TFEB regulates dendritic cell antigen presentation to modulate immune balance in asthma.

OBJECTIVE: Asthma stands as one of the most prevalent chronic respiratory conditions in children, with its pathogenesis tied to the actived antigen presentation by dendritic cells (DCs) and the imbalance within T cell subgroups. This study seeks to investigate the role of the transcription factor EB (TFEB) in modulating the antigen presentation process of DCs and its impact on the differentiation of T cell subgroups. METHODS: Bone marrow dendritic cells (BMDCs) were activated using house dust mites (HDM) and underwent RNA sequencing (RNA-seq) to pinpoint differentially expressed genes. TFEB mRNA expression levels were assessed in the peripheral blood mononuclear cells (PBMCs) of both healthy children and those diagnosed with asthma. In an asthma mouse model induced by HDM, the TFEB expression in lung tissue DCs was evaluated. Further experiments involved LV-shTFEB BMDCs co-cultured with T cells to explore the influence of TFEB on DCs' antigen presentation, T cell subset differentiation, and cytokine production. RESULTS: Transcriptomic sequencing identified TFEB as a significantly differentially expressed gene associated with immune system pathways and antigen presentation. Notably, TFEB expression showed a significant increase in the PBMCs of children diagnosed with asthma compared to healthy counterparts. Moreover, TFEB exhibited heightened expression in lung tissue DCs of HDM-induced asthmatic mice and HDM-stimulated BMDCs. Silencing TFEB resulted in the downregulation of MHC II, CD80, CD86, and CD40 on DCs. This action reinstated the equilibrium among Th1/Th2 and Th17/Treg cell subgroups, suppressed the expression of pro-inflammatory cytokines like IL-4, IL-5, IL-13, and IL-17, while augmenting the expression of the anti-inflammatory cytokine IL-10. CONCLUSION: TFEB might have a vital role in asthma's development by impacting the antigen presentation of DCs, regulating T cell subgroup differentiation, and influencing cytokine secretion. Its involvement could be pivotal in rebalancing the immune system in asthma. These research findings could potentially unveil novel therapeutic avenues for treating asthma.

Control of adaptive immunity by pattern recognition receptors.

One of the most significant conceptual advances in immunology in recent history is the recognition that signals from the innate immune system are required for induction of adaptive immune responses. Two breakthroughs were critical in establishing this paradigm: the identification of dendritic cells (DCs) as the cellular link between innate and adaptive immunity and the discovery of pattern recognition receptors (PRRs) as a molecular link that controls innate immune activation as well as DC function. Here, we recount the key events leading to these discoveries and discuss our current understanding of how PRRs shape adaptive immune responses, both indirectly through control of DC function and directly through control of lymphocyte function. In this context, we provide a conceptual framework for how variation in the signals generated by PRR activation, in DCs or other cell types, can influence T cell differentiation and shape the ensuing adaptive immune response.

Physical and functional interaction among Irf8 enhancers during dendritic cell differentiation.

The production of type 1 conventional dendritic cells (cDC1s) requires high expression of the transcription factor IRF8. Three enhancers at the Irf8 3' region function in a differentiation stage-specific manner. However, whether and how these enhancers interact physically and functionally remains unclear. Here, we show that the Irf8 3' enhancers directly interact with each other and contact the Irf8 gene body during cDC1 differentiation. The +56 kb enhancer, which functions from multipotent progenitor stages, activates the other 3' enhancers through an IRF8-dependent transcription factor program, that is, in trans. Then, the +32 kb enhancer, which operates in cDC1-committed cells, reversely acts in cis on the other 3' enhancers to maintain the high expression of Irf8. Indeed, mice with compound heterozygous deletion of the +56 and +32 kb enhancers are unable to generate cDC1s. These results illustrate how multiple enhancers cooperate to induce a lineage-determining transcription factor gene during cell differentiation.

Integrating bulk and single-cell sequencing data to construct a Scissor+ dendritic cells prognostic model for predicting prognosis and immune responses in ESCC.

