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关键词： Amyotrophic lateral sclerosis   MiR-223   Interleukin-1 beta   Inflammasome  

摘要： Background Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease identified by progressive motor neuron loss. NLRP3 inflammasomes induce inflammation and pyroptosis, which can lead to neurodegeneration, muscle atrophy, and respiratory decline. miR-223 targets NLRP3 and suppresses inflammasome formation. Here, miR-223, NLRP3 and IL-1 beta levels were evaluated as plasma biomarkers in the incidence and progression of ALS. Methods 32 ALS patients and 32 healthy subjects were assessed. In all patients, the functional disability was determined by Revised Amyotrophic Lateral Sclerosis Functional Rating Scale (ALSFRS-R), and the respiratory dysfunction was assessed by the percent predicted forced vital capacity (ppFVC) index in spirometry examination. Plasma levels of miR-223, NLRP3 and IL-1 beta were assessed in ALS and control groups. Results Compared to the healthy controls, ALS patients showed decreased miR-223 expression (P < 0.0001), increased NLRP3 expression (P = 0.0002) and increased IL-1 beta level (P = 0.0003). The areas under the ROC curves for miR-223, NLRP3 and IL-1 beta were 0.82, 0.76 and 0.75 respectively. The ALSFRS-R and ppFVC values were positively correlated with miR-223 and negatively correlated with NLRP3 and IL-1 beta levels. Conclusion Our results indicated that changes in miR-223, NLRP3 and IL-1 beta levels may correlate with the occurrence and functional progression of ALS. Additionally, therapeutic approaches based on miR-223 and inflammatory mediators can be proposed as effective strategies against disease progression.

关键词： MHC Class I   ProteinMPNN   direct preference optimization   protein deimmunization   protein design  

摘要： ProteinMPNN is widely used in protein design workflows due to its ability to identify amino acid sequences that fold into specific 3D protein structures. In our work, we adjust ProteinMPNN to design proteins for a given 3D protein structure with reduced immune-visibility to cytotoxic T lymphocytes that recognize proteins via the MHC-I pathway. To achieve this, we developed a novel framework that integrates Direct Preference Optimization (DPO)-a tuning method originally designed for large language models-with MHC-I peptide presentation predictions. This approach fosters the generation of designs with fewer MHC-I epitopes while preserving the protein's original structure. Our results demonstrate that DPO effectively reduces MHC-I visibility without compromising the structural integrity of the proteins. Source code: https://***/hcgasser/CAPE_MPNN.

关键词： immunoproteasome   IL-4RA   type 2 inflammation   precision-cut lung slices   airway hyperresponsiveness  

摘要： Introduction Immunoproteasome (IP) is induced by pro-inflammatory stimuli such as interferon gamma to regulate inflammation and immunity. Asthma patients with airway type 2 high inflammation (e.g., IL-13) demonstrate more eosinophils and airway hyperresponsiveness (AHR) with less interferon gamma. The role of IP in regulating airway eosinophilic inflammation and AHR has not been *** This study was aimed to determine how IP regulates type 2 inflammation and AHR using LMP7 (a subunit of IP) deficient mouse lungs, precision-cut lung slices (PCLS), and cultured human airway epithelial cells treated with IL-13 in the absence or presence of an IP inhibitor ONX-0914 or exogenous *** LMP7 KO mouse lungs had significantly more IL-4R alpha protein expression than the wildtype (WT) mice. Following IL-13 treatment in PCLS, LMP7 KO mice had significantly more airway contraction than WT mice, which was coupled with increased eotaxin-2 levels. IP inhibition by ONX-0914 in IL-13 treated human airway epithelial cells resulted in significantly more IL-4R alpha protein expression and eotaxin-3 release. IP inhibition in human PCLS significantly increased *** Collectively, these data demonstrated that IP promotes degradation of IL-4R alpha, while inhibits type 2 inflammation and AHR. Enhancement of IP expression or activity may serve as an alternative approach to reduce the severity of type 2 inflammation and AHR.

