Search Results (675)

Background: Type 1 diabetes (T1D) is an autoimmune disorder characterized by destruction of insulin-producing β-cells. While conventional insulin therapy manages hyperglycemia, it fails to halt autoimmunity. Oral immunotherapy targeting autoantigens like GAD65 offers potential for antigen-specific tolerance; however, its efficacy is limited by gastrointestinal degradation and poor mucosal uptake. Lactococcus lactis (L. lactis), a food-grade delivery vector, enables sustained antigen release and intestinal tract immune modulation, yet the differential transcriptomic mechanisms underlying mucosal versus systemic immune responses remain uncharacterized. Methods: Non-obese diabetic (NOD) mice were randomized into control and GAD65 groups, receiving oral PBS or the GAD65 recombinant L. lactis vaccine, respectively. Fasting blood glucose was monitored weekly. GAD65-specific IgA and IgG, along with immune tolerance-related factors, were quantified using ELISA. Lymphocyte subsets were analyzed by flow cytometry, alongside RNA sequencing and transcriptional profiling. Results: The study demonstrated that the orally administered GAD65-L. lactis vaccine could significantly induce GAD65-specific IgA antibody and TGF-β cytokine and alleviate hyperglycemia and diabetes symptoms in NOD mice. Our study facilitated the induction of GAD65-specific regulatory T cells within both intestinal lamina propria lymphocytes (LPLs) and splenic lymphocytes. Notably, antigen-specific tolerance was mainly observed in intestinal LPLs. Crucially, the immune responses elicited by the vaccine demonstrated significant disparities between intestinal LPLs and splenic lymphocytes, with intestinal LPLs exhibiting unique local immune tolerance transcriptomic profiles. Conclusions: Our findings have enhanced the comprehension of the mechanisms by which oral vaccines influence the interplay between mucosal and systemic immune responses, thereby establishing a foundational framework for the design of oral vaccines. This understanding is instrumental in advancing antigen-specific immune tolerance strategies for autoimmune diseases such as Type 1 Diabetes (T1D). Full article

Background: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of immature T cells, driven by dysregulated transcriptional networks and oncogenic signaling pathways. Here, we present the first comprehensive analysis of the expression and regulation of TSPAN32, a tetraspanin implicated in lymphocyte homeostasis, in T-ALL. Methods: Transcriptomic data from the Leukemia MILE study (GSE13159) were analyzed to assess TSPAN32 expression across leukemic subtypes. Gene Set Enrichment Analysis (GSEA) was performed to explore biological pathways associated with TSPAN32-correlated genes. For mechanistic validation, HPB-ALL cells were used as a model, with NOTCH signaling inhibited by γ-secretase inhibitor (GSI) treatment and TAL1–LMO1 overexpression induced through doxycycline-inducible lentiviral vectors. Gene expression changes were quantified by RT-qPCR. Results: TSPAN32 was frequently downregulated in T-ALL compared to healthy bone marrow, although expression was retained in a subset of cases. GSEA revealed that TSPAN32-correlated genes were inversely associated with cell cycle–related programs, consistent with its established role as a negative regulator of T cell proliferation. Mechanistically, TAL1–LMO1 overexpression strongly induced TSPAN32, while GSI-mediated NOTCH inhibition partially reactivated its expression. Interestingly, GSI treatment also increased TAL1 levels despite downregulating LMO1. Conversely, TAL1–LMO1 overexpression suppressed NOTCH1 and NOTCH3, highlighting a reciprocal regulatory interplay between NOTCH and TAL1/LMO1 oncogenic circuits that shapes TSPAN32 expression dynamics in T-ALL. Conclusions: This study identifies TSPAN32 as a novel transcriptional target under the influence of key leukemogenic pathways and suggests its potential role as a modulator of leukemic T cell proliferation, with implications for therapeutic strategies targeting TAL1 and NOTCH signaling. Full article

