Background Regulatory B cells (Bregs) suppress allergic inflammation primarily through interleukin-10 (IL-10), yet the intrinsic mechanisms constraining their regulatory capacity remain incompletely understood. Although the anti-apoptotic protein Mcl-1 is essential for B cell survival, its potential non-canonical roles in Breg-mediated immune regulation are unknown. Methods B cell-specific Mcl1ΔCD19mice and OVA-induced airway allergy models were used to assess Breg function. Translational studies employed lentiviral MCL1overexpression in primary human B cells, PBMC-reconstituted NSG mice, and clinical B cell profiling from asthma patients. Rigorous experimental controls were implemented to exclude confounding effects from B cell developmental defects and compensatory anti-apoptotic signaling. Results Compared with littermate controls, Mcl1ΔCD19mice exhibited significantly attenuated airway hyperresponsiveness, pulmonary eosinophilia, and antigen-specific IgE levels (all p < 0.01), accompanied by a 4.2-fold increase in the absolute number of IL-10+Bregs (p < 0.001). B cells isolated from Mcl1ΔCD19mice potently suppressed Th2 cytokine production and promoted type 1 regulatory T (Tr1) cell differentiation in an IL-10R-dependent manner. Mechanistically, Mcl-1 associates with the Il10promoter; this chromatin binding coincides with enrichment of the repressive histone mark H3K27me3 and reduced recruitment of RNA polymerase II. In humanized mice, enforced MCL1 overexpression in human B cells reduced IL-10 secretion by 65% and exacerbated airway inflammation. Furthermore, B cells from asthma patients displayed elevated endogenous MCL1 expression and correspondingly diminished IL-10 production. It should be noted that our human-subject analysis is limited by modest sample size, and further validation in larger patient cohorts is required. Conclusions Mcl-1 functions as an epigenetic suppressor of Il10transcription in B cells, independently of its canonical pro-survival role, thereby restraining Breg-mediated immune tolerance and promoting allergic airway inflammation. The B-cell Mcl-1/IL-10 axisrepresents a promising preclinical pathway for further investigation as a potential therapeutic target for allergic asthma, though pharmacological validation is still needed.
Background Persistent circulating SARS-CoV-2 spike protein has been proposed as a potential contributor to long-COVID pathophysiology. Whether administration of spike-targeting monoclonal antibodies is followed by changes in spike protein concentrations, autonomic receptor autoantibodies, or clinical outcomes remains uncertain. Methods In this prospective, non-randomized observational study, 21 participants were enrolled. Ten immunodeficient patients with long-COVID-associated ME/CFS and detectable circulating spike protein received Tixagevimab/Cilgavimab for SARS-CoV-2 pre-exposure prophylaxis. One patient subsequently withdrew consent, leaving nine patients for paired baseline and 30-day follow-up analyses. Eleven healthy individuals with previous SARS-CoV-2 infection but without long COVID provided descriptive reference data and were not considered an efficacy control group. Results β1-adrenergic receptor autoantibody concentrations decreased from 26.8 ± 18.2 to 21.2 ± 14.7 U/ml (unadjusted p = 0.003). No statistically significant changes were observed for β2-adrenergic or M3/M4-muscarinic receptor autoantibodies. Median spike protein concentrations changed from 63 pg/ml (IQR 9–127) to 26 pg/ml (IQR 10–120; p = 0.63), with individual decreases observed in five of nine patients. None of the assessed clinical outcomes changed significantly at group level. Changes in spike protein concentrations were not significantly correlated with changes in the assessed autonomic receptor autoantibodies. Conclusion Tixagevimab/Cilgavimab administration was followed by a nominally significant within-patient reduction in β1-adrenergic receptor autoantibody concentrations, but not by conclusive changes in circulating spike protein or clinical outcomes. These findings are preliminary and hypothesis-generating. The small sample size, non-randomized design, brief 30-day follow-up, inclusion of only immunodeficient patients, and absence of an untreated disease-matched control group preclude conclusions regarding therapeutic efficacy.
