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Nature Communications Mar 2024IgG4-related disease (IgG4-RD) has complex clinical manifestations ranging from fibrosis and inflammation to deregulated metabolism. The molecular mechanisms...
IgG4-related disease (IgG4-RD) has complex clinical manifestations ranging from fibrosis and inflammation to deregulated metabolism. The molecular mechanisms underpinning these phenotypes are unclear. In this study, by using IgG4-RD patient peripheral blood mononuclear cells (PBMCs), IgG4-RD cell lines and Usp25 knockout mice, we show that ubiquitin-specific protease 25 (USP25) engages in multiple pathways to regulate fibrotic and inflammatory pathways that are characteristic to IgG4-RD. Reduced USP25 expression in IgG4-RD leads to increased SMAD3 activation, which contributes to fibrosis and induces inflammation through the IL-1β inflammatory axis. Mechanistically, USP25 prevents ubiquitination of RAC1, thus, downregulation of USP25 leads to ubiquitination and degradation of RAC1. Decreased RAC1 levels result in reduced aldolase A release from the actin cytoskeleton, which then lowers glycolysis. The expression of LYN, a component of the B cell receptor signalosome is also reduced in USP25-deficient B cells, which might result in B cell activation deficiency. Altogether, our results indicate a potential anti-inflammatory and anti-fibrotic role for USP25 and make USP25 a promising diagnostic marker and potential therapeutic target in IgG4-RD.
Topics: Animals; Humans; Mice; B-Lymphocytes; Fibrosis; Immunoglobulin G4-Related Disease; Inflammation; Leukocytes, Mononuclear; Ubiquitin Thiolesterase
PubMed: 38521787
DOI: 10.1038/s41467-024-45977-7 -
Frontiers in Immunology 2023Nephrotic syndrome (NS) is a relatively rare and serious presentation of IgA nephropathy (IgAN) (NS-IgAN). Previous research has suggested that the pathogenesis of...
Nephrotic syndrome (NS) is a relatively rare and serious presentation of IgA nephropathy (IgAN) (NS-IgAN). Previous research has suggested that the pathogenesis of NS-IgAN may involve circulating immune imbalance and kidney injury; however, this has yet to be fully elucidated. To investigate the cellular and molecular status of NS-IgAN, we performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) and kidney cells from pediatric patients diagnosed with NS-IgAN by renal biopsy. Consistently, the proportion of intermediate monocytes (IMs) in NS-IgAN patients was higher than in healthy controls. Furthermore, flow cytometry confirmed that IMs were significantly increased in pediatric patients with NS. The characteristic expression of and MHC class II molecules and an increase in oxidative phosphorylation may be important features of IMs in NS-IgAN. Notably, we found that the expression level of was significantly increased in the CMs, IMs, and NCMs of patients with NS-IgAN. This may be related to kidney injury. Regulatory T cells (Tregs) are classified into two subsets of cells: Treg1 ( , , and ) and Treg2 ( , , and ). We found that the levels of Treg2 cells expressed significant levels of and , which may be related to the recovery of kidney injury. The state of NS in patients was closely related to podocyte injury. The expression levels of , , and genes related to epithelial-mesenchymal transition were significantly increased in podocytes from NS-IgAN patients. These represent key features of podocyte injury. Our analysis suggests that is significantly downregulated following injury and may represent a new marker for podocytes. In this study, we systematically analyzed molecular events in the circulatory system and kidney tissue of pediatric patients with NS-IgAN, which provides new insights for targeted therapy in the future.
Topics: Humans; Child; Glomerulonephritis, IGA; Nephrotic Syndrome; Leukocytes, Mononuclear; Receptors, CCR7; Kidney; HLA-DR Antigens
PubMed: 37908345
DOI: 10.3389/fimmu.2023.1231937 -
Frontiers in Immunology 2023Tumor-infiltrating myeloid cells (TIMs) are key regulators in tumor progression, but the similarity and distinction of their fundamental properties in pancreatic ductal...