Hepatocellular carcinoma (HCC) is the most common liver cancer, with poor survival rates in advanced stages due to late diagnosis, tumor heterogeneity, and therapy resistance. The tumor microenvironment (TME) in HCC has a crucial role in tumor progression, characterized by a complex interaction of immune cells, stromal components, and immunosuppressive signaling pathways. Chronic inflammation driven by viral infections, metabolic dysfunction, and alcohol consumption triggers an immunosuppressive TME, promoting immune evasion and tumor growth. Immune cell populations, such as myeloid-derived suppressor cells, regulatory T cells, and tumor-associated macrophages, contribute to immunosuppression, while cytotoxic T lymphocytes and natural killer cells exert anti-tumor effects. Recent advances in immunotherapy, mainly immune checkpoint inhibitors (ICIs) targeting programmed death-ligand 1 and programmed cell death protein 1 and cytotoxic T-lymphocyte-associated protein 4, have revolutionized HCC treatment, though response rates remain limited. Combined therapies using tyrosine kinase inhibitors, anti-angiogenic agents, and ICIs improve patient outcomes. This review discusses the immunological mechanisms contributing to HCC progression, the role of immune cell subsets in tumor evasion, and therapeutic interventions, from conventional treatments to advanced immunotherapies. Ongoing clinical trials, barriers to effective treatment, and future directions to enhance HCC management and patient survival will also be overviewed. Full article

Lactate dehydrogenase (LDH) is a serum biomarker well known to correlate with disease severity in atopic dermatitis (AD). The aim of this study was to explore the cutaneous immune responses and the clinical profile of AD patients in relation to serum LDH levels. To this end, 47 untreated, adult patients with moderate-to-severe AD were stratified by median levels of serum LDH. Circulating memory T-cell responses to house dust mite (HDM) extract, in the presence of autologous lesional epidermal cells, were compared between AD subgroups. The LDH^high group exhibited significantly higher IL-13, IL-5 and IL-9 in vitro responses confined to the cutaneous lymphocyte-associated antigen (CLA)⁺ subset compared to LDH^low patients. Clinically, LDH^high patients were younger and exhibited more severe disease, elevated eosinophil counts in their blood, increased total and specific IgE levels in their plasma, and a higher prevalence of allergic rhinitis. Our data suggests that high LDH levels identify a subgroup of AD patients with a specific immune and clinical profile, and highlight the potential of LDH as a clinical parameter that may enable patient stratification for treatment selection. Full article

Although Torque Teno Viruses (TTVs) were initially considered to be ubiquitous members of the mammalian virome, the finding that swine TTVs (TTSuV) can act as primary pathogens elevates the possible status of swine TTVs (TTSuVs) to an emerging swine pathogen. Since their discovery, the molecular mechanisms of TTV–host interactions remain largely unknown as robust in vitro culture systems and in vivo animal models have not been available. This study was undertaken to address some of these long-standing gaps. Recombinant TTSuV1 rescued from an infectious clone was used to infect C57BL/J6 mice. Infected mice seroconverted within 15 days post-infection and mounted virus neutralizing antibody responses. Viral DNA was detected in blood and lung tissue for the duration of the study. TTSuV1 isolated from the lung tissue of infected mice productively and serially infected PK-15 cells in vitro, indicating that the treatment produced viable, replicative viral particles in the host. TTSuV1 antigen was also detected by flow cytometry in lymphocytes, including the T and B lymphocyte subsets. Infected mice exhibited mild splenic hyperplasia and lymphopenia. The ability to respond to mitogenic stimuli was highly diminished in infected mice and a striking lack of virus-specific recall responses was observed for the 30-day duration of the study. Therefore, this study is the first to provide experimental evidence that recombinant TTSuV1 rescued from an infectious clone is infective and induces immune responses in laboratory mice. This model provides a critical tool for advancing research on TTV immunopathogenesis. Full article

