Driver mutation subtypes involve with differentiated immunophenotypes influencing pancreatic cancer outcomes.

Despite many studies focusing on the prognostic biomarkers in pancreatic adenocarcinomas (PAADs), there is ill-informed about the relationships between their genomic features and immune characteristics. Herein, we deeply investigated the involvement of major driver mutation subtypes with immunophenotypes impacting PAAD outcomes. Based on public data analyses of RNA expression-based immune subtypes in PAAD, in contrast to KRAS G12D & TP53 co-mutant patients with poor outcomes, the best immune subtype C3 (inflammatory) characterized by high Th1/Th2 ratio was relatively enriched in KRASnon-G12DTP53wt patients with better survival, whereas the inferior subtype C2 (IFN-γ dominant) with low Th1/Th2 ratio was more common in the former than in the latter. Moreover, contrary to the highly immunosuppressive microenvironment (high Treg, high ratio of Treg to tumor-specific CD4+ T cell) in KRASG12DTP53mut patients, KRASG12VTP53wt individuals exhibited an inflamed context profiled by multiplex immunohistochemistry. It could be responsible for their outstanding survival advantage over others in postsurgical PAAD patients receiving adjuvant chemotherapy as shown by our cohort. Together, KRASG12VTP53wt may be a promising biomarker for prognostic evaluation and screening certain candidates with PAAD to get desirable survival benefit from adjuvant chemotherapy.

Immune profiling and functional analysis of NK and T cells in ataxia telangiectasia.

Ataxia telangiectasia (AT) is a rare autosomal-recessive disorder characterized by profound neurodegeneration, combined immunodeficiency, and an increased risk for malignant diseases. Treatment options for AT are limited, and the long-term survival prognosis for patients remains grim, primarily due to the emergence of chronic respiratory pathologies, malignancies, and neurological complications. Understanding the dysregulation of the immune system in AT is fundamental for the development of novel treatment strategies. In this context, we performed a retrospective longitudinal immunemonitoring of lymphocyte subset distribution in a cohort of AT patients (n = 65). Furthermore, we performed FACS analyses of peripheral blood mononuclear cells from a subgroup of 12 AT patients to examine NK and T cells for the expression of activating and functional markers. We observed reduced levels of peripheral blood CD3+CD8+ cytotoxic T cells, CD3+CD4+ T helper cells, and CD19+ B cells, whereas the amount of CD3--CD56+ NK cells and CD3+CD56+ NKT-like cells was similar compared with age-matched controls. Notably, there was no association between the age-dependent kinetic of T-, B-, or NK-cell counts and the occurrence of malignancy in AT patients. Additionally, our results indicate an altered NK- and T-cell response to cytokine stimulation in AT with increased levels of TRAIL, FasL, and CD16 expression in NK cells, as well as an elevated activation level of T cells in AT with notably higher expression levels of IFN-γ, CD107a, TRAIL, and FasL. Together, these findings imply function alterations in AT lymphocytes, specifically in T and NK cells, shedding light on potential pathways for innovative therapies.

Mendelian randomization based on immune cells in diabetic nephropathy.

Background: DKD, a leading cause of chronic kidney and end-stage renal disease, lacks robust immunological research. Recent GWAS utilizing SNPs and CNVs has shed light on immune mechanisms of kidney diseases. However, DKD's immunological basis remains elusive. Our goal is to unravel cause-effect relationships between immune cells and DKD using Mendelian randomization. Methodology: We analyzed FinnGen data (1032 DKD cases, 451,248 controls) with 731 immunocyte GWAS summaries (MP=32, MFI=389, AC=118, RC=192). We employed forward and reverse Mendelian randomization to explore causal links between immune cell traits and DKD. Sensitivity analysis ensured robustness, heterogeneity checks, and FDR correction minimized false positives. Results: Our study explored the causal link between diabetic nephropathy (DKD) and immunophenotypes using two-sample Mendelian Randomization (MR) with IVW. Nine immunophenotypes were significantly associated with DKD at p<0.05 after FDR correction. Elevated CD24, CD3 in Treg subsets, CD39+ CD4+, and CD33- HLA DR- AC correlated positively with DKD risk, while CD27 in B cells and SSC-A in CD4+ inversely correlated. Notably, while none showed significant protection, further research on immune cells' role in DKD may provide valuable insights. Conclusion: The results of this study show that the immune cells are closely related to DKD, which may be helpful in the future clinical study.