INTRODUCTION
Tumor-infiltrating myeloid cells (TIMs) are key regulators in tumor progression, but the similarity and distinction of their fundamental properties in pancreatic ductal adenocarcinoma (PDAC) remain elusive.
METHOD
In this study, we conducted scRNA-seq data analysis of cells from 12 primary tumor (PT) tissues, 4 metastatic (Met) tumor tissues, 3 adjacent normal pancreas tissues (Para), and PBMC samples across 16 PDAC patients, and revealed a heterogeneous TIMs environment in PDAC.
RESULT
Systematic comparisons between tumor and non-tumor samples of myeloid lineages identified 10 necroptosis-associated genes upregulated in PDAC tumors compared to 5 upregulated in paratumor or healthy peripheral blood. A novel RTM (resident tissue macrophages), GLUL-SQSTM1- RTM, was found to act as a positive regulator of immunity. Additionally, HSP90AA1+HSP90AB1+ mast cells exhibited pro-immune characteristics, and JAK3+TLR4+ CD16 monocytes were found to be anti-immune. The findings were validated through clinical outcomes and cytokines analyses. Lastly, intercellular network reconstruction supported the associations between the identified novel clusters, cancer cells, and immune cell populations.
CONCLUSION
Our analysis comprehensively characterized major myeloid cell lineages and identified three subsets of myeloid-derived cells associated with necroptosis. These findings not only provide a valuable resource for understanding the multi-dimensional characterization of the tumor microenvironment in PDAC but also offer valuable mechanistic insights that can guide the design of effective immuno-oncology treatment strategies.
Topics: Humans; Cell Lineage; Single-Cell Gene Expression Analysis; Leukocytes, Mononuclear; Necroptosis; Pancreatic Neoplasms; Carcinoma, Pancreatic Ductal; Tumor Microenvironment
PubMed: 38149248
DOI: 10.3389/fimmu.2023.1263633 -
Inflammation Oct 2023Atherosclerosis (AS) is the main cause of cerebrovascular diseases, and macrophages play important roles in atherosclerosis. DExH-Box helicase 9 (DHX9), as a member of...
Atherosclerosis (AS) is the main cause of cerebrovascular diseases, and macrophages play important roles in atherosclerosis. DExH-Box helicase 9 (DHX9), as a member of DExD/H-box RNA helicase superfamily II, is identified as an autoantigen in the sera of systemic lupus erythematosus patients to trigger inflammation. The aim of this study was to investigate whether DHX9 is involved in AS development, especially in macrophages-mediated-inflammatory responses. We find that DHX9 expression is significantly increased in oxLDL or interferon-γ-treated macrophages and peripheral blood mononuclear cells (PBMCs) from patients with coronary artery disease (CAD). Knockdown of DHX9 inhibits lipid uptake and pro-inflammatory factors expression in macrophages, and ameliorates TNF-α-mediated monocyte adhesion capacity. Furthermore, we find that oxLDL stimulation promotes DHX9 interaction with p65 in macrophages, and further enhances the transcriptional activity of DHX9-p65-RNA Polymerase II complex to produce inflammatory factors. Moreover, using ApoE -/- mice fed with western diet to establish AS model, we find that knockdown of DHX9 mediated by adeno-associated virus-Sh-DHX9 through tail vein injection evidently alleviates AS progression in vivo. Finally, we also find that knockdown of DHX9 inhibits p65 activation, inflammatory factors expression, and the transcriptional activity of p65-RNA Polymerase II complex in PBMCs from patients with CAD. Overall, these results indicate that DHX9 promotes AS progression by enhancing inflammation in macrophages, and suggest DHX9 as a potential target for developing therapeutic drug.