Background/Objectives: Radiolabeled interleukin-2 (IL2) could allow for imaging activated T-lymphocytes in the tumor microenvironment (TME). The aims of this study were to assess the shelf life of a lyophilized kit containing THP-desIL2 to allow for the labeling of IL2 with ⁶⁸Ga at room temperature and to test the in vitro binding of ⁶⁸Ga-THP-desIL2 on different T-cell populations in order to determine which specific T-cell subset expresses the CD25 subunit of the IL2 receptor (IL2R). Methods: desIL2 was conjugated with THP and lyophilized. ⁶⁸Ga labeling was performed and several quality controls, including HPLC, iTLC and SDS-PAGE, were carried out at different storage times (1, 3 and 6 months) and temperatures (4 °C and −80 °C). Moreover, flow cytometric analysis on different T-cell populations and the in vitro and competitive binding of ⁶⁸Ga-THP-desIL2 were performed. Results: The lyophilized kit of THP-desIL2 was stable up to 6 months at −80 °C, preserving its sterility, integrity and acceptable values of labeling yield (51.80 ± 3.74%), radiochemical purity (>96%) and specific activity (5.59 ± 0.40 MBq/µg). Binding of ⁶⁸Ga-THP-desIL2 on activated lymphocytes was specific and exhibited a low dissociation constant from IL2R on stimulated Tregs (Kd: 10⁻⁹–10⁻¹⁰ mol/L). Conclusions: We assessed the shelf life of a lyophilized kit containing THP-desIL2 for the easy labeling of IL2 with ⁶⁸Ga at room temperature. The kit can be stored at −80 °C up to 6 months, thus facilitating the adoption of ⁶⁸Ga-THP-desIL2 into clinical practice. ⁶⁸Ga-THP-desIL2 showed high affinity and specificity for CD25 on activated T-lymphocytes, particularly Tregs, thus opening new opportunities for imaging immune cells trafficking in the TME. Full article

Human cutaneous leishmaniasis (CL) caused by Leishmania (Viannia) braziliensis is a complex parasitic disease marked by dynamic host–parasite interactions and immunomodulation. Extracellular vesicles (EV) derived from immune cells have emerged as key mediators of intercellular communication and potential biomarkers in infectious diseases. In this study, we combined a modified lymphocyte proliferation assay with nano-flow cytometry to quantify and phenotype EV released by CD4⁺, CD8⁺, and CD14⁺ cells in PBMC cultures from CL patients at different clinical stages: before treatment (PBT), during treatment (PDT), and post-treatment (PET) with antimonial. Healthy individuals (HI) were included as physiological controls. Upon stimulation with L. (V.) braziliensis antigens, we observed a distinct modulation of EV subsets. In the PBT group, CD4⁺ and CD14⁺ EV were significantly reduced, while CD8⁺ EV remained elevated. During PDT and PET, EV concentrations were restored across all subsets. These findings suggest that L. (V.) braziliensis selectively modulates the release of immune cell–derived EV, possibly as an immune evasion mechanism. The restoration of EV release following antimonial therapy highlights their potential as sensitive biomarkers for disease activity and treatment monitoring. This study offers novel insights into the immunoregulatory roles of EV in CL and underscores their relevance in host–parasite interactions. Full article

Systemic lupus erythematosus (SLE) is a severe autoimmune disease characterized by autoantibody production and multi-organ involvement. Anifrolumab, a monoclonal antibody targeting the type I interferon (IFN) receptor, has been approved for the treatment of SLE. Our aim was to investigate the long-term effects of inhibited type I IFN signaling on circulating follicular helper T subsets (T_FH), follicular regulatory T cells (T_FR), and B lymphocyte subpopulations, reflecting the ongoing germinal center reactions in SLE patients. Peripheral blood samples were obtained from ten SLE patients before the initiation of anifrolumab treatment, and at months 6 and 12 of the intervention period. Flow cytometry analysis was performed to assess the frequencies of circulating T_FH cell subsets, T_FR cells, and certain B cell subpopulations. Serological parameters, including autoantibody levels and complement components, were determined as part of the routine diagnostic evaluation. We observed a significant and sustained reduction in the percentage of activated circulating T_FH cells. Notably, the frequency of CXCR3⁻CCR6⁺ T_FH17 cells decreased, whereas the proportion of CXCR3⁺CCR6⁻ T_FH1 cells increased significantly. Furthermore, the proportion of the IgD⁻CD27⁻ double-negative B lymphocytes was also significantly reduced. These findings suggest that anifrolumab therapy attenuates T_FH cell activation, which may contribute to its clinical efficacy by modulating germinal center responses in SLE. Full article