Isolating Immune Cells from Mouse Brain and Skull.

Mounting evidence indicates that the immune response triggered by brain disorders (e.g., brain ischemia and autoimmune encephalomyelitis) occurs not only in the brain, but also in the skull. A key step toward analyzing changes in immune cell populations in both the brain and skull bone marrow after brain damage (e.g., stroke) is to obtain sufficient numbers of high-quality immune cells for downstream analyses. Here, two optimized protocols are provided for isolating immune cells from the brain and skull bone marrow. The advantages of both protocols are reflected in their simplicity, speed, and efficacy in yielding a large quantity of viable immune cells. These cells may be suitable for a range of downstream applications, such as cell sorting, flow cytometry, and transcriptomic analysis. To demonstrate the effectiveness of the protocols, immunophenotyping experiments were performed on stroke brains and normal brain skull bone marrow using flow cytometry analysis, and the results aligned with findings from published studies.

Genetically predicted metabolites mediate the association between immune cells and metabolic dysfunction-associated steatotic liver disease: a mendelian randomization study.

BACKGROUND: Existing studies have presented limited and disparate findings on the nexus between immune cells, plasma metabolites, and metabolic dysfunction-associated steatotic liver disease (MASLD). The aim of this study was to investigate the causal relationship between immune cells and MASLD. Additionally, we aimed to identify and quantify the potential mediating role of metabolites. METHODS: A Mendelian randomization (MR) analysis was conducted using two samples of pooled data from genome-wide association studies on MASLD that included 2568 patients and 409,613 control individuals. Additionally, a mediated MR study was employed to quantify the metabolite-mediated immune cell effects on MASLD. RESULTS: In this study, eight immunophenotypes were linked to the risk of MASLD, and thirty-five metabolites/metabolite ratios were linked to the occurrence of MASLD. Furthermore, a total of six combinations of immunophenotypic and metabolic factors demonstrated effects on the occurrence of MASLD, although the mediating effects of metabolites were not significant. CONCLUSION: Our study demonstrated that certain immunophenotypes and metabolite/metabolite ratios have independent causal relationships with MASLD. Furthermore, we identified specific metabolites/metabolite ratios that are associated with an increased risk of MASLD. However, their mediating role in the causal association between immunophenotypes and MASLD was not significant. It is important to consider immune and metabolic disorders among patients with MASLD in clinical practice.

FlowAtlas: an interactive tool for high-dimensional immunophenotyping analysis bridging FlowJo with computational tools in Julia.

As the dimensionality, throughput and complexity of cytometry data increases, so does the demand for user-friendly, interactive analysis tools that leverage high-performance machine learning frameworks. Here we introduce FlowAtlas: an interactive web application that enables dimensionality reduction of cytometry data without down-sampling and that is compatible with datasets stained with non-identical panels. FlowAtlas bridges the user-friendly environment of FlowJo and computational tools in Julia developed by the scientific machine learning community, eliminating the need for coding and bioinformatics expertise. New population discovery and detection of rare populations in FlowAtlas is intuitive and rapid. We demonstrate the capabilities of FlowAtlas using a human multi-tissue, multi-donor immune cell dataset, highlighting key immunological findings. FlowAtlas is available at https://github.com/gszep/FlowAtlas.jl.git.

An exploration of the causal relationship between 731 immunophenotypes and osteoporosis: a bidirectional Mendelian randomized study.

