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IntroductionAltered immune signatures are emerging as a central theme in neurodegenerative disease, yet little is known about immune responses in early-onset Alzheimer's disease (EOAD).MethodsWe examined single-cell RNA-sequencing (scRNA-seq) data from peripheral blood mononuclear cells (PBMCs) and droplet digital polymerase chain reaction (ddPCR) data from CD4 T cells from participants with EOAD and clinically normal controls.ResultsWe analyzed PBMCs from 16 individuals by scRNA-seq and discovered increased interferon signaling-associated gene (ISAG) expression and striking expansion of antiviral-like ISAGhi T cells in EOAD. Isolating CD4 T cells from 19 individuals, including four cases analyzed by scRNA-seq, we confirmed increased expression of ISAGhi marker genes. Publicly available cerebrospinal fluid leukocyte scRNA-seq data from late-onset mild cognitive impairment and AD also revealed increased expression of interferon-response genes.DiscussionAntiviral-like ISAGhi T cells are expanded in EOAD. Additional research into these cells and the role of heightened peripheral IFN signaling in neurodegeneration is warranted.HighlightsInterferon-responsive T cells expanded in early-onset Alzheimer's disease (AD). Increased interferon-associated gene expression present in early- and late-onset AD. Peripheral immune changes in T and NK cells driven by females with early-onset AD.

Inflammatory neuropathies, which include chronic inflammatory demyelinating polyneuropathy (CIDP) and Guillain Barré syndrome (GBS), result from autoimmune destruction of the PNS and are characterized by progressive weakness and sensory loss. CD4+ T cells play a key role in the autoimmune destruction of the PNS. Yet, key properties of pathogenic CD4+ T cells remain incompletely understood. Here, we used paired single-cell RNA-Seq (scRNA-Seq) and single-cell T cell receptor-sequencing (scTCR-Seq) of peripheral nerves from an inflammatory neuropathy mouse model to identify IL-21-expressing CD4+ T cells that were clonally expanded and multifunctional. These IL-21-expressing CD4+ T cells consisted of 2 transcriptionally distinct expanded cell populations, which expressed genes associated with T follicular helper (Tfh) and T peripheral helper (Tph) cell subsets. Remarkably, TCR clonotypes were shared between these 2 IL-21-expressing cell populations, suggesting a common lineage differentiation pathway. Finally, we demonstrated that IL-21 receptor-KO (IL-21R-KO) mice were protected from neuropathy development and had decreased immune infiltration into peripheral nerves. IL-21 signaling upregulated CXCR6, a chemokine receptor that promotes CD4+ T cell localization in peripheral nerves. Together, these findings point to IL-21 signaling, Tfh/Tph differentiation, and CXCR6-mediated cellular localization as potential therapeutic targets in inflammatory neuropathies.

CD4+ T cells survey and maintain immune homeostasis in the brain, yet their differentiation states and functional capabilities remain unclear. Our approach, combining single-cell transcriptomic analysis, ATAC-Seq, spatial transcriptomics, and flow cytometry, revealed a distinct subset of CCR7+ CD4+ T cells resembling lymph node central memory (TCM) cells. We observed chromatin accessibility at the CCR7, CD28, and BCL-6 loci, defining molecular features of TCM. Brain CCR7+ CD4+ T cells exhibited recall proliferation and interleukin-2 production ex vivo, showcasing their functional competence. We identified the skull bone marrow as a local niche for these cells alongside CNS border tissues. Sequestering TCM cells in lymph nodes using FTY720 led to reduced CCR7+ CD4+ T cell frequencies in the cerebrospinal fluid, accompanied by increased monocyte levels and soluble markers indicating immune activation. In macaques chronically infected with SIVCL757 and experiencing viral rebound due to cessation of antiretroviral therapy, a decrease in brain CCR7+ CD4+ T cells was observed, along with increased microglial activation and initiation of neurodegenerative pathways. Our findings highlight a role for CCR7+ CD4+ T cells in CNS immune surveillance, and their decline during chronic SIV highlights their responsiveness to neuroinflammation.

Immune checkpoint blockade (ICB) with PD-1/PD-L1 inhibition has revolutionized the treatment of non-small cell lung cancer (NSCLC). Durable responses, however, are observed only in a subpopulation of patients. Defective antigen presentation and an immunosuppressive tumor microenvironment (TME) can lead to deficient T cell recruitment and ICB resistance. We evaluate intratumoral (IT) vaccination with CXCL9- and CXCL10-engineered dendritic cells (CXCL9/10-DC) as a strategy to overcome resistance. IT CXCL9/10-DC leads to enhanced T cell infiltration and activation in the TME and tumor inhibition in murine NSCLC models. The antitumor efficacy of IT CXCL9/10-DC is dependent on CD4+ and CD8+ T cells, as well as CXCR3-dependent T cell trafficking from the lymph node. IT CXCL9/10-DC, in combination with ICB, overcomes resistance and establishes systemic tumor-specific immunity in murine models. These studies provide a mechanistic understanding of CXCL9/10-DC-mediated host immune activation and support clinical translation of IT CXCL9/10-DC to augment ICB efficacy in NSCLC.

BACKGROUNDPrecise stratification of patients with non-small cell lung cancer (NSCLC) is needed for appropriate application of PD-1/PD-L1 blockade therapy.METHODSWe measured soluble forms of the immune-checkpoint molecules PD-L1, PD-1, and CTLA-4 in plasma of patients with advanced NSCLC before PD-1/PD-L1 blockade. A prospective biomarker-finding trial (cohort A) included 50 previously treated patients who received nivolumab. A retrospective observational study was performed for patients treated with any PD-1/PD-L1 blockade therapy (cohorts B and C), cytotoxic chemotherapy (cohort D), or targeted therapy (cohort E). Plasma samples from all patients were assayed for soluble immune-checkpoint molecules with a highly sensitive chemiluminescence-based assay.RESULTSNonresponsiveness to PD-1/PD-L1 blockade therapy was associated with higher concentrations of these soluble immune factors among patients with immune-reactive (hot) tumors. Such an association was not apparent for patients treated with cytotoxic chemotherapy or targeted therapy. Integrative analysis of tumor size, PD-L1 expression in tumor tissue (tPD-L1), and gene expression in tumor tissue and peripheral CD8+ T cells revealed that high concentrations of the 3 soluble immune factors were associated with hyper or terminal exhaustion of antitumor immunity. The combination of soluble PD-L1 (sPD-L1) and sCTLA-4 efficiently discriminated responsiveness to PD-1/PD-L1 blockade among patients with immune-reactive tumors.CONCLUSIONCombinations of soluble immune factors might be able to identify patients unlikely to respond to PD-1/PD-L1 blockade as a result of terminal exhaustion of antitumor immunity. Our data suggest that such a combination better predicts, along with tPD-L1, for the response of patients with NSCLC.TRIAL REGISTRATIONUMIN000019674.FUNDINGThis study was funded by Ono Pharmaceutical Co. Ltd. and Sysmex Corporation.

Post-allogeneic hematopoietic cell transplant (HCT) immunosuppression regimens are given as graft-versus-host disease (GVHD) prophylaxis. Most GVHD prophylaxis regimens are based on calcineurin inhibitors (CNIs). Unfortunately, CNIs are associated with significant associated morbidity, frequently cannot be tolerated, and often need to be discontinued. There is no consensus as to which alternative immunosuppression should be used in cases where CNIs have to be permanently discontinued. Cytotoxic T-lymphocyte-associated protein 4-immunoglobulin (CTLA4-Ig) blocking agents are well tolerated and have been used extensively in patients with autoimmune disease and as post-transplant immunosuppression. There are two CTLA4-Ig agents: belatacept and abatacept. Belatacept is routinely used in adult kidney transplantation to prevent rejection and abatacept has been approved by the Food and Drug Administration (FDA) for GVHD prophylaxis in patients undergoing a matched or one allele-mismatched unrelated allogenic HCT. Herein, we describe a case in which abatacept was given off-label to replace tacrolimus GVHD prophylaxis in a patient with neurotoxicity undergoing haploidentical HCT. This case suggests that CTLA4-Ig blockade may be a good alternative to a CNI in cases where the CNI needs to be discontinued and warrants further investigation.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has created one of the largest global health crises in almost a century. Although the current rate of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections has decreased significantly, the long-term outlook of COVID-19 remains a serious cause of morbidity and mortality worldwide, with the mortality rate still substantially surpassing even that recorded for influenza viruses. The continued emergence of SARS-CoV-2 variants of concern (VOCs), including multiple heavily mutated Omicron sub-variants, has prolonged the COVID-19 pandemic and underscores the urgent need for a next-generation vaccine that will protect from multiple SARS-CoV-2 VOCs. METHODS: We designed a multi-epitope-based coronavirus vaccine that incorporated B, CD4+, and CD8+ T- cell epitopes conserved among all known SARS-CoV-2 VOCs and selectively recognized by CD8+ and CD4+ T-cells from asymptomatic COVID-19 patients irrespective of VOC infection. The safety, immunogenicity, and cross-protective immunity of this pan-variant SARS-CoV-2 vaccine were studied against six VOCs using an innovative triple transgenic h-ACE-2-HLA-A2/DR mouse model. RESULTS: The pan-variant SARS-CoV-2 vaccine (i) is safe , (ii) induces high frequencies of lung-resident functional CD8+ and CD4+ TEM and TRM cells , and (iii) provides robust protection against morbidity and virus replication. COVID-19-related lung pathology and death were caused by six SARS-CoV-2 VOCs: Alpha (B.1.1.7), Beta (B.1.351), Gamma or P1 (B.1.1.28.1), Delta (lineage B.1.617.2), and Omicron (B.1.1.529). CONCLUSION: A multi-epitope pan-variant SARS-CoV-2 vaccine bearing conserved human B- and T- cell epitopes from structural and non-structural SARS-CoV-2 antigens induced cross-protective immunity that facilitated virus clearance, and reduced morbidity, COVID-19-related lung pathology, and death caused by multiple SARS-CoV-2 VOCs.