Esophageal squamous cell carcinoma (ESCC) is characterized by molecular heterogeneity with various immune cell infiltration patterns, which have been associated with therapeutic sensitivity and resistance. In particular, dendritic cells (DCs) are recently discovered to be associated with prognosis and survival in cancer. However, how DCs differ among ESCC patients has not been fully comprehended. Recently, the advance of single-cell RNA sequencing (scRNA-seq) enables us to profile the cell types, states, and lineages in the heterogeneous ESCC tissues. Here, we dissect the ESCC tumor microenvironment at high resolution by integrating 192,078 single cells from 60 patients, including 4379 DCs. We then used Scissor, a method that identifies cell subpopulations from single-cell data that are associated bulk samples with genomic and clinical information, to stratify DCs into Scissorhi and Scissorlow subtypes. We applied the Scissorhi gene signature to stratify ESCC scRNAseq patient, and we found that PD-L1, TIGIT, PVR and IL6 ligand-receptor-mediated cell interactions existed mainly in Scissorhi patients. Finally, based on the Scissor results, we successfully developed a validated prognostic risk model for ESCC and further validated the reliability of the risk prediction model by recruiting 40 ESCC clinical patients. This information highlights the importance of these genes in assessing patient prognosis and may help in the development of targeted or personalized therapies for ESCC.

Regulation of Tumor Dendritic Cells by Programmed Cell Death 1 Pathways.

The advent of immune checkpoint blockade therapy has revolutionized cancer treatments and is partly responsible for the significant decline in cancer-related mortality observed during the last decade. Immune checkpoint inhibitors, such as anti-programmed cell death 1 (PD-1)/programmed cell death ligand 1 (PD-L1), have demonstrated remarkable clinical successes in a subset of cancer patients. However, a considerable proportion of patients remain refractory to immune checkpoint blockade, prompting the exploration of mechanisms of treatment resistance. Whereas much emphasis has been placed on the role of PD-L1 and PD-1 in regulating the activity of tumor-infiltrating T cells, recent studies have now shown that this immunoregulatory axis also directly regulates myeloid cell activity in the tumor microenvironment including tumor-infiltrating dendritic cells. In this review, I discuss the most recent advances in the understanding of how PD-1, PD-L1, and programmed cell death ligand 2 regulate the function of tumor-infiltrating dendritic cells, emphasizing the need for further mechanistic studies that could facilitate the development of novel combination immunotherapies for improved cancer patient benefit.

Checkpoint inhibitor-expressing lentiviral vaccine suppresses tumor growth in preclinical cancer models.

BACKGROUND: While immunotherapy has been highly successful for the treatment of some cancers, for others, the immune response to tumor antigens is weak leading to treatment failure. The resistance of tumors to checkpoint inhibitor therapy may be caused by T cell exhaustion resulting from checkpoint activation. METHODS: In this study, lentiviral vectors that expressed T cell epitopes of an experimentally introduced tumor antigen, ovalbumin, or the endogenous tumor antigen, Trp1 were developed. The vectors coexpressed CD40 ligand (CD40L), which served to mature the dendritic cells (DCs), and a soluble programmed cell death protein 1 (PD-1) microbody to prevent checkpoint activation. Vaccination of mice bearing B16.OVA melanomas with vector-transduced DCs induced the proliferation and activation of functional, antigen-specific, cytolytic CD8 T cells. RESULTS: Vaccination induced the expansion of CD8 T cells that infiltrated the tumors to suppress tumor growth. Vector-encoded CD40L and PD-1 microbody increased the extent of tumor growth suppression. Adoptive transfer demonstrated that the effect was mediated by CD8 T cells. Direct injection of the vector, without the need for ex vivo transduction of DCs, was also effective. CONCLUSIONS: This study suggests that therapeutic vaccination that induces tumor antigen-specific CD8 T cells coupled with a vector-expressed checkpoint inhibitor can be an effective means to suppress the growth of tumors that are resistant to conventional immunotherapy.

The functional role of L-fucose on dendritic cell function and polarization.