Background: There are complex interactions between osteoporosis and the immune system, and it has become possible to explore their causal relationship based on Mendelian randomization methods. Methods: Utilizing openly accessible genetic data and employing Mendelian randomization analysis, we investigated the potential causal connection between 731 immune cell traits and the risk of developing osteoporosis. Results: Ten immune cell phenotypes were osteoporosis protective factors and three immune cell phenotypes were osteoporosis risk factors. Specifically, the odds ratio (OR) of IgD+ CD24+ %B cell (B cell panel) risk on Osteoporosis was estimated to be 0.9986 (95% CI = 0.9978~0.9996, P<0.01). The OR of CD24+ CD27+ %B cell (B cell panel) risk on Osteoporosis was estimated to be 0.9991 (95% CI = 0.9984~0.9998, P = 0.021). The OR of CD33- HLA DR+AC (Myeloid cell panel) risk on Osteoporosis was estimated to be 0.9996 (95% CI = 0.9993~0.9999, P = 0.038). The OR of EM CD8br %CD8br (Maturation stages of T cell panel) risk on Osteoporosis was estimated to be 1.0004 (95% CI = 1.0000~1.0008, P = 0.045). The OR of CD25 on IgD+ (B cell panel) risk on Osteoporosis was estimated to be 0.9995 (95% CI = 0.9991~0.9999, P = 0.024). The OR of CD25 on CD39+ activated Treg+ (Treg panel) risk on Osteoporosis was estimated to be 1.001 (95% CI = 1.0001~1.0019, P = 0.038). The OR of CCR2 on CD62L+ myeloid DC (cDC panel) risk on Osteoporosis was estimated to be 0.9992 (95% CI = 0.9984~0.9999, P = 0.048). The OR of CCR2 on CD62L+ plasmacytoid DC (cDC panel) risk on Osteoporosis was estimated to be 0.9993 (95% CI = 0.9987~0.9999, P = 0.035). The OR of CD45 on CD33dim HLA DR+ CD11b- (Myeloid cell panel) risk on Osteoporosis was estimated to be 0.9988 (95% CI = 0.9977~0.9998, P = 0.031). The OR of CD45 on Mo MDSC (Myeloid cell panel) risk on Osteoporosis was estimated to be 0.9992 (95% CI = 0.9985~0.9998, P = 0.017). The OR of SSC-A on B cell (TBNK panel) risk on Osteoporosis was estimated to be 0.9986 (95% CI = 0.9972~0.9999, P = 0.042). The OR of CD11c on CD62L+ myeloid DC (cDC panel) risk on Osteoporosis was estimated to be 0.9987 (95% CI = 0.9978~0.9996, P<0.01). The OR of HLA DR on DC (cDC panel) risk on Osteoporosis was estimated to be 1.0007 (95% CI = 1.0002~1.0011, P<0.01). No causal effect of osteoporosis on immune cells was observed. Conclusions: Our study identified 13 unreported immune phenotypes that are causally related to osteoporosis, providing a theoretical basis for the bone immunology doctrine.

Flow Cytometric Assessment of Malignant Hematologic Disorders.

Multiparameter flow cytometry (MPF) is an essential component of the diagnostic workup of hematologic malignancies. Recently developed tools have expanded the utility of MPF in detecting T-cell clonality and myelomonocytic dysplasia. Minimal/measurable residual disease analysis has long been established as critical in the management of B-lymphoblastic leukemia and is emerging as a useful tool in myeloid malignancies. With the continued increased complexity of MPF assays, emerging tools for data collection and analysis will allow users to take full advantage of MPF in the diagnosis of hematologic disease.

Flow Cytometry-based Immune Phenotyping of T and B Lymphocytes in the Evaluation of Immunodeficiency and Immune Dysregulation.

There are approximately 500 congenital disorders that impair immune cell development and/or function. Patients with these disorders may present with a wide range of symptoms, including increased susceptibility to infection, autoimmunity, autoinflammation, lymphoproliferation, and/or atopy. Flow cytometry-based immune phenotyping of T and B lymphocytes plays an essential role in the evaluation of patients with these presentations. In this review, we describe the clinical utility of flow cytometry as part of a comprehensive evaluation of immune function and how this testing may be used as a diagnostic tool to identify underlying aberrant immune pathways, monitor disease activity, and assess infection risk.

A Nonenzymatic Procedure to Obtain Human Mesenchymal Stromal Cells from the Dermis.