Topics: Humans; Mice; Animals; DEAD-box RNA Helicases; Leukocytes, Mononuclear; RNA Polymerase II; Mice, Knockout, ApoE; Macrophages; Inflammation; Atherosclerosis; Neoplasm Proteins
PubMed: 37326773
DOI: 10.1007/s10753-023-01836-z -
Frontiers in Immunology 2023Despite treatment, hepatitis B surface antigen (HBsAg) persists in patients with chronic hepatitis B (CHB), suggesting the likely presence of the virus in the body. CD8...
Despite treatment, hepatitis B surface antigen (HBsAg) persists in patients with chronic hepatitis B (CHB), suggesting the likely presence of the virus in the body. CD8 T cell responses are essential for managing viral replication, but their effect on HBsAg levels remains unclear. We studied the traits of activated CD8 T cells and HBV-specific CD8 T cells in the blood of CHB patients undergoing nucleos(t)ide analog (NUC) therapy. For the transcriptome profiling of activated CD8 T cells in peripheral blood mononuclear cells (PBMCs), CD69 CD8 T cells were sorted from six donors, and single-cell RNA sequencing (scRNA-seq) analysis was performed. To detect HBV-specific CD8 T cells, we stimulated PBMCs from 26 donors with overlapping peptides covering the HBs, HBcore, and HBpol regions of genotype A/B/C viruses, cultured for 10 days, and analyzed via multicolor flow cytometry. scRNA-seq data revealed that CD8 T cell clusters harboring the transcripts involved in the cytolytic functions were frequently observed in donors with high HBsAg levels. Polyfunctional analysis of HBV-specific CD8 T cells utilized by IFN-γ/TNFα/CD107A/CD137 revealed that HBcore-specific cells exhibited greater polyfunctionality, suggesting that the quality of HBV-specific CD8 T cells varies among antigens. Moreover, a subset of HBcore-specific CD8 T cells with lower cytolytic potential was inversely correlated with HBsAg level. Our results revealed a stimulant-dependent qualitative difference in HBV-specific CD8 T cells in patients with CHB undergoing NUC therapy. Hence, the induction of HBcore-specific CD8 T cells with lower cytolytic potential could be a new target for reducing HBsAg levels.
Topics: Humans; Hepatitis B Surface Antigens; CD8-Positive T-Lymphocytes; Hepatitis B virus; Leukocytes, Mononuclear; Hepatitis B, Chronic
PubMed: 37920475
DOI: 10.3389/fimmu.2023.1257113 -
Journal of Translational Medicine Oct 2023Even after 3 years from SARS-CoV-2 identification, COVID-19 is still a persistent and dangerous global infectious disease. Significant improvements in our understanding...
BACKGROUND
Even after 3 years from SARS-CoV-2 identification, COVID-19 is still a persistent and dangerous global infectious disease. Significant improvements in our understanding of the disease pathophysiology have now been achieved. Nonetheless, reliable and accurate biomarkers for the early stratification of COVID-19 severity are still lacking. Long noncoding RNAs (LncRNAs) are ncRNAs longer than 200 nucleotides, regulating the transcription and translation of protein-coding genes and they can be found in the peripheral blood, thus holding a promising biomarker potential. Specifically, peripheral blood mononuclear cells (PBMCs) have emerged as a source of indirect biomarkers mirroring the conditions of tissues: they include monocytes, B and T lymphocytes, and natural killer T cells (NKT), being highly informative for immune-related events.
METHODS
We profiled by RNA-Sequencing a panel of 2906 lncRNAs to investigate their modulation in PBMCs of a pilot group of COVID-19 patients, followed by qPCR validation in 111 hospitalized COVID-19 patients.
RESULTS
The levels of four lncRNAs were found to be decreased in association with COVID-19 mortality and disease severity: HLA Complex Group 18-242 and -244 (HCG18-242 and HCG18-244), Lymphoid Enhancer Binding Factor 1-antisense 1 (LEF1-AS1) and lncCEACAM21 (i.e. ENST00000601116.5, a lncRNA in the CEACAM21 locus). Interestingly, these deregulations were confirmed in an independent patient group of hospitalized patients and by the re-analysis of publicly available single-cell transcriptome datasets. The identified lncRNAs were expressed in all of the PBMC cell types and inversely correlated with the neutrophil/lymphocyte ratio (NLR), an inflammatory marker. In vitro, the expression of LEF1-AS1 and lncCEACAM21 was decreased upon THP-1 monocytes exposure to a relevant stimulus, hypoxia.