Despite significant advances in the development and refinement of immunotherapies administered to combat cancer over the past decades, a number of barriers continue to limit their efficacy. One significant clinical barrier is the inability to mount initial immune responses towards the tumor. As dendritic cells are central initiators of immune responses in the body, the elucidation of mechanisms that can be therapeutically leveraged to enhance their functions to drive anti-tumor immune responses is urgently needed. Here, we report that the dietary sugar L-fucose can be used to enhance the immunostimulatory activity of dendritic cells (DCs). L-fucose polarizes immature myeloid cells towards specific DC subsets, specifically cDC1 and moDC subsets. In vitro, L-fucose treatment enhances antigen uptake and processing of DCs. Furthermore, our data suggests that L-fucose-treated DCs increase stimulation of T cell populations. Consistent with our functional assays, single-cell RNA sequencing of intratumoral DCs from melanoma- and breast tumor-bearing mice confirmed transcriptional regulation and antigen processing as pathways that are significantly altered by dietary L-fucose. Together, this study provides the first evidence of the ability of L-fucose to bolster DC functionality and provides rational to further investigate how L-fucose can be used to leverage DC function in order to enhance current immunotherapy.

Circulating immunome fingerprint in eosinophilic esophagitis is associated with clinical response to proton pump inhibitor treatment.

Objectives: The aim of the study was to characterize the circulating immunome of patients with EoE before and after proton pump inhibitor (PPI) treatment in order to identify potential non-invasive biomarkers of treatment response. Methods: PBMCs from 19 healthy controls and 24 EoE patients were studied using a 39-plex spectral cytometry panel. The plasmacytoid dendritic cell (pDC) population was differentially characterized by spectral cytometry analysis and immunofluorescence assays in esophageal biopsies from 7 healthy controls and 13 EoE patients. Results: Interestingly, EoE patients at baseline had lower levels of circulating pDC compared with controls. Before treatment, patients with EoE who responded to PPI therapy had higher levels of circulating pDC and classical monocytes, compared with non-responders. Moreover, following PPI therapy pDC levels were increased in all EoE patients, while normal levels were only restored in PPI-responding patients. Finally, circulating pDC levels inversely correlated with peak eosinophil count and pDC count in esophageal biopsies. The number of tissue pDCs significantly increased during active EoE, being even higher in non-responder patients when compared to responder patients pre-PPI. pDC levels decreased after PPI intake, being further restored almost to control levels in responder patients post-PPI. Conclusions: We hereby describe a unique immune fingerprint of EoE patients at diagnosis. Moreover, circulating pDC may be also used as a novel non-invasive biomarker to predict subsequent response to PPI treatment.

Neem leaf glycoprotein binding to Dectin-1 receptors on dendritic cell induces type-1 immunity through CARD9 mediated intracellular signal to NFκB.

BACKGROUND: A water-soluble ingredient of mature leaves of the tropical mahogany 'Neem' (Azadirachta indica), was identified as glycoprotein, thus being named as 'Neem Leaf Glycoprotein' (NLGP). This non-toxic leaf-component regressed cancerous murine tumors (melanoma, carcinoma, sarcoma) recurrently in different experimental circumstances by boosting prime antitumor immune attributes. Such antitumor immunomodulation, aid cytotoxic T cell (Tc)-based annihilation of tumor cells. This study focused on identifying and characterizing the signaling gateway that initiate this systemic immunomodulation. In search of this gateway, antigen-presenting cells (APCs) were explored, which activate and induce the cytotoxic thrust in Tc cells. METHODS: Six glycoprotein-binding C-type lectins found on APCs, namely, MBR, Dectin-1, Dectin-2, DC-SIGN, DEC205 and DNGR-1 were screened on bone marrow-derived dendritic cells from C57BL/6 J mice. Fluorescence microscopy, RT-PCR, flow cytometry and ELISA revealed Dectin-1 as the NLGP-binding receptor, followed by verifications through RNAi. Following detection of ß-Glucans in NLGP, their interactions with Dectin-1 were explored in silico. Roles of second messengers and transcription factors in the downstream signal were studied by co-immunoprecipitation, western blotting, and chromatin-immunoprecipitation. Intracellularization of FITC-coupled NLGP was observed by processing confocal micrographs of DCs. RESULTS: Considering extents of hindrance in NLGP-driven transcription rates of the cytokines IL-10 and IL-12p35 by receptor-neutralization, Dectin-1 receptors on dendritic cells were found to bind NLGP through the ligand's peripheral ß-Glucan chains. The resulting signal phosphorylates PKCδ, forming a trimolecular complex of CARD9, Bcl10 and MALT1, which in turn activates the canonical NFκB-pathway of transcription-regulation. Consequently, the NFκB-heterodimer p65:p50 enhances Il12a transcription and the p50:p50 homodimer represses Il10 transcription, bringing about a cytokine-based systemic-bias towards type-1 immune environment. Further, NLGP gets engulfed within dendritic cells, possibly through endocytic activities of Dectin-1. CONCLUSION: NLGP's binding to Dectin-1 receptors on murine dendritic cells, followed by the intracellular signal, lead to NFκB-mediated contrasting regulation of cytokine-transcriptions, initiating a pro-inflammatory immunopolarization, which amplifies further by the responding immune cells including Tc cells, alongside their enhanced cytotoxicity. These insights into the initiation of mammalian systemic immunomodulation by NLGP at cellular and molecular levels, may help uncovering its mode of action as a novel immunomodulator against human cancers, following clinical trials.