Background/Objectives: Restored CD4 absolute counts (CD4AC) and CD4/CD8 ratio in the setting of continuous antiretroviral treatment (ART) do not exclude a low-level immune activation associated with HIV reservoirs, microbial translocation, or the side effects of ART itself, which accelerates the aging of people living with HIV (PLHIV). To delineate biomarkers of incomplete immune restoration in PLHIV on successful ART, we evaluated T-lymphocyte mitochondrial parameters in relation to phenotypic markers of immune exhaustion and senescence. Methods: PLHIV with sustained viral suppression, CD4AC > 500 and CD4/CD8 ratio >0.9 on ART (n = 39) were compared to age-matched ART-naïve donors (n = 27) and HIV(–) healthy controls (HC, n = 35). CD4 and CD8 differentiation and effector subsets (CCR7/CD45RA and CD27/CD28), activation, exhaustion, and senescence markers (CD38, CD39 Treg, CD57, TIGIT, and PD-1) were determined by flow cytometry. Mitochondrial mass (MM) and membrane potential (MMP) of CD8 and CD4 T cells were evaluated with MitoTracker Green and Red flow cytometry dyes. Results: ART+PLHIV differed from HC by increased CD4 TEMRA (5.3 (2.1–8.8) vs. 3.2 (1.6–4.4), p < 0.05), persistent TIGIT+CD57–CD27+CD28– CD8+ subset (53.9 (45.5–68.9) vs. 40.1 (26.7–58.5), p < 0.05), and expanding preapoptotic TIGIT–CD57+CD8+ effectors (9.2 (4.3–21.8) vs. 3.0 (1.5–7.3), p < 0.01) in correlation with increased CD8+ MMP (2527 (1675–4080) vs.1477 (1280–1691), p < 0.01). These aberrations were independent of age, time to ART, or ART duration, and were combined with increasing CD4 T cell MMP and MM. Conclusions: In spite of recovered CD4AC and CD4/CD8 ratio, the increased CD8+ MMP, combined with elevated markers of exhaustion and senescence in ART+PLHIV, signals a malfunction of the CD8 effector pool that may compromise viral reservoir latency. Full article

Objective: The interactions between the central nervous system (CNS) and the immune system suggest that immune mechanisms may be effective in the pathogenesis of epilepsy and epileptic seizures. Although studies on the natural immune response and epilepsy are continuing, it is not yet clear whether the interaction of the current immune system is due to epilepsy itself or antiepileptic drugs (AEDs), since epileptic patients also use AEDs There are a limited number of studies that have reported an increased incidence of upper respiratory tract infections (URTIs) in patients during levetiracetam (LEV) treatment. Therefore, we aimed to report our experience regarding the effect of LEV monotherapy on the complete blood count (CBC), immunoglobulin (Ig) levels, and lymphocyte subgroups in the interictal period in children and adolescents with epilepsy. Methods: This study enrolled 31 children who presented with epilepsy and underwent LEV monotherapy for at least one year (patient group) and 43 healthy children (control group). The CBC parameters (hemoglobin (hb), lymphocytes, leukocytes, neutrophils, and platelets), Ig levels (IgA, IgM, IgG, and IgE), and lymphocyte subsets (CD3, CD4, CD8, CD4/CD8 ratio, CD19, CD56, NKT cells, and Treg cells) were measured and compared between the two groups. The patients were also investigated regarding the frequency and types of infections that they experienced in the first month and first year of the study, and these data were compared between the patient group and the control group. In addition, the same parameters and the frequency of infection were compared among the patient subgroups (focal and generalized seizures). Results: The results of the present study indicate that there were no significant differences in the CBC parameters, lymphocyte subsets, or Ig levels between the patient group and the control group. The comparison among the patient subgroups was similar; however, the CD4/CD8 ratio was lower in the patient subgroup with focal seizures. In addition, there were no significant differences in the frequency or type of infections experienced one month and one year of the study between the patient group and the control group, and likewise for the patient subgroups (focal and generalized seizures). Conclusions: The present study demonstrated that LEV monotherapy did not increase the incidence of infection, and there were no significant effects on the CBC or on the humoral or cellular immune system in epileptic children. These findings also suggest that the CD4/CD8 ratio among lymphocyte subgroups is lower in patients with focal seizures. However, the epilepsy subgroups had a relatively small sample size; therefore, further prospective studies involving a larger patient population are needed to establish the association between LEV monotherapy and lymphocyte subgroups in patients with epilepsy. Full article