Human mesenchymal stromal cells (MSCs) have gained significant interest as cell-based therapeutics for organ restoration in the field of regenerative medicine. More recently, substantial attention has been directed toward cell-free therapy, achieved through the utilization of soluble factors possessing trophic and immunomodulatory properties present in the MSC secretome. This collection of soluble factors can be found either freely in the secretome or packed within its vesicular fraction, known as extracellular vesicles (EVs). MSCs can be derived from various tissue sources, each involving different extraction methods and yielding varying cell amounts. In this study, we describe a nonenzymatic procedure for a straightforward isolation of MSCs from the fetal dermis and the adult dermis. The results demonstrate the isolation of a cell population with a uniform MSC immunophenotype from the earliest passages (approximately 90% positive for the classical MSC markers CD90, CD105, and CD73, while negative for the hematopoietic markers CD34 and CD45, as well as HLA-DR). Additionally, we describe the procedures for cell expansion, banking, and secretome collection.

Culture and Immunomodulation of Equine Muscle-Derived Mesenchymal Stromal Cells: A Comparative Study of Innovative 2D versus 3D Models Using Equine Platelet Lysate.

Muscle-derived mesenchymal stromal cells (mdMSCs) hold great promise in regenerative medicine due to their immunomodulatory properties, multipotent differentiation capacity and ease of collection. However, traditional in vitro expansion methods use fetal bovine serum (FBS) and have numerous limitations including ethical concerns, batch-to-batch variability, immunogenicity, xenogenic contamination and regulatory compliance issues. This study investigates the use of 10% equine platelet lysate (ePL) obtained by plasmapheresis as a substitute for FBS in the culture of mdMSCs in innovative 2D and 3D models. Using muscle microbiopsies as the primary cell source in both models showed promising results. Initial investigations indicated that small variations in heparin concentration in 2D cultures strongly influenced medium coagulation with an optimal proliferation observed at final heparin concentrations of 1.44 IU/mL. The two novel models investigated showed that expansion of mdMSCs is achievable. At the end of expansion, the 3D model revealed a higher total number of cells harvested (64.60 ± 5.32 million) compared to the 2D culture (57.20 ± 7.66 million). Trilineage differentiation assays confirmed the multipotency (osteoblasts, chondroblasts and adipocytes) of the mdMSCs generated in both models with no significant difference observed. Immunophenotyping confirmed the expression of the mesenchymal stem cell (MSC) markers CD-90 and CD-44, with low expression of CD-45 and MHCII markers for mdMSCs derived from the two models. The generated mdMSCs also had great immunomodulatory properties. Specific immunological extraction followed by enzymatic detection (SIEFED) analysis demonstrated that mdMSCs from both models inhibited myeloperoxidase (MPO) activity in a strong dose-dependent manner. Moreover, they were also able to reduce reactive oxygen species (ROS) activity, with mdMSCs from the 3D model showing significantly higher dose-dependent inhibition compared to the 2D model. These results highlighted for the first time the feasibility and efficacy of using 10% ePL for mdMSC expansion in novel 2D and 3D approaches and also that mdMSCs have strong immunomodulatory properties that can be exploited to advance the field of regenerative medicine and cell therapy instead of using FBS with all its drawbacks.

Machine learning-based derivation and validation of three immune phenotypes for risk stratification and prognosis in community-acquired pneumonia: a retrospective cohort study.

Background: The clinical presentation of Community-acquired pneumonia (CAP) in hospitalized patients exhibits heterogeneity. Inflammation and immune responses play significant roles in CAP development. However, research on immunophenotypes in CAP patients is limited, with few machine learning (ML) models analyzing immune indicators. Methods: A retrospective cohort study was conducted at Xinhua Hospital, affiliated with Shanghai Jiaotong University. Patients meeting predefined criteria were included and unsupervised clustering was used to identify phenotypes. Patients with distinct phenotypes were also compared in different outcomes. By machine learning methods, we comprehensively assess the disease severity of CAP patients. Results: A total of 1156 CAP patients were included in this research. In the training cohort (n=809), we identified three immune phenotypes among patients: Phenotype A (42.0%), Phenotype B (40.2%), and Phenotype C (17.8%), with Phenotype C corresponding to more severe disease. Similar results can be observed in the validation cohort. The optimal prognostic model, SuperPC, achieved the highest average C-index of 0.859. For predicting CAP severity, the random forest model was highly accurate, with C-index of 0.998 and 0.794 in training and validation cohorts, respectively. Conclusion: CAP patients can be categorized into three distinct immune phenotypes, each with prognostic relevance. Machine learning exhibits potential in predicting mortality and disease severity in CAP patients by leveraging clinical immunological data. Further external validation studies are crucial to confirm applicability.