The role of dendritic cells in MASH: friends or foes?

Dendritic cells (DCs) are major antigen-presenting cells that connect innate and adaptive immunity. Hepatic DCs are less activated and contribute to maintain the tolerogenic environment of the liver under steady state. Several studies indicated DCs in metabolic dysfunction-associated steatohepatitis (MASH), representing a substantial burden on healthcare systems due to its association with liver-related morbidity and mortality. Studies highlighted the potential disease-promoting role of liver DCs in the development of MASH while other experimental systems suggested their protective role. This review discusses this controversy and the current understanding of how DCs affect the pathogenesis of MASH.

Glioblastoma-instructed microglia transition to heterogeneous phenotypic states with phagocytic and dendritic cell-like features in patient tumors and patient-derived orthotopic xenografts.

BACKGROUND: A major contributing factor to glioblastoma (GBM) development and progression is its ability to evade the immune system by creating an immune-suppressive environment, where GBM-associated myeloid cells, including resident microglia and peripheral monocyte-derived macrophages, play critical pro-tumoral roles. However, it is unclear whether recruited myeloid cells are phenotypically and functionally identical in GBM patients and whether this heterogeneity is recapitulated in patient-derived orthotopic xenografts (PDOXs). A thorough understanding of the GBM ecosystem and its recapitulation in preclinical models is currently missing, leading to inaccurate results and failures of clinical trials. METHODS: Here, we report systematic characterization of the tumor microenvironment (TME) in GBM PDOXs and patient tumors at the single-cell and spatial levels. We applied single-cell RNA sequencing, spatial transcriptomics, multicolor flow cytometry, immunohistochemistry, and functional studies to examine the heterogeneous TME instructed by GBM cells. GBM PDOXs representing different tumor phenotypes were compared to glioma mouse GL261 syngeneic model and patient tumors. RESULTS: We show that GBM tumor cells reciprocally interact with host cells to create a GBM patient-specific TME in PDOXs. We detected the most prominent transcriptomic adaptations in myeloid cells, with brain-resident microglia representing the main population in the cellular tumor, while peripheral-derived myeloid cells infiltrated the brain at sites of blood-brain barrier disruption. More specifically, we show that GBM-educated microglia undergo transition to diverse phenotypic states across distinct GBM landscapes and tumor niches. GBM-educated microglia subsets display phagocytic and dendritic cell-like gene expression programs. Additionally, we found novel microglial states expressing cell cycle programs, astrocytic or endothelial markers. Lastly, we show that temozolomide treatment leads to transcriptomic plasticity and altered crosstalk between GBM tumor cells and adjacent TME components. CONCLUSIONS: Our data provide novel insights into the phenotypic adaptation of the heterogeneous TME instructed by GBM tumors. We show the key role of microglial phenotypic states in supporting GBM tumor growth and response to treatment. Our data place PDOXs as relevant models to assess the functionality of the TME and changes in the GBM ecosystem upon treatment.