Objective and design: The exact immunological mechanism of widespread chronic inflammatory skin disorder psoriasis has not been fully established. CD11b+Gr.1+ myeloid-derived cells are immature heterogeneous cells with T-cell suppressive property in neoplasia; however, influence of these cells on adaptive immunity is highly contextual; therefore, we dubbed these cells as myeloid-derived adjuster cells (MDAC). We studied imiquimod induced psoriasis in mouse model and evaluated for the first time the RORγt-NFAT1 axis in MDACs and the function, differentiation and interaction of these cells with T cells. Materials and methods: The status of T cells and MDACs; their functionality and differentiation properties, and the roles of RORγt and NFAT1 in MDACs were evaluated using flow cytometry, qRT-PCR and confocal imaging. Results: We found gradual increase in T cells and MDACs and an increase in the number of IL17 -secreting MDACs and T cells in the skin of psoriatic animals. We also noted that MDAC differentiation is biased toward M1 macrophages and DCs which perpetuate inflammation. We found that psoriatic MDACs were unable to suppress T-cell proliferation or activation but seemingly helped these T cells produce more IL17. Inhibition of the RORγt/NFAT1 axis in MDACs increased the suppressive nature of MDACs, allowing these cells to suppress the activity of psoriatic T-cells. Conclusion: Our results indicate that altered MDAC properties in psoriatic condition sustains pathological inflammation and RORγt and NFAT1 as promising intervention target for psoriasis management. [ABSTRACT FROM AUTHOR]

Objective: The objective is to characterize transcriptomic profiles and immune cell composition and distribution in juvenile idiopathic arthritis (JIA) synovial biopsies, assess for associations of these features with clinical parameters, and compare JIA and rheumatoid arthritis (RA) synovial features. Methods: RNA sequencing (RNASeq) was performed on 24 samples, with pathway analysis and inference of relative abundance of immune cell subsets based on gene expression data. Two multiplex fluorescence immunohistochemistry (IHC) panels were performed on 28 samples (including 13 on which RNASeq was performed), staining for CD206− classical and CD206+ nonclassical macrophages, and CD8+ and CD4+ T and B lymphocytes. Data were compared to a published series of early RA synovial biopsies. Results: Pathway analysis of the most variably expressed genes (n = 339) identified a B and plasma cell signature as the main driver of heterogeneity in JIA synovia, with strong overlap between JIA and RA synovitis. Multiplex IHC confirmed heterogeneity of immune cell infiltration. M1‐like macrophage–rich synovial lining was associated with greater lining hypertrophy and higher (CD45+) pan–immune cell and CD8+ T cell infiltration. Conclusion: Our study indicates significant similarities between JIA and RA synovitis. Similar to RA, JIA synovia may be broadly categorized into two groups: (1) those with an inflammatory/adaptive immune transcriptomic signature, M1‐like macrophage and CD8+ T cell infiltration, and thicker, M1‐like macrophage–rich synovial lining, and (2) those with an M2‐like macrophage transcriptomic signature, greater M2/M1‐like macrophage ratios, and thinner, M2‐like macrophage–rich synovial lining. Synovial features were not significantly associated with clinical parameters, likely because of group size and heterogeneity. [ABSTRACT FROM AUTHOR]

Objective: Glucocorticoid‐induced tumor necrosis factor receptor superfamily–related protein (GITR), with its ligand (GITRL), plays an important role in CD4+ T cell–mediated autoimmunity. This study aimed to investigate the underlying mechanisms of GITRL in primary Sjögren syndrome (pSS). Methods: Patients with pSS and healthy controls were recruited. Serum GITRL and Th17‐related cytokines were determined. RNA sequencing was performed to decipher key signal pathways. Nonobese diabetes (NOD) mice were adopted as experimental Sjögren models and recombinant adeno‐associated virus (rAAV) transduction was conducted to verify the therapeutic potentials of targeting GITRL in vivo. Results: Serum GITRL was significantly higher in patients with pSS and showed a positive correlation with leukopenia, thrombocytopenia, autoantibodies, lung involvement, and disease activity. Serum GITRL was correlated with Th17‐related cytokines. GITRL promoted the expansion of Th17 and Th17.1 cells. Expansion of granulocyte‐macrophage colony‐stimulating factor positive (GM‐CSF+) CD4+ T cells induced by GITRL could be inhibited by blockade of GITRL. Moreover, GM‐CSF could stimulate GITRL expression on monocytes. RNA sequencing revealed mammalian target of rapamycin complexes 1 (mTORC1) might be the key modulator. The increased phosphorylation of S6 and STAT3 and the expansion of Th17 and Th17.1 cells induced by GITRL were effectively inhibited by rapamycin, suggesting a GITRL–mTORC1–GM‐CSF positive loop in pathogenic Th17 response in pSS. Administration of an rAAV vector expressing short hairpin RNA targeting GITRL alleviated disease progression in NOD mice. Conclusion: Our results identified the pathogenic role of GITRL in exacerbating disease activity and promoting pathogenic Th17 response in pSS through a GITRL–mTORC1–GM‐CSF loop. These findings suggest GITRL might be a promising therapeutic target in the treatment of pSS. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease that mainly affects the joints. Studies have shown that memory CD4+ T cells play an important role in the pathogenesis of RA. This study investigated the expression and function of CD305 on human memory CD4+ T cells and the effects of CD305 activating antibody on collagen-induced arthritis. The results showed that CD305 expression was significantly decreased on circulating memory CD4+ T cells from patients with RA and its mean fluorescence intensity (MFI) was negatively correlated with DAS28. Moreover, CD305 inhibited the activation of memory CD4+ T cells by down-regulating CD69 and CD25 and the production of IFN-γ, IL-4, and IL-17A induced by anti-CD3/CD28 antibodies. In addition, activation of CD305 inhibited the severity of disease in collagen-induced arthritis. In summary, CD305 reduction may mediate the excessive activation of memory CD4+ T cells and participate in the development of RA. It can be used as a predictive marker of disease activity and has potential medicinal value in the treatment of RA. • CD305 decreases prominently on circulating memory CD4+ T cells and is negatively correlated with DAS28 of patients with RA. • CD305 participates in the abnormal activation of memory CD4+ T cells in patients with RA. • CD305 inhibits secretion of IFN-γ, IL-4 and IL-17A by memory CD4+ T cells. • CD305 attenuates arthritis severity in Collagen-induced arthritis. [ABSTRACT FROM AUTHOR]

HIV-1 chronically infects host CD4+ T lymphocytes and further affects a variety of immune cells, including CD8+ T cells. In our previous study, by analyzing unbiased high-dimensional single-cell RNA-seq data (scRNA-seq), we found that the frequency of GZMK+CD8+ T cells expressing granzyme K (GZMK) was increased in people living with HIV-1 (PLWHs). However, the phenotypic and functional characteristics of these cells in chronic HIV-1 infection and their correlation with disease are not well understood. In this study, we conducted a comprehensive analysis of scRNA-seq and matched T-cell receptor repertoire (TCR) sequencing data to delve into the characterizations of GZMK+CD8+ T cells, which was further validated by flow cytometry. We observed heterogeneity within the GZMK+CD8+ T cells, which could be further subdivided into a GZMK+GZMB- subset and a GZMK+GZMB+ subset, with the latter being significantly enriched in PLWHs. The GZMK+GZMB+ cells are a unique subset within CD8+ T cells, characterized by high proliferation, activation, inflammatory response, clone transition, etc., and are one of the differentiation endpoints by pseudotemporal analysis of CD8+αβ T cells. Despite being predominantly composed of effector memory T cells (Tem), similar to the GZMK+GZMB- subset, the GZMK+GZMB+ subset exhibits differentiation at a later stage than the GZMK+GZMB- subset. We also observed that the frequency/count of GZMK+GZMB+CD8+ T cells was negatively correlated with CD4/CD8 ratio, and positively correlated with HIV DNA, IP-10, and MIG levels in PLWHs. In vitro experiments demonstrate that GZMK can potentiate the stimulatory effects of lipopolysaccharide (LPS) on THP-1 macrophages via the TLR-4 pathway, significantly enhancing the secretion of IP-10, MIG, and MCP-1, as well as increasing the proportion of TNF-α+ cells. In conclusion, in PLWHs, GZMK+GZMB+CD8+ T cells are a highly reactive and inflammatory-inducing subset that may be associated with systemic inflammation. • We characterized GZMK+CD8+ T cells and identified a new subset in HIV-1 patients. • We used scRNA-seq, TCR-seq, flow cytometry, and functional experiments. • The GZMK+GZMB+ subset is a subset of GZMK+ T cells that is enriched in HIV-1 patients. • The GZMK+GZMB+ subset is characterized by proliferation, activation, inflammation. • The GZMK+GZMB+ subset is highly reactive and associated with systemic inflammation. [ABSTRACT FROM AUTHOR]