CONCLUSION
The identified COVID-19-lncRNAs are proposed as potential innovative biomarkers of COVID-19 severity and mortality.
Topics: Humans; COVID-19; Leukocytes, Mononuclear; RNA, Long Noncoding; SARS-CoV-2; Biomarkers; Patient Acuity
PubMed: 37884975
DOI: 10.1186/s12967-023-04497-6 -
Immunological link between periodontitis and type 2 diabetes deciphered by single-cell RNA analysis.Clinical and Translational Medicine Dec 2023Type 2 diabetes mellitus (DM) is a complex metabolic disorder that causes various complications, including periodontitis (PD). Although a bidirectional relationship has...
BACKGROUND
Type 2 diabetes mellitus (DM) is a complex metabolic disorder that causes various complications, including periodontitis (PD). Although a bidirectional relationship has been reported between DM and PD, their immunological relationship remains poorly understood. Therefore, this study aimed to compare the immune response in patients with PD alone and in those with both PD and DM (PDDM) to expand our knowledge of the complicated connection between PD and DM.
METHODS
Peripheral blood mononuclear cells were collected from 11 healthy controls, 10 patients with PD without DM, and six patients with PDDM, followed by analysis using single-cell RNA sequencing. The differences among groups were then compared based on intracellular and intercellular perspectives.
RESULTS
Compared to the healthy state, classical monocytes exhibited the highest degree of transcriptional change, with elevated levels of pro-inflammatory cytokines in both PD and PDDM. DM diminished the effector function of CD8+ T and natural killer (NK) cells as well as completely modified the differentiation direction of these cells. Interestingly, a prominent pathway, RESISTIN, which is known to increase insulin resistance and susceptibility to diabetes, was found to be activated under both PD and PDDM conditions. In particular, CAP1+ classical monocytes from patients with PD and PDDM showed elevated nuclear factor kappa B-inducing kinase activity.
CONCLUSIONS
Overall, this study elucidates how the presence of DM contributes to the deterioration of T/NK cell immunity and the immunological basis connecting PD to DM.
Topics: Humans; Diabetes Mellitus, Type 2; Leukocytes, Mononuclear; Periodontitis; Cytokines; Killer Cells, Natural
PubMed: 38082425
DOI: 10.1002/ctm2.1503 -
Communicable Diseases Intelligence... Nov 2023The Australian Partnership for Preparedness Research on InfectiouS disease Emergencies (APPRISE) has developed a virtual biobank to support infectious disease research...
The Australian Partnership for Preparedness Research on InfectiouS disease Emergencies (APPRISE) has developed a virtual biobank to support infectious disease research in Australia. The virtual biobank (https://apprise.biogrid.org.au) integrates access to existing distributed infectious disease biospecimen collections comprising multiple specimen types, including plasma, serum, and peripheral blood mononuclear cells. Through the development of a common data model, multiple collections can be searched simultaneously via a secure web portal. The portal enhances the visibility and searchability of existing collections within their current governance and custodianship arrangements. The portal is easily scalable for integration of additional collections.
Topics: Humans; Australia; Biological Specimen Banks; Leukocytes, Mononuclear; Specimen Handling; Communicable Diseases
PubMed: 37968063
DOI: 10.33321/cdi.2023.47.66 -
Frontiers in Immunology 2023Natural killer (NK) cells play an important role in immune rejection in solid organ transplantation. To mitigate human NK cell activation in xenotransplantation,...