Immunophenotyping and Therapeutic Insights from Chronic Mucocutaneous Candidiasis Cases with STAT1 Gain-of-Function Mutations.

PURPOSE: Heterozygous STAT1 Gain-of-Function (GOF) mutations are the most common cause of chronic mucocutaneous candidiasis (CMC) among Inborn Errors of Immunity. Clinically, these mutations manifest as a broad spectrum of immune dysregulation, including autoimmune diseases, vascular disorders, and malignancies. The pathogenic mechanisms of immune dysregulation and its impact on immune cells are not yet fully understood. In treatment, JAK inhibitors have shown therapeutic effectiveness in some patients. METHODS: We analyzed clinical presentations, cellular phenotypes, and functional impacts in five Taiwanese patients with STAT1 GOF. RESULTS: We identified two novel GOF mutations in 5 patients from 2 Taiwanese families, presenting with symptoms of CMC, late-onset rosacea, and autoimmunity. The enhanced phosphorylation and delayed dephosphorylation were displayed by the patients' cells. There are alterations in both innate and adaptive immune cells, including expansion of CD38+HLADR +CD8+ T cells, a skewed activated Tfh cells toward Th1, reduction of memory, marginal zone and anergic B cells, all main functional dendritic cell lineages, and a reduction in classical monocyte. Baricitinib showed therapeutic effectiveness without side effects. CONCLUSION: Our study provides the first comprehensive clinical and molecular characteristics in STAT1 GOF patient in Taiwan and highlights the dysregulated T and B cells subsets which may hinge the autoimmunity in STAT1 GOF patients. It also demonstrated the therapeutic safety and efficacy of baricitinib in pediatric patient. Further research is needed to delineate how the aberrant STAT1 signaling lead to the changes in cellular populations as well as to better link to the clinical manifestations of the disease.

MRI features of breast cancer immunophenotypes with a focus on luminal estrogen receptor low positive invasive carcinomas.

To compare the magnetic resonance imaging (MRI) features of different immunophenotypes of breast carcinoma of no special type (NST), with special attention to estrogen receptor (ER)-low-positive breast cancer. This retrospective, single-centre, Institutional Review Board (IRB)-approved study included 398 patients with invasive breast carcinoma. Breast carcinomas were classified as ER-low-positive when there was ER staining in 1-10% of tumour cells. Pretreatment MRI was reviewed to assess the tumour imaging features according to the 5th edition of the Breast Imaging Reporting and Data System (BI-RADS) lexicon. Of the 398 cases, 50 (12.6%) were luminal A, 191 (48.0%) were luminal B, 26 (6.5%) were luminal ER-low positive, 64 (16.1%) were HER2-overexpressing, and 67 (16.8%) were triple negative. Correlation analysis between MRI features and tumour immunophenotype showed statistically significant differences in mass shape, margins, internal enhancement and the delayed phase of the kinetic curve. An oval or round shape and rim enhancement were most frequently observed in triple-negative and luminal ER-low-positive tumours. Spiculated margins were most common in luminal A and luminal B tumours. A persistent kinetic curve was more frequent in luminal A tumours, while a washout curve was more common in the triple-negative, HER2-overexpressing and luminal ER-low-positive immunophenotypes. Multinomial regression analysis showed that luminal ER-low-positive tumours had similar results to triple-negative tumours for almost all variables. Luminal ER-low-positive tumours present with similar MRI findings to triple-negative tumours, which suggests that MRI can play a fundamental role in adequate radiopathological correlation and therapeutic planning in these patients.