Aging is a gradual, inevitable physiologic process. The organ aging is related to the persistence of chronic inflammation, but the understanding of inflammatory state during renal aging is lacking currently. Single-cell transcriptome sequencing was performed on aging mouse kidney to reveal the molecular phenotype and composition changes of different cell types. In the early stage of aging, immune cells such as T, B cells and mononuclear macrophages increased in kidney. The molecular state of T cells in aging kidney changed and polarized. Among them, we identified a group of GZMK + CD8 + T cells with high expression of Eomes , Pdcd1 and Ifng and a group of Il17a + T cells with high expression of Il17a and Il23r. Moreover, the cytokines and inflammations can aggravate tissue damage eventually. Furthermore, we found the interaction between different types of epithelial cells and T cells increased during the renal aging. These results identify the changes of T cells in the early stage of aging kidney and suggest that GZMK+CD8+ T cells might be a potential target to ameliorate age-associated dysfunctions of kidney(Graphical Abstract). [Display omitted] • Kidney undergoes the most severe structural and functional deterioration with aging. • Organ aging is related to chronic inflammation, and persist inflammation hinders intrinsic cell repair. • Immune cells increased in the early stage of kidney aging. • The molecular state of T cells in aging kidney changed and polarized. • The crosstalk between different types of epithelial cells and immune cells increased. [ABSTRACT FROM AUTHOR]

Chronic hepatitis B virus (HBV) infection is a significant global public health concern, and the clearance of HBV is closely linked to the activity of HBV‐specific T cells, which is regulated by various co‐suppressor molecules. Cytotoxic T lymphocyte‐associated antigen 4 (CTLA‐4) is among these co‐suppressor molecules which induces T cell exhaustion by competitively inhibiting CD28 and dampening the function of HBV‐specific T cells. CTLA‐4 also plays a role in the regulation of T helper (Th) cell differentiation and influences cytokine release. In addition, CTLA‐4 can impact glucose metabolism in hepatocellular carcinoma through its interaction with T regulatory (Treg) cells. This review aims to provide a comprehensive overview of the existing literature related to the role of CTLA‐4 in HBV patients across different subsets of T cells. Additionally, we propose a discussion on the possible mechanisms through which CTLA‐4 may contribute to HBV infection, as well as the development of HBV‐induced cirrhosis and hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

Biomedical research has witnessed significant strides in manufacturing chimeric antigen receptor T cell (CAR-T) therapies, marking a transformative era in cellular immunotherapy. Nevertheless, existing manufacturing methods for autologous cell therapies still pose several challenges related to cost, immune cell source, safety risks, and scalability. These challenges have motivated recent efforts to optimize process development and manufacturing for cell therapies using automated closed-system bioreactors and models created using artificial intelligence. Simultaneously, non-viral gene transfer methods like mRNA, CRISPR genome editing, and transposons are being applied to engineer T cells and other immune cells like macrophages and natural killer cells. Alternative sources of primary immune cells and stem cells are being developed to generate universal, allogeneic therapies, signaling a shift away from the current autologous paradigm. These multifaceted innovations in manufacturing underscore a collective effort to propel this therapeutic approach toward broader clinical adoption and improved patient outcomes in the evolving landscape of cancer treatment. Here, we review current CAR immune cell manufacturing strategies and highlight recent advancements in cell therapy scale-up, automation, process development, and engineering. [ABSTRACT FROM AUTHOR]

Background Psoriasis is a T cell-mediated chronic inflammatory skin condition characterized by the interaction of T cells with various cell types, forming an inflammatory microenvironment that sustains psoriatic inflammation. Homeostasis of these tissue-resident T cells is supported by fibroblasts, the primary structural cells in the dermis. In psoriasis, there is increased expression of matrix metalloproteinase 2 (MMP2), mediating structural alterations in skin tissues and modulating inflammation. Additionally, the CD100–plexin-B2 (PLXNB2) axis is known to enhance psoriasis inflammation via keratinocytes, and CD103 levels are associated with the severity of psoriasis upon relapse. Objectives To elucidate the role of fibroblasts and the MMP2–CD100 axis in modulating psoriasis inflammation. Methods CD100 expression and function in psoriasis were assessed using immunofluorescence, enzyme-linked immunosorbent assay, single-cell transcriptome sequencing, cellular interaction analyses and quantitative reverse transcriptase polymerase chain reaction. CD8+ T cells from people with psoriasis were isolated using magnetic beads, to investigate the regulatory effect of MMP2 on CD100 expression on their membranes. Single-cell transcriptome sequencing, spatial transcriptome sequencing, mimetic timing analysis, immunofluorescence and flow cytometry were used to determine the origin of MMP2 and its impact on CD103+ CD8+ T cells. The hypotheses were further validated in vivo using MMP2 and CD100 inhibitors. Results Soluble CD100 (sCD100) was significantly upregulated in both psoriatic lesions and peripheral blood, amplifying psoriasis inflammation by promoting the production of inflammatory cytokines by keratinocytes, fibroblasts and endothelial cells via the sCD100–PLXNB2 axis. Fibroblasts that highly expressed MMP2 (MMP2hi) exacerbated psoriasis symptoms by facilitating CD100 shedding from CD8+ T-cell membranes. Additionally, it was shown that fibroblasts enhance the upregulation of the CD8+ T-cell residency factor CD103 in co-cultures with CD8+ T cells. Inhibitors targeting MMP2 and CD100 were effective in reducing inflammation in an imiquimod-induced psoriasis model. Conclusions Our findings underscore the pivotal role of MMP2hi fibroblasts in the amplification and recurrence of inflammatory responses in psoriasis. These fibroblasts augment psoriasis inflammation through the CD100–PLXNB2 axis by facilitating CD100 shedding on CD8+ T-cell membranes and by upregulating CD103, thereby enhancing CD8+ T-cell residency. [ABSTRACT FROM AUTHOR]

Aim: Neural activity in the olfactory bulb (OB) can represent odor information during different brain and behavioral states. For example, the odor responses of mitral/tufted (M/T) cells in the OB change during learning of odor‐discrimination tasks and, at the network level, beta power increases and the high gamma (HG) power decreases during odor presentation in such tasks. However, the neural mechanisms underlying these observations remain poorly understood. Here, we investigate whether serotonergic modulation from the dorsal raphe nucleus (DRN) to the OB is involved in shaping activity during the learning process in a go/no‐go task in mice. Methods: Fiber photometry was used to record the population activity of DRN serotonergic neurons during a go/no‐go task. In vivo electrophysiology was used to record neural activity (single units and local field potentials) in the OB during the go/no‐go task. Real‐time place preference (RTPP) and intracranial light administration in a specific subarea (iClass) tests were used to assess the ability of mice to encoding reward information. Results: Odor‐evoked population activity in serotonergic neurons in the DRN was shaped during the learning process in a go/no‐go task. In the OB, neural activity from oscillations to single cells showed complex, learning‐associated changes and ability to encode information during an odor discrimination task. However, these properties were not observed after ablation of DRN serotonergic neurons. Conclusion: The activity of neural networks and single cells in the OB, and their ability to encode information about odor value, are shaped by serotonergic projections from the DRN. [ABSTRACT FROM AUTHOR]

B cells play a central role in the pathogenesis of systemic sclerosis (SSc). Most B-cell studies have focused on their pathological role as antibody producers. However, in addition to immunoglobulin secretion, these cells have a wide range of functions in the immune response, including antigen presentation to T cells and cytokine production. Importantly, not all B-cell subsets promote the immune response. Regulatory B cells (Bregs) attenuate inflammation and contribute to the maintenance of immune tolerance. However, effector B cells (Beffs) positively modulate the immune response through the production of various cytokines. In SSc, Bregs are insufficient and/or dysfunctional. B-cell-targeting biologics have been trialled with promising results in the treatment of SSc. These therapies can affect Bregs or Beffs, which can potentially limit their long-term efficacy. Future strategies might involve the modulation of effector B cells in combination with the stimulation of regulatory subsets. Additionally, the monitoring of individual B-cell subsets in patients may lead to the discovery of novel biomarkers that could help predict disease relapse or progression. The purpose of this review is to summarize the relevant literatures and explain how Bregs and Beffs jointly participate in the pathogenesis of SSc. [ABSTRACT FROM AUTHOR]