First-generation vaccine candidates against leishmaniasis, based on chemically or physically inactivated whole parasite, have attracted considerable attention owing to their feasibility, scalability of production, and comparatively low manufacturing costs. Vaccine formulations typically require immunostimulatory compounds to elicit robust cell-mediated immunity. Traditionally, adjuvants have been incorporated into vaccine formulation to augment their immunogenic potential. Recent advances over the past decade have yielded novel adjuvant system capable of stimulating both antibody-mediated and cell-mediated immune pathways. Recent advances in adjuvant development have facilitated the design of several promising vaccine candidates against leishmaniasis. Therefore, in the present study, we tested tomatine (identified as a steroidal glycoalkaloid, a saponin subgroup), for its ability to induce cell-mediated immunity that confer protection against leishmaniasis. Tomatine, a glycoalkaloid based adjuvant, possesses the ability to elicit cytotoxic T-lymphocyte responses. It shows remarkable potential as an effective adjuvant candidate for vaccine applications targeting infectious pathogens. Our results demonstrated that tomatine combined with the first-generation antigen induced durable central (CD44highCD62Lhigh) and effector (CD44highCD62Llow) memory CD4⁺ and CD8⁺ T-cell responses in both pre-challenge and post-challenge mice, indicating the development of long-lasting protective cellular immunity. Significant protective immunity was further demonstrated in the vaccinated animals, as evidenced by remarkably declined parasite burden, increased levels of T cells (CD4+, CD8+), IFN-γ, GM-CSF, effector molecules i.e. reactive oxygen species and nitric oxide and heightened NF-kB and iNOS gene expression. Collectively, these findings highlight tomatine as an effective immunomodulatory adjuvant capable of inducing cell-mediated protective immunity against leishmaniasis.
Background The classical adenosinergic pathway (CD39/CD73/A2BR) plays a pivotal role in regulating tumor immune evasion in hepatocellular carcinoma (HCC). However, the prognostic significance of the specific spatial coordination of these enzymes within the tumor microenvironment remains elusive. Methods We retrospectively included 92 patients with HCC who underwent curative hepatectomy. Multiplex immunohistochemistry was used to assess the in situ expression of CD39, CD73, A2BR, and cytokeratin (CK). Digital pathology was employed to segment the tissue into tumor parenchyma (CK+) and stromal regions (CK−). Associations between spatial phenotypes and survival outcomes were evaluated using Kaplan–Meier and Cox proportional hazards models. Furthermore, the transcriptomic profiles of ENTPD1 (CD39), NT5E (CD73), and ADORA2B (A2BR) were analyzed in the TCGA-LIHC cohort (n = 371) to externally validate the protein-level findings. Results Although the densities of CD39, CD73, and A2BR were higher in the tumor parenchyma than those in the stroma, multivariate Cox regression revealed that only stromal CD39⁺CD73⁺A2BR⁺ co-expression phenotype emerged as an independent protective factor for overall survival (hazard ratio [HR] = 0.300, P = 0.035). Conversely, the stromal CD39⁻CD73⁺A2BR⁻ phenotype was identified as an independent risk factor (HR = 2.93, P = 0.023). Correlation analysis further demonstrated a strong positive correlation between CD73 and A2BR within the stroma (r = 0.803, P < 0.001). External validation in TCGA-LIHC corroborated the mIHC findings. Conclusion The spatial organization of the adenosine pathway, particularly within the stroma, is associated with HCC prognosis and may provide greater prognostic information than individual marker expression. These findings provide a potential basis for spatially informed risk stratification and may inform future studies of metabolic immunotherapy in HCC.
Objectives Tween-80 can induce severe pseudo-allergic reactions, but its mechanism is unclear. Mas-related G protein coupled receptor X2 (MRGPRX2) is a novel receptor that mediated pseudo-allergic reactions. The study aimed to determine whether MRGPRX2 signaling is involved in Tween-80-induced pseudo-allergic reactions. Methods With the treatment of Tween-80, degranulation of LAD2 mast cells transfected with specific MRGPRX2 siRNA was assayed by spectrophotometry, and paw swelling and extravasation were determined in wild-type and MrgprB2 knockout mice. Ca2+ mobilization and protein expression of signaling molecules were assayed by western blotting and laser confocal microscopy. Results Tween-80 caused pseudo-allergic reactions in WT mice, but not in Kitw-sh/w-sh mice, indicating mast cell dependency. In passive cutaneous pseudo-allergic test, Tween-80 vascular extravasation, paw swelling, mast cell degranulation, plasma histamine, and body temperature loss in wild‑type mice, whereas all these effects were inhibited in MrgprB2‑knockout mice, demonstrating that MrgprB2 (the mouse ortholog of MRGPRX2) is involved in the reaction. Consistently, in LAD2 cells, silencing MRGPRX2 with siRNA suppressed Tween‑80‑induced degranulation and Ca²⁺ influx. Tween‑80 also increased intracellular Ca²⁺ in MRGPRX2‑expressing HEK293 cells but not in HEK293 cells. Furthermore, Tween‑80 triggered Ca²⁺ mobilization and phosphorylation of Lyn, PLCγ, IP₃R, PKC, and p38 in LAD2 cells, and these responses were diminished by MRGPRX2 knockdown. Conclusion MRGPRX2 signaling is involved in Tween-80-induced pseudo-allergic reactions, likely through the Lyn/PLCγ/PKC/IP₃R and p38 pathways, whose activation appears to depend on MRGPRX2 and may contribute to mast cell degranulation. MRGPRX2 may be a potential therapeutic target for Tween-80-induced pseudo-allergic reactions.