Molecular screening of transitional B cells as a prognostic marker of improved graft outcome and reduced rejection risk in kidney transplant.

Introduction: Understanding immune cell dynamics in kidney transplantation may provide insight into the mechanisms of rejection and improve patient management. B cells have gained interest with a special relevance of the "regulatory" subsets and their graft outcome prognostic value. In this study, we aimed to prove that the direct immunophenotyping and target gene expression analysis of kidney transplant patients' fresh whole blood will help to identify graft rejection risk and assist in the monitoring of kidney transplanted patients. Methods: We employed flow cytometry and qPCR techniques to characterize B and T cell subsets within fresh whole blood samples, with particular emphasis on transitional B cells (TrB) identified as CD19+CD24hiCD38hi. TrB are a relevant population in the context of kidney transplantation and are closely associated with regulatory B cells (Bregs) in humans. Patients were monitored, tracking pertinent clinical parameters and kidney-related events, including alterations in graft function and episodes of biopsy proven rejection. Results: Higher percentages of TrB cells at 3 months after transplantation were positively associated with better graft outcomes and lower biopsy-proven acute rejection risk. Furthermore, a novel panel of B cell regulatory associated genes was validated at 3 months post-transplantation by qPCR analysis of peripheral blood mononuclear cell (PBMC) mRNA, showing high predictive power of graft events and prognostic value. Discussion: These findings suggest that monitoring TrB may provide interesting patient management information, improve transplant outcomes, and allow for personalized drug regimens to minimize clinical complications.

Identification of single-cell blasts in pediatric acute myeloid leukemia using an autoencoder.

Pediatric acute myeloid leukemia (AML) is an aggressive blood cancer with a poor prognosis and high relapse rate. Current challenges in the identification of immunotherapy targets arise from patient-specific blast immunophenotypes and their change during disease progression. To overcome this, we present a new computational research tool to rapidly identify malignant cells. We generated single-cell flow cytometry profiles of 21 pediatric AML patients with matched samples at diagnosis, remission, and relapse. We coupled a classifier to an autoencoder for anomaly detection and classified malignant blasts with 90% accuracy. Moreover, our method assigns a developmental stage to blasts at the single-cell level, improving current classification approaches based on differentiation of the dominant phenotype. We observed major immunophenotype and developmental stage alterations between diagnosis and relapse. Patients with KMT2A rearrangement had more profound changes in their blast immunophenotypes at relapse compared to patients with other molecular features. Our method provides new insights into the immunophenotypic composition of AML blasts in an unbiased fashion and can help to define immunotherapy targets that might improve personalized AML treatment.

Clonal analysis of fetal hematopoietic stem/progenitor cells reveals how post-transplantation capabilities are distributed.

It has been proposed that adult hematopoiesis is sustained by multipotent progenitors (MPPs) specified during embryogenesis. Adult-like hematopoietic stem cell (HSC) and MPP immunophenotypes are present in the fetus, but knowledge of their functional capacity is incomplete. We found that fetal MPP populations were functionally similar to adult cells, albeit with some differences in lymphoid output. Clonal assessment revealed that lineage biases arose from differences in patterns of single-/bi-lineage differentiation. Long-term (LT)- and short-term (ST)-HSC populations were distinguished from MPPs according to capacity for clonal multilineage differentiation. We discovered that a large cohort of long-term repopulating units (LT-RUs) resides within the ST-HSC population; a significant portion of these were labeled using Flt3-cre. This finding has two implications: (1) use of the CD150+ LT-HSC immunophenotype alone will significantly underestimate the size and diversity of the LT-RU pool and (2) LT-RUs in the ST-HSC population have the attributes required to persist into adulthood.

DIAPH1-Deficiency is Associated with Major T, NK and ILC Defects in Humans.