Overexpression of T‐cell immunoglobulin and mucin‐domain containing‐3 (TIM‐3) on T cells has been observed in smokers. However, whether and how galectin‐9 (Gal‐9)/TIM‐3 signal between T‐regulatory cells (Tregs) and type 17 helper (Th17) cells contributes to tobacco smoke‐induced airway inflammation remains unclear. Here, we aimed to explore the role of the Gal‐9/TIM‐3 signal between Tregs and Th17 cells during chronic tobacco smoke exposure. Tregs phenotype and the expression of TIM‐3 on CD4+ T cells were detected in a mouse model of experimental emphysema. The role of TIM‐3 in CD4+ T cells was explored in a HAVCR2−/− mouse model and in mice that received recombinant anti‐TIM3. The crosstalk between Gal‐9 and Tim‐3 was evaluated by coculture Tregs with effector CD4+ T cells. We also invested the expression of Gal‐9 in Tregs in patients with COPD. Our study revealed that chronic tobacco smoke exposure significantly reduces the frequency of Tregs in the lungs of mice and remarkably shapes the heterogeneity of Tregs by downregulating the expression of Gal‐9. We observed a pro‐inflammatory but restrained phenotypic transition of CD4+ T cells after tobacco smoke exposure, which was maintained by TIM‐3. The restrained phenotype of CD4+ T cells was perturbed when TIM‐3 was deleted or neutralised. Tregs from the lungs of mice with emphysema displayed a blunt ability to inhibit the differentiation and proliferation of Th17 cells. The inhibitory function of Tregs was partially restored by using recombinant Gal‐9. The interaction between Gal‐9 and TIM‐3 inhibits the differentiation of Th17 cells and promotes apoptosis of CD4+ T cells, possibly by interfering with the expression of retinoic acid receptor‐related orphan receptor gamma t. The expression of Gal‐9 in Tregs was reduced in patients with COPD, which was associated with Th17 response and lung function. These findings present a new paradigm that impairment of Gal‐9/Tim‐3 crosstalk between Tregs and Th17 cells during chronic tobacco smoke exposure promotes tobacco smoke‐induced airway/lung inflammation. [ABSTRACT FROM AUTHOR]

Lung adenocarcinoma (LUAD) is the most prevalent subtype of lung cancer, and the early detection and diagnosis of this disease are crucial in reducing mortality rates. The timely diagnosis of LUAD is essential for controlling tumour development and enabling early surgical treatment. GPR56 is a vital G protein‐coupled receptor and its role in T lymphocytes has received considerable attention. However, its function in B cells remains unclear. This study aimed to investigate the significance of GPR56 in LUAD. We found that GPR56 exhibited a significant increase in circulating plasmablasts and a decrease in new memory B cells. GPR56 expression in B cells was significantly reduced after LPS stimulation and the proportion of HLA‐DR+ and CD40+ proportions were also decreased in GPR56+ B cells after stimulation. Additionally, GPR56 exhibited significant down‐regulation in circulating B cell subsets of early‐stage LUAD patients, and there were significant correlations between GPR56+ B cell subsets and tumour markers. In conclusion, GPR56 could reflect the hypoactivation state of B cells and the decreased proportion of GPR56+ B cell subset in LUAD patients can signify the active humoral immunity in vivo. The expression of GPR56 in B cells could potentially hold value in the early diagnosis of LUAD. [ABSTRACT FROM AUTHOR]

Advances in single‐cell level analytical techniques, especially cytometric approaches, have led to profound innovation in biomedical research, particularly in the field of clinical immunology. This has resulted in an expansion of high‐dimensional data, posing great challenges for comprehensive and unbiased analysis. Conventional manual analysis is thus becoming untenable to handle these challenges. Furthermore, most newly developed computational methods lack flexibility and interoperability, hampering their accessibility and usability. Here, we adapted Seurat, an R package originally developed for single‐cell RNA sequencing (scRNA‐seq) analysis, for high‐dimensional flow cytometric data analysis. Based on a 20‐marker antibody panel and analyses of T‐cell profiles in both adult blood and cord blood (CB), we showcased the robust capacity of Seurat in flow cytometric data analysis, which was further validated by Spectre, another high‐dimensional cytometric data analysis package, and conventional manual analysis. Importantly, we identified a unique CD8+ T‐cell population defined as CD8+CD45RA+CD27+CD161+ T cell that was predominantly present in CB. We characterised its IFN‐γ‐producing and potential cytotoxic properties using flow cytometry experiments and scRNA‐seq analysis from a published dataset. Collectively, we identified a unique human CB CD8+CD45RA+CD27+CD161+ T‐cell subset and demonstrated that Seurat, a widely used package for scRNA‐seq analysis, possesses great potential to be repurposed for cytometric data analysis. This facilitates an unbiased and thorough interpretation of complicated high‐dimensional data using a single analytical pipeline and opens a novel avenue for data‐driven investigation in clinical immunology. [ABSTRACT FROM AUTHOR]

Members of the Protein kinases D (PKD) family are described as regulators of T cell responses. From the two T cell‐expressed isoforms PKD2 and PKD3, so far mainly the former was thoroughly investigated and is well understood. Recently, we have investigated also PKD3 using conventional as well as conditional T cell‐specific knockout models. These studies suggested PKD3 to be a T cell‐extrinsic regulator of the cells' fate. However, these former model systems did not take into account possible redundancies with the highly homologous PKD2. To overcome this issue and thus properly unravel PKD3's T cell‐intrinsic functions, here we additionally used a mouse model overexpressing a constitutively active isoform of PKD3 specifically in the T cell compartment. These transgenic mice showed a slightly higher proportion of central memory T cells in secondary lymphoid organs and blood. This effect could not be explained via differences upon polyclonal stimulation in vitro, however, may be connected to the observed developmental aberrances in the CD8 single positive compartment during thymic development. Lastly, the observed alterations in the CD8+ T cell compartment did not impact proper immune response upon immunization with ovalbumin or in a subcutaneous tumour model suggesting only a small to absent biological relevance. Taking together the knowledge of all our published studies on PKD3 in the T cell compartment, we now conclude that T cell‐intrinsic PKD3 is a fine‐tuner of central memory T cell as well as CD8 single positive thymocyte development. [ABSTRACT FROM AUTHOR]

Cytokines of the common‐γ receptor chain (γc) family are crucial for T‐cell differentiation and dysregulation of γc cytokine pathways is involved in the pathogenesis of autoimmune diseases. There is increasing evidence that the availability of the γc receptor (CD132) for the associated receptor chains has implications for T‐cell functions. Here we studied the influence of differential γc expression on the expression of the IL‐2Rα (CD25), the IL‐7Rα (CD127) and the differentiation of activated naïve T cells. We fine‐tuned the regulation of γc expression in human primary naïve T cells by lentiviral transduction using small hairpin (sh)RNAs and γc cDNA. Differential γc levels were then analysed for effects on T‐cell phenotype and function after activation. Differential γc expression markedly affected IL‐2Rα and IL‐7Rα expression on activated naïve T cells. High γc expression (γc‐high) induced significantly higher expression of IL‐2Rα and re‐expression of IL‐7Rα after activation. Inhibition of γc caused lower IL‐2Rα/IL‐7Rα expression and impaired proliferation of activated naïve T cells. In contrast, γc‐high T cells secreted significantly higher concentrations of effector cytokines (i.e., IFN‐γ, IL‐6) and showed higher cytokine‐receptor induced STAT5 phosphorylation during initial stages as well as persistently higher pSTAT1 and pSTAT3 levels after activation. Finally, accelerated transition towards a CD45RO expressing effector/memory phenotype was seen especially for CD4+ γc‐high naïve T cells. These results suggested that high expression of γc promotes expression of IL‐2Rα and IL‐7Rα on activated naïve T cells with significant effects on differentiation and effector cytokine expression. [ABSTRACT FROM AUTHOR]

Immune checkpoint inhibitor therapy can provide significant clinical benefit in patients with certain cancer types including melanoma; however, objective responses are only observed for a subset of patients. Mucosal melanoma is a rare melanoma subtype associated with a poor prognosis and, compared with cutaneous melanoma, is significantly less responsive to immune checkpoint inhibitors. Spontaneous canine tumours have emerged as valuable models to inform human cancer studies. In contrast to human melanoma, most canine melanomas are mucosal—an incidence that may be leveraged to better understand the subtype in humans. However, a more comprehensive understanding of the immune landscape of the canine disease is required. Here, we quantify tumour infiltrative T and myeloid cells in canine mucosal (n = 13) and cutaneous (n = 5) melanomas using immunohistochemical analysis of CD3 and MAC387 expression, respectively. Gene expression analysis using the Canine IO NanoString panel was also performed to identify genes and pathways associated with immune cell infiltration. T and myeloid cell densities were variable with geometric means of 158.7 cells/mm2 and 166.7 cells/mm2, respectively. Elevated T cell infiltration was associated with increased expression of cytolytic genes as well as genes encoding the coinhibitory checkpoint molecules PD‐1, CTLA‐4, TIM‐3 and TIGIT; whereas increased myeloid cell infiltration was associated with elevated expression of protumourigenic cytokines. These data provide a basic characterization of the tumour microenvironment of canine malignant melanoma and suggest that, like human melanoma, inherent variability in anti‐tumour T cell responses exists and that a subset of canine melanomas may respond better to immunomodulation. [ABSTRACT FROM AUTHOR]