Obese visceral adipose dendritic cells downregulate regulatory T cell development through IL-33.

Regulatory T cells (Tregs) residing in visceral adipose tissue (VAT) play a pivotal role in regulating tissue inflammation and metabolic dysfunction associated with obesity. However, the specific phenotypic and functional characteristics of Tregs in obese VAT, as well as the regulatory mechanisms shaping them, remain elusive. This study demonstrates that obesity selectively reduces Tregs in VAT, characterized by restrained proliferation, heightened PD-1 expression, and diminished ST2 expression. Additionally, obese VAT displays distinctive maturation of dendritic cells (DCs), marked by elevated expressions of MHC-II, CD86, and PD-L1, which are inversely correlated with VAT Tregs. In an in vitro co-culture experiment, only obese VAT DCs, not macrophages or DCs from subcutaneous adipose tissue (SAT) and spleen, result in decreased Treg differentiation and proliferation. Furthermore, Tregs differentiated by obese VAT DCs exhibit distinct characteristics resembling those of Tregs in obese VAT, such as reduced ST2 and IL-10 expression. Mechanistically, obesity lowers IL-33 production in VAT DCs, contributing to the diminished Treg differentiation. These findings collectively underscore the critical role of VAT DCs in modulating Treg generation and shaping Treg phenotype and function during obesity, potentially contributing to the regulation of VAT Treg populations.

Immune microniches shape intestinal Treg function.

The intestinal immune system is highly adapted to maintaining tolerance to the commensal microbiota and self-antigens while defending against invading pathogens1,2. Recognizing how the diverse network of local cells establish homeostasis and maintains it in the complex immune environment of the gut is critical to understanding how tolerance can be re-established following dysfunction, such as in inflammatory disorders. Although cell and molecular interactions that control T regulatory (Treg) cell development and function have been identified3,4, less is known about the cellular neighbourhoods and spatial compartmentalization that shapes microorganism-reactive Treg cell function. Here we used in vivo live imaging, photo-activation-guided single-cell RNA sequencing5-7 and spatial transcriptomics to follow the natural history of T cells that are reactive towards Helicobacter hepaticus through space and time in the settings of tolerance and inflammation. Although antigen stimulation can occur anywhere in the tissue, the lamina propria-but not embedded lymphoid aggregates-is the key microniche that supports effector Treg (eTreg) cell function. eTreg cells are stable once their niche is established; however, unleashing inflammation breaks down compartmentalization, leading to dominance of CD103+SIRPα+ dendritic cells in the lamina propria. We identify and validate the putative tolerogenic interaction between CD206+ macrophages and eTreg cells in the lamina propria and identify receptor-ligand pairs that are likely to govern the interaction. Our results reveal a spatial mechanism of tolerance in the lamina propria and demonstrate how knowledge of local interactions may contribute to the next generation of tolerance-inducing therapies.

Sexual dimorphism in skin immunity is mediated by an androgen-ILC2-dendritic cell axis.

Males and females exhibit profound differences in immune responses and disease susceptibility. However, the factors responsible for sex differences in tissue immunity remain poorly understood. Here, we uncovered a dominant role for type 2 innate lymphoid cells (ILC2s) in shaping sexual immune dimorphism within the skin. Mechanistically, negative regulation of ILC2s by androgens leads to a reduction in dendritic cell accumulation and activation in males, along with reduced tissue immunity. Collectively, our results reveal a role for the androgen-ILC2-dendritic cell axis in controlling sexual immune dimorphism. Moreover, this work proposes that tissue immune set points are defined by the dual action of sex hormones and the microbiota, with sex hormones controlling the strength of local immunity and microbiota calibrating its tone.

Single-cell sequencing analysis and multiple machine-learning models revealed the cellular crosstalk of dendritic cells and identified FABP5 and KLRB1 as novel biomarkers for psoriasis.