Loss of function mutations in Diaphanous related formin 1 (DIAPH1) are associated with seizures, cortical blindness, and microcephaly syndrome (SCBMS) and are recently linked to combined immunodeficiency. However, the extent of defects in T and innate lymphoid cells (ILCs) remain unexplored. Herein, we characterized the primary T, natural killer (NK) and helper ILCs of six patients carrying two novel loss of function mutation in DIAPH1 and Jurkat cells after DIAPH1 knockdown. Mutations were identified by whole exome sequencing. T-cell immunophenotyping, proliferation, migration, cytokine signaling, survival, and NK cell cytotoxicity were studied via flow cytometry-based assays, confocal microscopy, and real-time qPCR. CD4+ T cell proteome was analyzed by mass spectrometry. p.R351* and p.R322*variants led to a significant reduction in the DIAPH1 mRNA and protein levels. DIAPH1-deficient T cells showed proliferation, activation, as well as TCR-mediated signaling defects. DIAPH1-deficient PBMCs also displayed impaired transwell migration, defective STAT5 phosphorylation in response to IL-2, IL-7 and IL-15. In vitro generation/expansion of Treg cells from naïve T cells was significantly reduced. shRNA-mediated silencing of DIAPH1 in Jurkat cells reduced DIAPH1 protein level and inhibited T cell proliferation and IL-2/STAT5 axis. Additionally, NK cells from patients had diminished cytotoxic activity, function and IL-2/STAT5 axis. Lastly, DIAPH1-deficient patients' peripheral blood contained dramatically reduced numbers of all helper ILC subsets. DIAPH1 deficiency results in major functional defects in T, NK cells and helper ILCs underlining the critical role of formin DIAPH1 in the biology of those cell subsets.

Causal role of immune cells in aplastic anemia: Mendelian randomization (MR) study.

Prior research has identified associations between immune cells and aplastic anaemia (AA); however, the causal relationships between them have not been conclusively established. A two-sample Mendelian randomisation analysis was conducted to investigate the causal link between 731 immune cell signatures and AA risk using publicly available genetic data. Four types of immune signatures, including relative cell, absolute cell (AC), median fluorescence intensities and morphological parameters, were considered sensitivity analyses were also performed to verify the robustness of the results and assess potential issues such as heterogeneity and horizontal pleiotropy. Following multiple test adjustments using the False Discovery Rate (FDR) method, no statistically significant impact of any immunophenotype on AA was observed. However, twelve immunophenotypes exhibited a significant correlation with AA without FDR correction (p of IVW < 0.01), of which eight were harmful to AA: CD127- CD8br %T cell (Treg panel), CD25 on IgD + CD38dim (B cell panel), CD38 on naive-mature B cell (B cell panel), CD39 + resting Treg % CD4 Treg (Treg panel), CD39 + secreting Treg AC (Treg panel), CD8 on CD28 + CD45RA- CD8br (Treg panel), HLA DR + NK AC (TBNK panel), Naive DN (CD4-CD8-) AC (Maturation stages of T cell panel); and four were protective to AA: CD86 on CD62L + myeloid DC (cDC panel), DC AC (cDC panel), DN (CD4-CD8-) NKT %T cell (TBNK panel), and TD CD4 + AC (Maturation stages of T cell panel). The results of this study demonstrate a close link between immune cells and AA by genetic means, thereby improving the current understanding of the interaction between immune cells and AA risk and providing guidance for future clinical research.

Ultrasound-Guided Fine Needle Biopsy of Human Axillary Lymph Nodes to Assess B Cell Responses to Vaccination.

Ultrasound-guided fine needle biopsy, also known as fine needle aspiration, of human axillary lymph nodes is a safe and effective procedure to assess the immune response within the lymph nodes following vaccination. Once acquired, lymph node cells can be characterized via flow cytometric immunophenotyping and/or single-cell RNA sequencing for gene expression and T and B cell receptors. Analysis of the immune cells from the lymph nodes enables the investigation of T and B cells that may interact at this site. These interactions may lead to germinal center formation and expansion, critical for the generation of effective immunity to vaccination. Directly studying the dynamic processes and interaction of the key cells has been challenging in humans due to the anatomically protected location of these cells. Here, we describe the methods involved in ultrasound-guided fine needle biopsy of human axillary lymph nodes in response to vaccination and subsequent analyses of the B cell populations.