It is urgent to explore factors affecting immunotherapy efficacy to benefit non‐small cell lung cancer (NSCLC) patient survival. Bioinformatics predicted genes associated with programmed cell death ligand 1 (PD‐L1) expression and analysed phospholipase A2 group IID (PLA2G2D) expression in NSCLC. BODIPY 493/503 dye staining and kits detected lipids, triglycerides, and phospholipids in H1299 cells, respectively. Extracellular vesicles (EVs) were extracted for morphology and size assessment using electron microscopy. Western blot assayed CD9, CD63, HSP90, EVs‐PD‐L1, PD‐L1, and PLA2G2D expression. CCK‐8, LDH, and ELISA tested proliferation and toxicity of CD8+ T cells, interleukin‐2, and interferon‐gamma secretion, respectively. PLA2G2D, PD‐L1, and Ki67 expression was detected by immunohistochemistry. Immunofluorescence assayed PLA2G2D localisation and CD8+ T cell content. Flow cytometry assessed PD‐L1 and CD8 expression. In NSCLC, upregulated EVs‐PD‐L1 and clinical characteristics showed a strong correlation. H1299 cells with overexpression PD‐L1 significantly reduced proliferation, toxicity of CD8+ T cells, and interleukin‐2 and interferon‐gamma levels. Bioinformatics revealed positive correlations between PLA2G2D and overexpressed PD‐L1. PLA2G2D was expressed in macrophages and dendritic cells in NSCLC tissue. Overexpression PLA2G2D (oe‐PLA2G2D) increased lipids, triglycerides, and phospholipids contents in H1299 cells. oe‐PLA2G2D significantly reduced proliferation, toxicity of CD8+ T cells, and interleukin‐2 and interferon‐gamma levels. si‐PD‐L1 restored inhibition of oe‐PLA2G2D on CD8+ T cells. oe‐PLA2G2D significantly increased mice tumour volume and weight, upregulated expression of blood EVs‐PD‐L1 and tissue PD‐L1, PLA2G2D, Ki67, and decreased CD8+ T cell content. PLA2G2D facilitated immune escape in NSCLC by regulating CD8+ T cell immune function by upregulating EVs‐PD‐L1. [ABSTRACT FROM AUTHOR]

Mucosal‐associated invariant T (MAIT) cells are innate‐like T cells that can be activated by microbial antigens and cytokines and are abundant in mucosal tissues including the colon. MAIT cells have cytotoxic and pro‐inflammatory functions and have potentials for use as adoptive cell therapy. However, studies into their anti‐cancer activity, including their role in colon cancer, are limited. Using an animal model of colon cancer, we showed that peritumoral injection of in vivo‐expanded MAIT cells into RAG1−/− mice with MC38‐derived tumours inhibits tumour growth compared to control. Multiplex cytokine analyses showed that tumours from the MAIT cell‐treated group have higher expression of markers for eosinophil‐activating cytokines, suggesting a potential association between eosinophil recruitment and tumour inhibition. In a human peripheral leukocyte co‐culture model, we showed that leukocytes stimulated with MAIT ligand showed an increase in eotaxin‐1 production and activation of eosinophils, associated with increased cancer cell killing. In conclusion, we showed that MAIT cells have a protective role in a murine colon cancer model, associated with modulation of the immune response to cancer, potentially involving eosinophil‐associated mechanisms. Our results highlight the potential of MAIT cells for non‐donor restricted colon cancer immunotherapy. [ABSTRACT FROM AUTHOR]

Mesoporous silica nanoparticles (MSNs) are physically and chemically stable inorganic nanomaterials that have been attracting much attention as carriers for drug delivery systems in the field of nanomedicine. In the present study, we investigated the potential of MSN vaccines that incorporate antigen peptides for use in cancer immunotherapy. In vitro experiments demonstrated that fluorescently labeled MSNs accumulated in a line of mouse dendritic cells (DC2.4 cells), where the particles localized to the cytosol. These observations could suggest that MSNs have potential for use in delivering the loaded molecules into antigen-presenting cells, thereby stimulating the host acquired immune system. In vivo experiments demonstrated prolonged survival in mice implanted with ovalbumin (OVA)-expressing lymphoma cells (E.G7-OVA cells) following subcutaneous inoculation with MSNs incorporating OVA antigen peptides. Furthermore, OVA-specific immunoglobulin G antibodies and cytotoxic T lymphocytes were detected in the serum and the spleen cells, respectively, of mice inoculated with an MSN-OVA vaccine, indicating the induction of antigen-specific responses in both the humoral and cellular immune systems. These results suggested that the MSN therapies incorporating antigen peptides may serve as novel vaccines for cancer immunotherapy. [ABSTRACT FROM AUTHOR]

The specific biology of the male breast remains relatively unexplored in spite of the increasing global prevalence of male breast cancer. Delineation of the microenvironment of the male breast is restricted by the low availability of human samples and a lack of characterisation of appropriate animal models. Unlike the mouse, the male ovine gland persists postnatally. We suggest that the male ovine mammary gland constitutes a promising adjunctive model for the male breast. In this study, we evaluate the male ovine mammary gland microenvironment, comparing intact and neutered males. Assessment of the glandular histo‐anatomy highlights the resemblance of the male gland to that of neonatal female sheep and confirms the presence of rudimentary terminal duct lobular units. Irrespective of neutered status, cell proliferation in epithelial and stromal compartments is similarly low in males, and cell proliferation in epithelial cells and in the intralobular stroma is significantly lower than in pubertal female sheep. Between 42% and 72% of the luminal mammary epithelial cells in the male gland express the androgen receptor and expression is significantly reduced by neutering. Luminal epithelial cells within the intact and neutered male gland also express oestrogen receptor alpha, but minimal progesterone receptor expression is observed. The distribution of leukocytes within the ducts and stroma is similar to the mammary gland of female sheep and females of other species. Both macrophages and T lymphocytes are intercalated in the epithelial bilayer and are more abundant in the intralobular stroma than the interlobular stroma, suggesting that they may have a protective immunological function within the vestigial glandular tissue of the male sheep. Mast cells are also observed within the stroma. These cells cluster near the glandular tissue and are frequently located adjacent to blood vessels. The abundance of mast cells is significantly higher in intact males compared to neutered males, suggesting that hormone signalling may impact mast cell recruitment. In this study, we demonstrate the utility of the male ovine mammary gland as a model for furthering our knowledge of postnatal male mammary biology. [ABSTRACT FROM AUTHOR]

Diffuse large B‐cell lymphoma (DLBCL) is the most frequent lymphoma. MIC‐A and MIC‐B are the natural ligands for NKG2D, a receptor expressed in NK cells. MIC‐A soluble isoforms (sMICA) have been described in different malignancies. Objectives: To analyze lymphocyte subsets and sMIC‐A in germinal center DLBCL. Materials and Methods: sMICA, sMICB, and peripheral blood lymphocyte subsets (CD4+, CD8+, NK, NKT, γδ T cells, and dendritic cells) were analyzed in 59 patients and 60 healthy donors. Results: Patients had decreased numbers of type 1 and type 2 dendritic cells, NK, iNKT, CD4 T, and CD8 T cells, and higher levels of sMIC‐A. The 2‐year PFS for high IPI scores and high sMIC‐A was 24% and 28%, respectively. The 2‐year OS for high IPI scores and high sMIC‐A was 42% and 33%. The 2‐year PFS and OS for patients not achieving response to treatment were 0% and 10%, respectively. The MICPI score (one point each for high IPI score and high sMIC‐A) showed that those patients summing two points had worse PSF and OS. Conclusions: Patients with DLBCL have decreased numbers of peripheral lymphocyte subsets and high levels of sMIC‐A. The addition of sMIC‐A to IPI could improve its prognostic relevance. [ABSTRACT FROM AUTHOR]

A comprehensive meta-analysis on the composition of circulating immune cells from both the myeloid and the lymphoid lines including specialized subsets in blood and cerebrospinal fluid (CSF) of patients with psychotic disorders compared with healthy control participants has been lacking. Multiple databases (PubMed, EMBASE, Cochrane Library, Web of Science, ClinicalTrials.gov, and PsycINFO) were searched for eligible studies up until October 18, 2022. All studies investigating circulating immune cells in the blood and CSF from patients with psychotic disorders (ICD-10: F20 and F22-29) compared with healthy control participants were included. A total of 86 studies were included in the meta-analysis. In the blood, the following categories of immune cells were elevated: leukocyte count (31 studies, standardized mean difference [SMD] = 0.35; 95% CI, 0.24 to 0.46), granulocyte count (4 studies, SMD = 0.57; 95% CI, 0.12 to 1.01), neutrophil granulocyte count (21 studies, SMD = 0.32; 95% CI, 0.11 to 0.54), monocyte count (23 studies, SMD = 0.40; 95% CI, 0.23 to 0.56), and B lymphocyte count (10 studies, SMD = 0.26; 95% CI, 0.04 to 0.48). Additionally, the neutrophil/lymphocyte ratio (23 studies, SMD = 0.40; 95% CI, 0.19 to 0.60), the monocyte/lymphocyte ratio (9 studies, SMD = 0.31; 95% CI, 0.04 to 0.57), and the platelet/lymphocyte ratio (10 studies, SMD = 0.23; 95% CI, 0.03 to 0.43) were elevated. The CSF cell count showed a similar tendency but was not significantly elevated (3 studies, SMD = 0.14; 95% CI, −0.04 to 0.32). The results indicate a broad activation of the immune system in psychotic disorders, with cells from both the myeloid and the lymphoid line being elevated. However, CSF analyses were lacking in most of the studies, and many studies were hampered by insufficient adjustment for confounding factors such as body mass index and smoking. [ABSTRACT FROM AUTHOR]