Natural killer (NK) cells play an important role in immune rejection in solid organ transplantation. To mitigate human NK cell activation in xenotransplantation, introducing inhibitory ligands on xenografts genetic engineering of pigs may protect the graft from human NK cell-mediated cytotoxicity and ultimately improve xenograft survival. In this study, non-classical HLA class I molecules HLA-E and HLA-G were introduced in an immortalized porcine liver endothelial cell line with disruption of five genes (, , , , and ) encoding three major carbohydrate xenoantigens (αGal, Neu5Gc, and Sda) and swine leukocyte antigen class I (SLA-I) molecules. Expression of HLA-E and/or HLA-G on pig cells were confirmed by flow cytometry. Endogenous HLA-G molecules as well as exogenous HLA-G VL9 peptide could dramatically enhance HLA-E expression on transfected pig cells. We found that co-expression of HLA-E and HLA-G on porcine cells led to a significant reduction in human NK cell activation compared to the cells expressing HLA-E or HLA-G alone and the parental cell line. NK cell activation was assessed by analysis of CD107a expression in CD3CD56 population gated from human peripheral blood mononuclear cells. CD107a is a sensitive marker of NK cell activation and correlates with NK cell degranulation and cytotoxicity. HLA-E and/or HLA-G on pig cells did not show reactivity to human sera IgG and IgM antibodies. This study demonstrated that co-expression of HLA-E and HLA-G on genetically modified porcine endothelial cells provided a superior inhibition in human xenoreactive NK cells, which may guide further genetic engineering of pigs to prevent human NK cell mediated rejection.
Topics: Animals; Humans; Swine; HLA-G Antigens; Leukocytes, Mononuclear; Cytotoxicity, Immunologic; Endothelial Cells; Killer Cells, Natural; HLA-E Antigens
PubMed: 37529053
DOI: 10.3389/fimmu.2023.1217809 -
RMD Open Aug 2023To assess to what extent leflunomide (LEF) and hydroxychloroquine (HCQ) therapy in patients with primary Sjögren's syndrome (RepurpSS-I) targets type I IFN-associated...
Leflunomide/hydroxychloroquine combination therapy targets type I IFN-associated proteins in patients with Sjögren's syndrome that show potential to predict and monitor clinical response.
OBJECTIVES
To assess to what extent leflunomide (LEF) and hydroxychloroquine (HCQ) therapy in patients with primary Sjögren's syndrome (RepurpSS-I) targets type I IFN-associated responses and to study the potential of several interferon associated RNA-based and protein-based biomarkers to predict and monitor treatment.
METHODS
In 21 patients treated with LEF/HCQ and 8 patients treated with placebo, blood was drawn at baseline, 8, 16 and 24 weeks. IFN-signatures based on RNA expression of five IFN-associated genes were quantified in circulating mononuclear cells and in whole blood. MxA protein levels were measured in whole blood, and protein levels of CXCL10 and Galectin-9 were quantified in serum. Differences between responders and non-responders were assessed and receiver operating characteristic analysis was used to determine the capacity of baseline expression and early changes (after 8 weeks of treatment) in biomarkers to predict treatment response at the clinical endpoint.
RESULTS
IFN-signatures in peripheral blood mononuclear cell and whole blood decreased after 24 weeks of LEF/HCQ treatment, however, changes in IFN signatures only poorly correlated with changes in disease activity. In contrast to baseline IFN signatures, baseline protein concentrations of galectin-9 and decreases in circulating MxA and Galectin-9 were robustly associated with clinical response. Early changes in serum Galectin-9 best predicted clinical response at 24 weeks (area under the curve 0.90).
CONCLUSIONS
LEF/HCQ combination therapy targets type-I IFN-associated proteins that are associated with strongly decreased B cell hyperactivity and disease activity. IFN-associated Galectin-9 is a promising biomarker for treatment prediction and monitoring in pSS patients treated with LEF/HCQ.
Topics: Humans; Biomarkers; Hydroxychloroquine; Interferon Type I; Leflunomide; Leukocytes, Mononuclear; Proteins; RNA; Sjogren's Syndrome
PubMed: 37532471
DOI: 10.1136/rmdopen-2023-002979