Background: Psoriasis is an immune-mediated disorder influenced by environmental factors on a genetic basis. Despite advancements, challenges persist, including the diminishing efficacy of biologics and small-molecule targeted agents, alongside managing recurrence and psoriasis-related comorbidities. Unraveling the underlying pathogenesis and identifying valuable biomarkers remain pivotal for diagnosing and treating psoriasis. Methods: We employed a series of bioinformatics (including single-cell sequencing data analysis and machine learning techniques) and statistical methods to integrate and analyze multi-level data. We observed the cellular changes in psoriatic skin tissues, screened the key genes Fatty acid binding protein 5 (FABP5) and The killer cell lectin-like receptor B1 (KLRB1), evaluated the efficacy of six widely prescribed drugs on psoriasis treatment in modulating the dendritic cell-associated pathway, and assessed their overall efficacy. Finally, RT-qPCR, immunohistochemistry, and immunofluorescence assays were used to validate. Results: The regulatory influence of dendritic cells (DCs) on T cells through the CD70/CD27 signaling pathway may emerge as a significant facet of the inflammatory response in psoriasis. Notably, FABP5 and KLRB1 exhibited up-regulation and co-localization in psoriatic skin tissues and M5-induced HaCaT cells, serving as potential biomarkers influencing psoriasis development. Conclusion: Our study analyzed the impact of DC-T cell crosstalk in psoriasis, elucidated the characterization of two biomarkers, FABP5 and KLRB1, in psoriasis, and highlighted the promise and value of tofacitinib in psoriasis therapy targeting DCs.

Genome-wide screening identifies Trim33 as an essential regulator of dendritic cell differentiation.

The development of dendritic cells (DCs), including antigen-presenting conventional DCs (cDCs) and cytokine-producing plasmacytoid DCs (pDCs), is controlled by the growth factor Flt3 ligand (Flt3L) and its receptor Flt3. We genetically dissected Flt3L-driven DC differentiation using CRISPR-Cas9-based screening. Genome-wide screening identified multiple regulators of DC differentiation including subunits of TSC and GATOR1 complexes, which restricted progenitor growth but enabled DC differentiation by inhibiting mTOR signaling. An orthogonal screen identified the transcriptional repressor Trim33 (TIF-1γ) as a regulator of DC differentiation. Conditional targeting in vivo revealed an essential role of Trim33 in the development of all DCs, but not of monocytes or granulocytes. In particular, deletion of Trim33 caused rapid loss of DC progenitors, pDCs, and the cross-presenting cDC1 subset. Trim33-deficient Flt3+ progenitors up-regulated pro-inflammatory and macrophage-specific genes but failed to induce the DC differentiation program. Collectively, these data elucidate mechanisms that control Flt3L-driven differentiation of the entire DC lineage and identify Trim33 as its essential regulator.

Analysis of Human Peripheral Blood Mononuclear Cells in Patients with Neurodegenerative Diseases.

The role of immune system in the progression of neurodegenerative diseases has been studied for decades in animal models. However, invasive studies in human subjects remain controversial due to the heterogeneity of the presentation of different diagnostic categories at different stages of the disease. Peripheral blood mononuclear cells (PBMCs) contain immune cells including dendritic cells (DCs), monocytes, macrophages, and T lymphocytes. Isolating PBMCs from whole blood samples collected from patients provides a minimally invasive method for analyzing the immune system's function in patients with neurodegenerative diseases. By isolating single cell types from patients' peripheral blood, in vitro analyses can be conducted including RNA sequencing, immunofluorescence, and phagocytic analysis. In this chapter, we discuss PBMC separation and isolation of macrophages in pure culture in vitro. We also outline methods for performing RNA-seq on cultured macrophages and other techniques for investigating the role of macrophages in neurodegenerative disease pathophysiology.

A Case of Blastic Plasmacytoid Dendritic Cell Neoplasm with Orbital Tumor as the Initial Symptom.