Background: This study aimed to assess how acupoint catgut-embedding therapy influencesTh2-type immune response and the infiltration of CD4+ and CD8+ cells in DNCB-induced atopic dermatitis in BALB/c mice. It also conducted an initial examination of the underlying molecular mechanisms. Methods: Seventy-two mice were randomly divided intofourgroups: normal control, DNCB-induced atopic dermatitis model (AD), AD with acupoint catgut-embedding treatment (ADA), and AD with sham-acupoint catgut-embedding treatment. After DNCB challenge to induce AD, the ADA group received acupoint catgut-embedding therapy treatment at Zusanli (ST 36) and Quchi (LI 11) acupoints everyother week from day 8. Mice in the AD with sham-acupoint catgut-embedding treatment group underwent the same procedure as the ADA group but without catgut implantation. Severity was assessed using SCORAD on treatment days 1, 10, and 20. On day 18, nine miceper group were euthanized, and the remaining on day 28. Histopathological changes were observed using hematoxylin-eosin and immunohistochemistry staining. TNF-α, IL-4, IL-6, and IL-13 levels were analyzed by ELISA, and GATA3 and STAT6 protein levels by western blot. Results: After 20 days of acupoint catgut-embedding therapy treatment, mice showed reduced dermatitis scores compared to DNCB-induced AD-like mice. Significant decreases occurred in serum IL-4, IL-6, IL-13, and TNF-α levels. Skin analysis revealed marked reductions in CD4+ and CD8+ cell infiltration, as well as GATA3 and STAT6 protein levels. Conclusion: Acupoint catgut-embedding therapy may effectively alleviate atopic dermatitis by suppressing Th2 immune responses via the STAT6-GATA3 pathway and reducing CD4+and CD8+ T cell infiltration in skin lesions. [ABSTRACT FROM AUTHOR]

Hepatitis B virus (HBV) remains an important healthcare challenge, leading to liver diseases like cirrhosis and cancer. In response, we created a prophylactic and therapeutic HBV vaccine by integrating HBcAg and HBsAg from HBV genotype B into Norovirus (NoV) GII.2 P domain (PdomGII.2-HBV) for enhanced intranasal delivery. This vaccine also aimed to simultaneously prevent NoV infection, which causes gastroenteritis. Since the selected HBV epitopes have undergone extensive research and are tailored to the Indonesian population, this study focused on identifying NoV epitopes and assessing T cell epitopes coverage of the PdomGII.2-HBV for the Indonesian population. Following that, we expressed the PdomGII.2-HBV protein using Escherichia coli BL21(DE3) and employed a gentle solubilization technique for protein purification. Our in-silico analysis identified two B cell epitopes, along with 15 CD4+T cell epitopes and 35 CD8+T cell epitopes within the GII.2 P domain. These T cell epitopes cover 100% of the Javanese-Sundanese population's HLA allele variations, which constituted the largest demographic group in Indonesia. Subsequently, we successfully purified the presumed PdomGII.2-HBV protein, revealing a molecular weight of 39.5 kDa. Following the successful expression and purification of the presumed PdomGII.2-HBV protein, it is evident that this vaccine design has significant potential, warranting further study. [ABSTRACT FROM AUTHOR]

Human papillomavirus (HPV) L1 major capsid protein generates a highly immunogenic virus like particles (VLPs), which have been used as the main component of its prophylactic vaccine. However, the neutralizing antibodies against L1 VLPs are mostly type specific and may not be effective to prevent infection from different strains of HPV. On the other hand, HPV L2 minor capsid protein has low antigenic variation, thus can induce cross-neutralization. This study aims to obtain HPV 52 L1/L2 chimeric protein, which is designed based on HPV type 52 as one of the most circulated high-risk types in Indonesia, to develop a broad-spectrum HPV vaccine. Substitution of HPV 52 H4 helix L1 region with an HPV 52 L2 epitope was carried out using overlap extension PCR. HPV 52 L1/L2 chimeric gene was constructed into pET-SUMO expression vector and expressed in Escherichia coli BL21 (DE3). Bioinformatics modeling suggested that L2 epitope was located inside of the loop region in monomer form, and on the contrary, it was located outside of the pentamer surface. Furthermore, B cell and T cell epitopes predictions were conducted using Immune Epitope Database (IEDB) analysis. The B cell epitopes prediction revealed eleven potential epitopes, whereas the T cell epitopes prediction showed seven potential epitopes for each MHC class I and MHC class II. This study showed that HPV 52 L1/L2 chimeric protein has the potential to induce cross-neutralizing antibodies and can be developed as a promising candidate for a new HPV vaccine. [ABSTRACT FROM AUTHOR]

Tumor immunotherapies targeting PD-(L)1 exhibit anti-tumor efficacy in only 10–30% of patients with various cancers. Literature has demonstrated that a "hot tumor" which contains high T lymphocytes in the tumor microenvironment exhibits a better response to immunotherapies than a "cold tumor." This study aimed to investigate whether tumor-intrinsic IFNα and CXCL10 determine the recruitment and activation of CD8+ T cells to become "hot tumor." In this study, we found that CXCL10 overexpressed in a variety of tumors including lung, colon, and liver tumors with a correlation with PD-L1. High PD-L1 and CXCL10 are associated with better survival rates in tumor patients receiving immunotherapies. IFNs-downstream transcriptional factor IRF-1 and STAT1 were correlated with PD-L1 and CXCL10 expression. We demonstrated that IRF-1 and STAT1 were both bound with the promoters of PD-L1 and CXCL10, sharing the same signaling pathway and determining IFNs-mediated PD-L1 and CXCL10 expression. In addition, IFNα significantly increased activation marker IFNγ in PBMCs, promoting M1 type monocyte differentiation, CD4+ T, and CD8+ T cell activation. Particularly, we found that CD8+ T lymphocytes abundantly expressed CXCR3, a receptor of CXCL10, by flow cytometry, indicating that tumor-intrinsic CXCL10 potentially recruited CD8+ T in tumor microenvironment. To demonstrate the hypothesis, immunotherapy-sensitive CT26 and immunotherapy-resistant LL/2 were used and we found that CT26 cells exhibited higher IFNα, IFNγ, CXCL10, and PD-L1 levels compared to LL/2, leading to higher IFNγ expression in mouse splenocytes. Moreover, we found that CD8+ T cells were recruited by CXCL10 in vitro, whereas SCH546738, an inhibitor of CXCR3, inhibited T cell migration and splenocytes-mediated anti-tumor effect. We then confirmed that CT26-derived tumor was sensitive to αPD-L1 immunotherapy and LL/2-tumor was resistant, whereas αPD-L1 significantly increased T lymphocyte activation marker CD107a in CT26-derived BALB/c mice. In conclusion, this study revealed that CXCL10 expression is correlated with PD-L1 in tumors, sharing the same signaling pathway and associating with better immunotherapeutic efficacy. Further evidence in the syngeneic tumor models demonstrated that immunotherapy-sensitive CT26 intrinsically exhibited higher IFNα and CXCL10 compared to immunotherapy-resistant LL/2 to recruit and activate CD8+ T cells in the tumor microenvironment, exhibiting "hot tumor" characteristic of sensitizing αPD-L1 immunotherapies. [ABSTRACT FROM AUTHOR]

Γδ T cell infiltration into tumours usually correlates with improved patient outcome, but both tumour-promoting and tumoricidal effects of γδ T cells have been documented. Human γδ T cells can be divided into functionally distinct subsets based on T cell receptor (TCR) Vδ usage. Still, the contribution of these different subsets to tumour immunity remains elusive. Here, we provide a detailed γδ T cell profiling in colon tumours, using mass and flow cytometry, mRNA quantification, and TCR sequencing. δ chain usage in both the macroscopically unaffected colon mucosa and tumours varied considerably between patients, with substantial fractions of Vδ1, Vδ2, and non-Vδ1 Vδ2 cells. Sequencing of the Vδ complementarity-determining region 3 showed that almost all non-Vδ1 Vδ2 cells used Vδ3 and that tumour-infiltrating γδ clonotypes were unique for every patient. Non-Vδ1Vδ2 cells from colon tumours expressed several activation markers but few NK cell receptors and exhaustion markers. In addition, mRNA analyses showed that non-Vδ1 Vδ2 cells expressed several genes for proteins with tumour-promoting functions, such as neutrophil-recruiting chemokines, Galectin 3, and transforming growth factor-beta induced. In summary, our results show a large variation in γδ T cell subsets between individual tumours, and that Vδ3 cells make up a substantial proportion of γδ T cells in colon tumours. We suggest that individual γδ T cell composition in colon tumours may contribute to the balance between favourable and adverse immune responses, and thereby also patient outcome. [ABSTRACT FROM AUTHOR]