Background: Regulatory T cells (Tregs) are the main suppressor cells that maintain immune tolerance and prevent autoimmunity. Changes in Treg number or function are implicated in a wide range of autoimmune and inflammatory (AI/I) diseases, with or without underlying inborn errors of immunity (IEI). Understanding the phenotypic profiles of Treg subsets and their associations with immune dysregulation is crucial to identifying potential robust and holistic biomarkers for disease activity. Methods: We examined peripheral blood mononuclear cells from 40 patients diagnosed with various autoimmune/inflammatory diseases, including those with genetically confirmed inborn errors of immunity (IEIs), and compared these samples to those from 38 healthy controls of the same age. Utilizing multiparametric flow cytometry, we measured multiple Treg sub-populations and investigated their correlations with lymphocyte subset profiles and the diversity of autoantibodies. We applied advanced statistical and machine learning techniques, such as t-SNE, k-means clustering, and ROC analysis, to analyze immunophenotypic patterns in the patients. Results: Among all Treg sub-populations, only CD4⁺CD127^lowCD25^highFOXP3⁺ Tregs showed a significant decrease in patients compared to healthy controls (p < 0.05), while other Treg phenotypes did not differ. FOXP3 expression showed reduced intensity in patients and demonstrated diagnostic potential (AUC = 0.754). Notably, this Treg subset negatively correlated with CD19⁺ B cell percentages and positively correlated with the diversity of circulating autoantibodies. Unsupervised clustering revealed three distinct immunophenotypic profiles, highlighting heterogeneity among patients and underlining FOXP3-centered immune dysregulation. Conclusions: Our results presented that patients have an impairment in the CD4⁺CD127^lowCD25^highFOXP3⁺ regulatory T cell subset, which is identified by significantly decreased frequency and decreased expression of FOXP3. Immunological heterogeneity among patients was further uncovered by unsupervised clustering, highlighting the critical role that FOXP3-centered regulatory failure plays in the pathophysiology of illness. The combined evaluation of these three immunological factors, centered around FOXP3, holds promise as an integrative tool for monitoring disease progression across various autoimmune and immunodeficient contexts. Full article

CD8+ T lymphocytes (CTLs) act as serial killers of infected or malignant cells by releasing large amounts of interferon-gamma (IFNγ) and granzymes. Although IFNγ is a pleiotropic cytokine with diverse immunomodulatory functions, its precise spatiotemporal regulation and role in CTL-mediated cytotoxicity remain incompletely understood. Using wild-type and granzyme B-mTFP knock-in mice, we employed a combination of in vitro approaches, including T cell isolation and culture, plate-bound anti-CD3e stimulation, degranulation assays, flow cytometry, immunofluorescence, and structured illumination microscopy, to investigate IFNγ dynamics in CTLs. IFNγ expression in CTLs was rapid, transient, and strictly dependent on T cell receptor (TCR) activation. We identified two functionally distinct IFNγ-producing subsets: IFNγ^high (IFNγ^hi) and IFNγ^low (IFNγ^lo) CTLs. IFNγ^hi CTLs exhibited an effector/effector memory phenotype, significantly elevated CD107a surface expression (a marker of lytic granule exocytosis), and higher colocalization with cis-Golgi and granzyme B compared to IFNγ^lo CTLs. Furthermore, CRTAM, an early activation marker, correlated with IFNγ expression in naive CTLs. Our findings establish a link between elevated IFNγ production and enhanced CTL cytotoxicity, implicating CRTAM as a potential regulator of early CTL activation and IFNγ induction. These insights provide a foundation for optimizing T cell-based immunotherapies against infections and cancers. Full article

