted T Cells in Clearing Bartonella Infection.

The importance of unconventional T cells for mucosal immunity is firmly established but for systemic bacterial infection remains less well defined. In this study, we explored the role of various T cell subsets in murine Bartonella infection, which establishes persistent bacteremia unless controlled by antibacterial Abs. We found that αβ T cells are essential for Ab production against and clearance of B. taylorii, whereas MHC class I (MHC-I)- or MHC class II (MHC-II)-deficient mice eliminated B. taylorii infection with normal kinetics. Similarly, animals lacking either CD1d or MR1 suppressed bacteremia with normal kinetics. Interestingly, mice with a combined deficiency of either MHC-II and CD1d or MHC-II and MR1 failed to clear the infection, indicating that the combination of CD1d- and MR1-restricted T cells can compensate for the lack of MHC-II in this model. Our data document a previously underappreciated contribution of unconventional T cells to the control of systemic bacterial infection, supposedly as helper cells for antibacterial Ab production.

IFNγ mediates the resistance of tumor cells to distinct NK cell subsets

BackgroundImmune checkpoint blockade targeting the adaptive immune system has revolutionized the treatment of cancer. Despite impressive clinical benefits observed, patient subgroups remain non-responsive underscoring the necessity for combinational therapies harnessing...

Traumatic brain injury alters the effects of class II invariant peptide (CLIP) antagonism on chronic meningeal CLIP + B cells, neuropathology, and neurobehavioral impairment in 5xFAD mice

BackgroundTraumatic brain injury (TBI) is a significant risk factor for Alzheimer’s disease (AD), and accumulating evidence supports a role for adaptive immune B and T cells in both TBI and AD pathogenesis. We previously identified B cell and major histocompatibility complex class II (MHCII)-associated invariant chain peptide (CLIP)-positive B cell expansion after TBI. We also showed that antagonizing CLIP binding to the antigen presenting groove of MHCII after TBI acutely reduced CLIP + splenic B cells and was neuroprotective. The current study investigated the chronic effects of antagonizing CLIP in the 5xFAD Alzheimer’s mouse model, with and without TBI.Methods12-week-old male wild type (WT) and 5xFAD mice were administered either CLIP antagonist peptide (CAP) or vehicle, once at 30 min after either sham or a lateral fluid percussion injury (FPI). Analyses included flow cytometric analysis of immune cells in dural meninges and spleen, histopathological analysis of the brain, magnetic resonance diffusion tensor imaging, cerebrovascular analysis, and assessment of motor and neurobehavioral function over the ensuing 6 months.Results9-month-old 5xFAD mice had significantly more CLIP + B cells in the meninges compared to age-matched WT mice. A one-time treatment with CAP significantly reduced this population in 5xFAD mice. Importantly, CAP also improved some of the immune, histopathological, and neurobehavioral impairments in 5xFAD mice over the ensuing six months. Although FPI did not further elevate meningeal CLIP + B cells, it did negate the ability of CAP to reduce meningeal CLIP + B cells in the 5xFAD mice. FPI at 3 months of age exacerbated some aspects of AD pathology in 5xFAD mice, including further reducing hippocampal neurogenesis, increasing plaque deposition in CA3, altering microgliosis, and disrupting the cerebrovascular structure. CAP treatment after injury ameliorated some but not all of these FPI effects.

Cytotoxic CD4+ T Cells Are Induced during Infection with Chlamydia trachomatis.

Chlamydia trachomatis is the most common cause of bacterial sexually transmitted infection in both men and women. Immunity to C. trachomatis involves many cell types, but CD4+ T cells play a key role in protecting the host during natural infection. Specifically, IFN-γ production by CD4+ T cells is the main effector responsible for bacterial clearance, yet the exact mechanism by which IFN-γ confers protection is poorly defined. In our efforts to define the specific mechanisms for bacterial clearance, we now show that IFN-γ upregulates expression of MHC class II (MHCII) on nonhematopoietic cells during C. trachomatis infection invivo. We also find that MHCII expression on epithelial cells of the upper genital tract contributes to the efficient clearance of bacteria mediated by pathogen-specific CD4+ Th1 cells. As we further cataloged the protective mechanisms of C. trachomatis-specific CD4+ T cells, we found that the T cells also express granzyme B (GzmB) when coincubated with infected cells. In addition, during C. trachomatis infection of mice, primed activated-naive CD4+ Th1 cells displayed elevated granzyme transcripts (GzmA, GzmB, GzmM, GzmK, GzmC) compared with memory CD4+ T cells invivo. Finally, using intracellular cytokine staining and a GzmB-/- mouse strain, we show that C. trachomatis-specific CD4+ Th1 cells express GzmB upon Ag stimulation, and that this correlates with Chlamydia clearance invivo. Together these results have led us to conclude that Chlamydia-specific CD4+ Th1 cells develop cytotoxic capacity through engagement with nonhematopoietic MHCII, and this correlates to C. trachomatis clearance.