Background: Vulvar squamous cell carcinoma (VSCC) arises after an HPV infection or the mutation of p53 or other driver genes and is treated by mutilating surgery and/or (chemo) radiation, with limited success and high morbidity. In-depth information on the immunological make up of VSCC is pivotal to assess whether immunotherapy may form an alternative treatment. Methods: A total of 104 patient samples, comprising healthy vulva (n = 27) and VSCC (n = 77), were analyzed. Multispectral immunofluorescence (15 markers) was used to study both the myeloid and lymphoid immune cell composition, and this was linked to differences in transcriptomics (NanoString nCounter, 1258 genes) and in survival (Kaplan–Meier analyses). Results: Healthy vulva and VSCC are both well infiltrated but with different subpopulations of lymphoid and myeloid cells. In contrast to the lymphoid cell infiltrate, the density and composition of the myeloid cell infiltrate strongly differed per VSCC molecular subtype. A relative strong infiltration with epithelial monocytes (HLADR−CD11c−CD14+CD68−CD163−CD33−) was prognostic for improved survival, independent of T cell infiltration, disease stage or molecular subtype. A strong infiltration with T cells and/or monocytes was associated with drastic superior survival: 5-year survival > 90% when either one is high, versus 40% when both are low (p < 0.001). Conclusion: A hot myeloid and/or lymphoid infiltrate predicts excellent survival in VSCC. Based on the response of similarly high-infiltrated other tumor types, we have started to explore the potential of neoadjuvant checkpoint blockade in VSCC. [ABSTRACT FROM AUTHOR]

T cell receptor-engineered T cells (TCR-Ts) therapy is promising for cancer immunotherapy. Most studies have focused on identifying tumor-specific T cell receptors (TCRs) through predicted tumor neoantigens. However, current algorithms for predicting tumor neoantigens are unreliable and many neoantigens are derived from non-coding regions. Thus, the technological platform for identifying tumor-specific TCRs using natural antigens expressed on tumor cells is urgently needed. In this study, tumor organoids-enriched tumor infiltrating lymphocytes (oeT) were obtained by repeatedly stimulation of autologous patient-derived organoids (PDO) in vitro. The oeT cells specifically responded to autologous tumor PDO by detecting CD137 expression and the secretion of IFN-γ using enzyme-linked immunospot assay. The measurement of oeT cell-mediated killing of three-dimensional organoids was conducted using a caspase3/7 flow cytometry assay kit. Subsequently, tumor-specific T cells were isolated based on CD137 expression and their TCRs were identified through single-cell RT-PCR analysis. The specificity cytotoxicity of TCRs were confirmed by transferring to primary peripheral blood T cells. The co-culture system proved highly effective in generating CD8+ tumor-specific oeT cells. These oeT cells effectively induced IFN-γ secretion and exhibited specificity in killing autologous tumor organoids, while not eliciting a cytotoxic response against normal organoids. The analysis conducted by TCRs revealed a significant expansion of T cells within a specific subset of TCRs. Subsequently, the TCRs were cloned and transferred to peripheral blood T cells generation engineered TCR-Ts, which adequately recognized and killed tumor cell in a patient-specific manner. The co-culture system provided an approach to generate tumor-specific TCRs from tumor-infiltrating lymphocytes of patients with colorectal cancer, and tumor-specific TCRs can potentially be used for personalized TCR-T therapy. [ABSTRACT FROM AUTHOR]

In addition to their immunosuppressive effect, cytostatics conditioning prior to adoptive therapy such as chimeric antigen receptor (CAR) T cells may play a role in debulking and remodeling the tumor microenvironment. We investigated in vitro the killing efficacy and impact of treosulfan and fludarabine on ovarian cancer cells expressing mesothelin (MSLN) and effect on MSLN-targeting CAR T cells. Treosulfan and fludarabine had a synergetic effect on killing of SKOV3 and OVCAR4 cells. Sensitivity to the combination of treosulfan and fludarabine was increased when SKOV3 cells expressed MSLN and when OVCAR4 cells were tested in hypoxia, while MSLN cells surface expression by SKOV3 and OVCAR4 cells was not altered after treosulfan or fludarabine exposure. Exposure to treosulfan or fludarabine (10 µM) neither impacted MSLN-CAR T cells degranulation, cytokines production upon challenge with MSLN + OVCAR3 cells, nor induced mitochondrial defects. Combination of treosulfan and fludarabine decreased MSLN-CAR T cells anti-tumor killing in normoxia but not hypoxia. In conclusion, treosulfan and fludarabine killed MSLN + ovarian cancer cells without altering MSLN-CAR T cells functions (at low cytostatics concentration) even in hypoxic conditions, and our data support the use of treosulfan and fludarabine as conditioning drugs prior to MSLN-CAR T cell therapy. [ABSTRACT FROM AUTHOR]

Cutaneous squamous cell carcinoma (CSCC) is the second most common malignant tumor of the skin. B7 homolog 4 (B7-H4) and B7-H5 (B7 homolog 5) are associated with a variety of tumors. Investigate the potential role of B7-H4 and B7-H5 in regulating the tumorigenesis and progression of CSCC. B7-H4 and B7-H5 transcriptome data were collected from GEO and TCGA databases and subjected to bioinformatical analysis by protein–protein interaction (PPI) network, functional enrichment analysis, immune analysis, and drug–gene interaction prediction analysis. We characterized the expression of B7-H4 and B7-H5 in carcinoma tissues of CSCC patients by immunohistochemistry. Meanwhile, the clinical correlation of B7-H4 and B7-H5 in CSCC was explored by statistical analysis. B7-H4 and B7-H5 genes were under-expressed in CSCC and correlated with tumor staging. According to GO and KEGG Pathway enrichment analysis, B7-H4, and B7-H5 can regulate the proliferation and activation of T cells, lymphocytes, and monocytes, and the expression of cytokines, such as IL-6 and IL-10, in CSCC. B7-H4 and B7-H5 are also jointly involved in the occurrence and development of CSCC via the JAK-STAT and Notch signaling pathways. We found that B7-H4 and B7-H5 proteins were abnormally highly expressed in CSCC tissue and correlated with tumor size and stage. Our findings offer new insights into the pathogenesis of CSCC and suggest that B7-H4 and B7-H5 are novel tissue biomarkers and promising therapeutic targets for CSCC. [ABSTRACT FROM AUTHOR]

In different types of cancer treatments, cancer-specific T cells are required for effective anticancer immunity, which has a central role in cancer immunotherapy. However, due to the multiple inhibitions of CD8+ T cells by tumor-related immune cells, CD8+ T-cell mediated antitumor immunotherapy has not achieved breakthrough progress in the treatment of solid tumors. Receptors for sialic acid (SA) are highly expressed in tumor-associated immune cells, so SA-modified nanoparticles are a drug delivery nanoplatform using tumor-associated immune cells as vehicles. To relieve the multiple inhibitions of CD8+ T cells by tumor-associated immune cells, we prepared SA-modified doxorubicin liposomes (SL-DOX, Scheme 1A). In our study, free SA decreased the toxicity of SL-DOX to tumor-associated immune cells. Compared with common liposomes, SL-DOX could inhibit tumor growth more effectively. It is worth noting that SL-DOX could not only kill tumor-related neutrophils and monocytes to relieve the multiple inhibitions of CD8+ T cells but also induce immunogenic death of tumor cells to promote the infiltration and differentiation of CD8+ T cells (Scheme 1B). Therefore, SL-DOX has potential value for the clinical therapeutic effect of CD8+ T cells mediating anti-tumor immunotherapy. [ABSTRACT FROM AUTHOR]

Mitochondria play a multitude of essential roles within mammalian cells, and understanding how they control immunity is an emerging area of study. Lymphocytes, as integral cellular components of the adaptive immune system, rely on mitochondria for their function, and mitochondria can dynamically instruct their differentiation and activation by undergoing rapid and profound remodelling. Energy homeostasis and ATP production are often considered the primary functions of mitochondria in immune cells; however, their importance extends across a spectrum of other molecular processes, including regulation of redox balance, signalling pathways, and biosynthesis. In this review, we explore the dynamic landscape of mitochondrial homeostasis in T and B cells, and discuss how mitochondrial disorders compromise adaptive immunity. [ABSTRACT FROM AUTHOR]