Background/Objectives: We examined the in vivo effects of successive treatments with the histone deacetylase (HDAC) inhibitor romidepsin in patients with cutaneous T-cell lymphoma (CTCL), using changes in gene expression in peripheral blood mononuclear cells (PBMCs). Methods: Exploiting data from a highly responsive CTCL patient through 12 months of treatment, we identified a malignant cell predictor (MCP), a gene signature associated with the diminishing numbers of circulating malignant cells. Results: The MCP was successfully validated in the patient’s relapse sample 9 months after treatment was terminated and via an independent set of CTCL patient samples. Conclusions: The MCP set of genes contained novel CTCL markers, including membrane-associated proteins not normally expressed in lymphocytes. A subclass of those markers was also detectable in residual malignant cells undetected by flow cytometry in remission samples from a patient who relapsed 10 months later. We identified a subset of transcriptional regulators, miRNAs and methylation patterns associated with the effect of progressive treatments revealing potential mechanisms of transcriptional dysregulation and functional effects in the malignant cells. We demonstrate a role for transcriptional activator HLF, over-expressed in malignant cells, and downregulated transcriptional-suppressor and immune-modulator NFIL3, as regulators of CTCL-specific genes. Full article

HIV-associated lymphoma (HAL) is a heterogeneous and highly aggressive group of malignancies. Although antiretroviral therapy (ART) has significantly prolonged the survival of people living with HIV (PLWH), the risk of malignancy secondary to HIV infection remains higher than in HIV-negative individuals, with HAL being among the most frequent. The pathogenesis of HAL is complex, involving multifactorial interactions. In current clinical practice, HAL faces a double challenge: the lack of effective biological risk warning systems and the lack of precise prognostic stratification tools. In recent years, the construction of multidimensional biomarker systems has shown critical value in the comprehensive management of HAL. This review aims to systematically summarize recent advances in circulating biomarkers for HAL, focusing on the potential applications of immune environment indicators, such as inflammatory cytokine profiles and microbial translocation markers, as well as serum protein profiles, lymphocyte subsets, extracellular vesicles (EVs), circulating microRNAs (miRNAs), and viral biomarkers. These biomarkers offer promising avenues for early risk prediction, therapeutic monitoring, and prognostic evaluation. Developing an assessment system based on multidimensional biomarkers will optimize early risk stratification, enable precise prognostic classification, and support personalized therapeutic strategies, thereby providing a novel theoretical basis and practical direction for the clinical management of HAL. Full article

The traditional paradigm of multiple sclerosis (MS) as a T cell-mediated disorder has been challenged by the effectiveness of monoclonal antibodies (mAbs) targeting CD20-expressing lymphocytes. Although these are mostly represented by B cells, the CD20 marker is expressed by 2–6% of T cells (CD20+ T), which are effectively depleted in serum and cerebrospinal fluid of MS patients by anti-CD20 mAbs. CD20+ T cells are characterized by a pro-inflammatory phenotype and increased potential for migrating and invading the central nervous system (CNS) compared to CD20− T cells. Furthermore, CD20+ T cells are detected within brain inflammatory lesions from MS patients and actively participate in the experimental MS model. This review aims to summarize the current knowledge on CD20+ T cells, from their identification and characterization to evidence of depletion by disease-modifying treatments (DMTs), likely contributing to therapeutic efficacy. Conflicting hypotheses on the origin and development of CD20+ T cells will also be discussed, as well as evidence from clinical and preclinical studies supporting their pathogenetic role in MS. Full article