A monoclonal antibody that recognizes a unique 13-residue epitope in the cytoplasmic tail of HLA-E

The Class I MHC molecule (MHC-I) HLA-E presents peptides that are derived from the signal sequences, either those of other MHC-I products, or of viral type I membrane glycoproteins. Monoclonal antibodies with proven specificity for HLA-E, and with no cross-reactions with other MHC-I products, have yet to be described. To obtain anti-HLA-E-specific antibodies suitable for a range of applications, we generated monoclonal antibodies against a unique feature of HLA-E: its cytoplasmic tail. We created an immunogen by performing an enzymatically catalyzed transpeptidation reaction to obtain a fusion of the cytoplasmic tail of HLA-E with a nanobody that recognizes murine Class II MHC (MHC-II) products. We obtained a mouse monoclonal antibody that recognizes a 13-residue stretch in the HLA-E cytoplasmic tail. We cloned the genes that encode this antibody in expression vectors to place an LPETG sortase recognition motif at the C-terminus of the heavy and light chains. This arrangement allows the site-specific installation of fluorophores or biotin at these C-termini. The resulting immunoglobulin preparations, labeled with 4 equivalents of a fluorescent or biotinylated payload of choice, can then be used for direct immunofluorescence or detection of the tag by fluorescence or by streptavidin-based methods. We also show that the 13-residue sequence can serve as an epitope tag, independent of the site of its placement within a protein’s sequence. The antibody can be used diagnostically to stain for HLA-E on patient tumor samples, it can be used as an antibody-epitope tag for extracellular proteins, and it enables research into the unique role of the cytoplasmic tail of HLA-E.

Andrographolide exhibits antinociceptive effects in neuropathic rats via inhibiting class Ⅱ MHC associated response and regulating synaptic plasticity

BackgroundNeuropathic pain (NP) due to nerve injury, disrupts neural plasticity by triggering the release of inflammatory mediators. Alongside the hypothesis that neuro-inflammation contributes to this disruption, Andrographolide (Andro), a traditional bioactive compound derived from Andrographis paniculata, has garnered attention for its potent anti-inflammatory properties. However, whether Andro could ameliorate NP by regulating neuroinflammation remains unknown. PurposeThis study aimed to investigate whether and how Andro regulates neuroinflammation and alleviates NP. MethodsThe analgesic effects of Andro on NP were evaluated using both the spinal nerve ligation (SNL) and formalin rat models. A combination of network pharmacology, RNA sequencing, and experimental validation was employed to elucidate the underlying mechanism behind Andro's analgesic effects. Additionally, various techniques such as functional ultrasound, immunohistochemistry, quantitative real-time polymerase chain reaction (qPCR), patch clamp, and electron microscopy were employed to investigate the specific neural cell types, neural functions, and changes in neural plasticity influenced by Andro. ResultsNetwork pharmacology analysis unveiled the crucial roles played by shared targets of Andro and pain in regulating pain-related inflammation, including microglia activation, neuroinflammation, immune modulation, and synaptic transmission. Furthermore, we confirmed Andro's superior efficacy in pain relief compared to the traditional analgesic drug, Gabapentin. In these models, Andro was observed to modulate the haemodynamic response triggered by SNL. Transcriptome analysis and molecular docking studies indicated the involvement of major histocompatibility complex class II (MHCII) genes (Db1, Da, and Bb). Electron microscopy revealed improvements in synaptic ultrastructure, and electrophysiological investigations showed a selective reduction in glutamatergic transmission in neuropathic rats after following Andro treatment. The integration of systems pharmacology analysis and biological validation collectively demonstrated that the mechanism of pain relief involves immune modulation, enhancement of synaptic plasticity, and precise regulation of excitatory neurotransmission. ConclusionIn conclusion, this study has demonstrated that Andro, by targeting MHCII genes, may serve as a promising therapeutic candidate for neuropathic pain.