Bispecific T‐cell Engagers (TCEs) are promising anti‐cancer treatments that bind to both the CD3 receptors on T cells and an antigen on the surface of tumor cells, creating an immune synapse, leading to killing of malignant tumor cells. These novel therapies have unique development challenges, with specific safety risks of cytokine release syndrome. These on‐target adverse events fortunately can be mitigated and deconvoluted from efficacy via innovative dosing strategies, making clinical pharmacology key in the development of these therapies. This review assesses dose selection and the role of quantitative clinical pharmacology in the development of the first eight approved TCEs. Model informed drug development (MIDD) strategies can be used at every stage to guide TCE development. Mechanistic modeling approaches allow for (1) efficacious yet safe first‐in‐human dose selection as compared with in vitro minimum anticipated biological effect level (MABEL) approach; (2) rapid escalation and reducing number of patients with subtherapeutic doses through model‐based adaptive design; (3) virtual testing of different step‐up dosing regimens that may not be feasible to be evaluated in the clinic; and (4) selection and justification of the optimal clinical step‐up and full treatment doses. As the knowledge base around TCEs continues to grow, the relevance and utilization of MIDD strategies for supporting the development and dose optimization of these molecules are expected to advance, optimizing the benefit–risk profile for cancer patients. [ABSTRACT FROM AUTHOR]

Despite enormous efforts being invested in the development of novel therapies for brain malignancies, there remains a dire need for effective treatments, particularly for pediatric glioblastomas. Their poor prognosis has been attributed to the fact that conventional therapies target tumoral cells, but not glioblastoma stem cells (GSCs). GSCs are characterized by self-renewal, tumorigenicity, poor differentiation, and resistance to therapy. These characteristics represent the fundamental tools needed to recapitulate the tumor and result in a relapse. The mechanisms by which GSCs alter metabolic cues and escape elimination by immune cells are discussed in this article, along with potential strategies to harness effector immune cells against GSCs. As cellular immunotherapy is making significant advances in a variety of cancers, leveraging this underexplored reservoir may result in significant improvements in the treatment options for brain malignancies. [ABSTRACT FROM AUTHOR]

BACKGROUND: With the increasing popularity of plant protein‐based diets, soy proteins are favored as the most important source of plant protein worldwide. However, potential food allergy risks limit their use in the food industry. This work aims to reveal the mechanism of β‐conglycinin‐induced food allergy, and to explore the regulatory mechanism of heat treatment and high hydrostatic pressure (HHP) treatment in a BALB/c mouse model. RESULTS: Our results showed that oral administration of β‐conglycinin induced severe allergic symptoms in BALB/c mice, but these symptoms were effectively alleviated through heat treatment and HHP treatment. Moreover, β‐conglycinin stimulated lymphocyte proliferation and differentiation; a large number of cytokines interleukin (IL)‐4, IL‐5, IL‐10, IL‐12 and IL‐13 were released and interferon γ secretion was inhibited, which disrupted the Th1/Th2 immune balance and promoted the differentiation and proliferation of naive T cells into Th2‐type cells. CONCLUSION: Heat/non‐heat treatment altered the conformation of soybean protein, which significantly reduced allergic reactions in mice. This regulatory mechanism may be associated with Th1/Th2 immune balance. Our results provide data support for understanding the changes in allergenicity of soybean protein within the food industry. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Background: Immunoglobulin G subclass deficiencies (IgGsd) comprise a wide clinical spectrum from no symptoms to repeated respiratory infections and risk for the development of lung damage. Our aims were to investigate whether the immunological phenotype of IgGsd patients on and off immunoglobulin replacement therapy (IgRT) was reflected in the clinical features of IgGsd. Method: Thirty patients with IgGsd were included in this prospective study of 18 months of IgRT, followed by 7-18 months of IgRT discontinuation. Blood samples were collected when patients were on and off IgRT and compared with samples from 34 cross-sectional healthy controls. An in-depth lymphocyte phenotyping was performed by flow cytometry and plasma levels of immune checkpoints were assessed. Results: IgG3 subclass deficiency was most common. Patients with IgGsd had decreased levels of activated T cells and B cells and plasma levels of negative immune checkpoint molecules correlated negatively with T cell and B cell activation. The decreased T cell activation level was unaffected by IgRT, while the B cell activation was partly restored. Of note, decreased levels of activated regulatory T cells (Tregs) were found in IgGsd patients and was partly restored during IgRT. The profile of comorbidities did not associate with Treg levels. Discussion: IgGsd is associatedwith decreased B cell and T cell activation including Tregs, and increased plasma levels of negative immune checkpoint molecules. The consequence of reduced activated Tregs in IgGsd remains unclear. Decreased immune cell activation was partly restored during IgRT, demonstrating that IgRT may contribute to improved immune function in patients with IgGsd. [ABSTRACT FROM AUTHOR]

The meninges, choroid plexus (CP) and blood-brain barrier (BBB) are recognized as important gateways for peripheral immune cell trafficking into the central nervous system (CNS). Accumulation of peripheral immune cells in brain parenchyma can be observed during aging and Alzheimer's disease (AD). However, the mechanisms by which peripheral immune cells enter the CNS through these three pathways and how they interact with resident cells within the CNS to cause brain injury are not fully understood. In this paper, we review recent research on T cells recruitment in the brain during aging and AD. This review focuses on the possible pathways through which T cells infiltrate the brain, the evidence that T cells are recruited to the brain, and how infiltrating T cells interact with the resident cells in the CNS during aging and AD. Unraveling these issues will contribute to a better understanding of the mechanisms of aging and AD from the perspective of immunity, and hopefully develop new therapeutic strategies for brain aging and AD. [ABSTRACT FROM AUTHOR]

Genetic TNFAIP3 (A20) inactivation is a classical somatic lymphoma lesion and the genomic trait in haploinsufficiency of A20 (HA20). In a cohort of 34 patients with HA20, we show that heterozygous TNFAIP3 loss skews immune repertoires toward lymphocytes with classical self-reactive antigen receptors typically found in B and T cell lymphomas. This skewing was mediated by a feed-forward tumor necrosis factor (TNF)/A20/nuclear factor κB (NF-κB) loop that shaped pre-lymphoma transcriptome signatures in clonally expanded B (CD81, BACH2, and NEAT1) or T (GATA3, TOX, and PDCD1) cells. The skewing was reversed by anti-TNF treatment but could also progress to overt lymphoma. Analysis of conditional TNFAIP3 knock-out mice reproduced the wiring of the TNF/A20/NF-κB signaling axis with permissive antigen receptors and suggested a distinct regulation in B and T cells. Together, patients with the genetic disorder HA20 provide an exceptional window into A20/TNF/NF-κB-mediated control of immune homeostasis and early steps of lymphomagenesis that remain clinically unrecognized. [ABSTRACT FROM AUTHOR]

Background Methods Results Conclusions Awareness of age‐related features of carcinogenesis and the importance of cellular immunity is crucial for developing effective antitumor therapies for specific patient groups.In this study, we examined different populations of cancer stem cells (CSCs) and circulating tumor cells (CTCs) in "young" (8‐10 weeks) and "aged" (80‐82 weeks) C57BL/6 male mice. We used an orthotopic model of Lewis lung carcinoma (LLC) to evaluate the effectiveness of cell therapy targeting lung cancer through reprogrammed CD8‐positive T cells (rCD8+ T cells) in mice from two different ages.The findings revealed that tumor progression with age is primarily caused by impaired recruitment of T cells to the lungs. Additionally, a lower number of CTCs and CSCs were observed in younger mice compared to the older mice. The antitumor effect of rCD8+ T cells in aged mice was found to be inferior to that in young mice, which can be attributed to the reduced impact of therapy on specific CSCs populations.These results offer new insights into the treatment of lung cancer using rCD8+ T cells. Considering the age‐related characteristics influencing disease progression, this therapy has the potential to significantly enhance the effectiveness of treatment methods. [ABSTRACT FROM AUTHOR]

Personalized neoantigen therapy has shown long‐term and stable efficacy in specific patient populations. However, not all patients have sufficient levels of neoantigens for treatment. Although somatic mutations are commonly found in tumours, a significant portion of these mutations do not trigger an immune response. Patients with low mutation burdens continue to exhibit unresponsiveness to this treatment. We propose a design paradigm for neoantigen vaccines by utilizing the highly immunogenic unnatural amino acid p‐nitrophenylalanine (pNO2Phe) for sequence alteration of somatic mutations that failed to generate neoepitopes. This enhances the immunogenicity of the mutations and transforms it into a suitable candidate for immunotherapy. The nitrated altered epitope vaccines designed according to this paradigm is capable of activating circulating CD8+ T cells and inducing immune cross‐reactivity against autologous mutated epitopes in different MHC backgrounds (H‐2Kb, H‐2Kd, and human HLA‐A02:01), leading to the elimination of tumour cells carrying the mutation. After immunization with the altered epitopes, tumour growth was significantly inhibited. It is noteworthy that nitrated epitopes induce tumour‐infiltrating macrophages to differentiate into the M1 phenotype, surprisingly enhancing the MHC II molecule presenting pathway of macrophages. Nitrated epitope‐treated macrophages have the potential to cross‐activate CD4+ and CD8+ T cells, which may explain why pNO2Phe can enhance the immunogenicity of epitopes. Meanwhile, the immunosuppressive microenvironment of the tumour is altered due to the activation of macrophages. The nitrated neoantigen vaccine strategy enables the design of vaccines targeting non‐immunogenic tumour mutations, expanding the pool of potential peptides for personalized and shared novel antigen therapy. This approach provides treatment opportunities for patients previously ineligible for new antigen vaccine therapy. [ABSTRACT FROM AUTHOR]