BACKGROUND: Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematologic malignancy arising from precursor dendritic cells. It is a rare and challenging clinical presentation. For decades, there has been no treatment course for managing BPDCN and its overall prognosis is poor. METHODS AND RESULTS: We report a 27-year-old man who was admitted to the hospital due to an orbital tumor as the first symptom. Progressive enlargement of the orbital tumor was accompanied by multiple purple circular nodules on the body trunk. Pathological confirmation of BPDCN after resection of the orbital mass. Bone marrow smear and flow cytometry on examination indicate AML-M5. Performance of chemotherapy and peripheral blood autologous stem cell transplantation. CONCLUSIONS: The clinical manifestations of blastic plasmacytoid dendritic cell neoplasms are diverse. The diagnosis of BPDCN can be difficult due to overlapping morphologic, immunophenotypic, and clinical features of other hematologic AML. Relapsed and refractory BPDCN remains an elusive therapeutic challenge. The future of new targeted therapeutic drugs is expected.

Dendritic cell targeting peptide plus Salmonella FliCd flagellin fused outer membrane protein H (OmpH) demonstrated increased efficacy against infections caused by different Pasteurella multocida serogroups in mouse models.

As the major outer membrane protein (OMP) presents in the Pasteurella multocida envelope, OmpH was frequently expressed for laboratory assessments of its immunogenicity against P. multocida infections, but the results are not good. In this study, we modified OmpH with dendritic cell targeting peptide (Depeps) and/or Salmonella FliCd flagellin, and expressed three types of recombinant proteins with the MBP tag (rDepeps-FliC-OmpH-MBP, rDepeps-OmpH-MBP, rFliC-OmpH-MBP). Assessments in mouse models revealed that vaccination with rDepeps-FliC-OmpH-MBP, rDepeps-OmpH-MBP, or rFliC-OmpH-MBP induced significant higher level of antibodies as well as IFN-γ and IL-4 in murine sera than vaccination with rOmpH-MBP (P < 0.5). Vaccination with the three modified proteins also provided increased protection (rDepeps-FliC-OmpH-MBP, 70 %; rDepeps-OmpH-MBP, 50 %; rFliC-OmpH-MBP, 60 %) against P. multocida serotype D compared to vaccination with rOmpH-MBP (30 %). In mice vaccinated with different types of modified OmpHs, a significantly decreased bacterial strains were recovered from bloods, lungs, and spleens compared to rOmpH-MBP-vaccinated mice (P < 0.5). Notably, our assessments also demonstrated that vaccination with rDepeps-FliC-OmpH-MBP provided good protection against infections caused by a heterogeneous group of P. multocida serotypes (A, B, D). Our above findings indicate that modification with DCpep and Salmonella flagellin could be used as a promising strategy to improve vaccine effectiveness.

The immunologic abnormalities in patients with paroxysmal nocturnal hemoglobinuria are associated with disease progression.

OBJECTIVES: To suggest the presence of a hyperimmune state in patients, and indicate that immune system attack on glycosylphosphatidylinositol (+) (GPI+) cells while escaping GPI- cell immunity. METHODS: We retrospective the immune cell subtypes in peripheral blood from 25 patients visiting Tianjin Medical University General Hospital, Tianjin, China, with classical paroxysmal nocturnal hemoglobinuria (PNH) and 50 healthy controls. RESULTS: The total CD3+ and CD3+CD8+ cell levels were higher in patients with PNH. The CD3+ cells are positively, correlated with lactate dehydrogenase (LDH; r=0.5453, p=0.0040), indirect bilirubin (r=0.4260, p=0.0379) and Flear- cells in monocytes (r=0.4099, p=0.0303). However, a negative correlation was observed between CD3+ cells and hemoglobin (r= -0.4530, p=0.0105). The total CD19+ cells decreased in patients, and CD19+ cells were negatively correlated with LDH (r= -0.5640, p=0.0077) and Flear- cells in monocytes (r= -0.4432, p=0.0341). Patients showed an increased proportion of total dendritic cells (DCs), with a higher proportion of myeloid DCs (mDCs) within the DC population. Moreover, the proportion of mDC/DC was positively correlated with CD59- cells (II + III types) in red cells (r=0.7941, p=0.0004), Flear- cells in granulocytes (r=0.5357, p=0.0396), and monocytes (r=0.6445, p=0.0095). CONCLUSION: Our results demonstrated that immune abnormalities are associated with PNH development.