关键词： SARS-CoV-2   Omicron   cellular immune response   CD8 T-cell epitope   conserved epitope  

摘要： Introduction Recent surveillance has identified the emergence of the SARS-CoV-2 Omicron ariant, which exhibits the ability to evade multiple neutralizing antibodies generated by prior infection or vaccination. However, significant knowledge gaps remain regarding the CD8 T-cell immune reactivity to the Omicron variant. This study aims to evaluate the characteristics of HLA-A2-restricted CD8 T-cell epitopes from the Omicron variant and analyze epitope-specific CD8 T-cell responses to SARS-CoV-2 inactivated vaccines. Methods We conducted a comprehensive analysis of CD8 T-cell responses to SARS-CoV-2 inactivated vaccines, focusing on HLA-A2-restricted epitopes derived from the Omicron variant. Mutant epitopes were evaluated for their impact on antigen presentation and CD8 T-cell immune reactivity. Additionally, we screened for epitopes that exhibited reduced CD8 T-cell responses following the emergence of the Omicron variant. Results Our findings revealed that mutant epitopes in the Omicron variant led to escape from antigen presentation and diminished CD8 T-cell immune responses. We identified two epitopes associated with decreased CD8 T-cell reactivity post-Omicron variant emergence. Notably, we discovered an S protein epitope, 67A>V, which demonstrated similar proportions of CD8 T-cell specificity between the ancestral and mutant strains, suggesting its conservation and potential immunogenicity for vaccine development. Furthermore, the third dose of the inactivated vaccine significantly increased the number of epitope-specific CD8 T cells, underscoring the importance of booster doses in enhancing cellular immune responses against the Omicron variant. Discussion This study highlights the ability of the Omicron variant to evade CD8 T-cell immune responses through epitope mutations, while also identifying conserved epitopes with potential utility in vaccine design. The observed increase in epitope-specific CD8 T cells following a booster dose emphasizes the critical rol

关键词： 重组慢病毒载体   淋巴细胞激活基因   免疫抑制性受体   慢性病毒感染  

摘要： 目的 构建人淋巴细胞激活基因3(lymphocyte activation gene 3,LAG3)慢病毒重组表达载体,并转染小鼠成纤维细胞3T3以获得LAG3优势表达的单克隆细胞株.方法 提取人外周血单核细胞总RNA并逆转录为cDNA,以高保真DNA聚合酶PCR扩增LAG3胞外区和跨膜区基因序列.PCR产物采用限制性内切酶Eco RⅠ和NotⅠ双酶切后,与慢病毒载体pTSB-copGFP连接构建重组表达载体pTSB-LAG3-copGFP,并转化大肠埃希菌DH5α.PCR法筛选出阳性克隆菌,提取质粒DNA测序验证.pTSB-LAG3-copGFP重组载体与包装质粒psPAX2、pMD2.0G共同转染人胚肾293T细胞以包装、分泌病毒,收集病毒感染3T3细胞,采用嘌呤霉素筛选的同时有限稀释法获取LAG3稳定表达的3T3单克隆细胞.实时荧光定量PCR、免疫荧光法和流式细胞术分别检测细胞中LAG3 mRNA的转录及LAG3蛋白的表达.结果 重组pTSB-LAG3-copGFP慢病毒载体测序结果显示,扩增LAG3基因序列与GenBank中LAG3标准序列(序列号NM_002286.6)比对仅在跨膜区1 697 bp碱基位存在一处His氨基酸密码子同义突变.pTSB-LAG3-copGFP包装病毒感染3T3细胞经嘌呤霉素筛选,共获得20株LAG3+copGFP+-3T3单克隆细胞,均可检测出LAG3 mRNA的转录,其中MC-6单克隆细胞株LAG3转录水平最高.免疫荧光法与流式细胞术检测结果提示LAG3蛋白在MC-6中有效表达并分布于细胞膜和胞质细胞器膜上.结论 成功构建pTSB-LAG3-copGFP慢病毒载体,筛选获得淋巴细胞激活因子3高效表达的LAG3+copGFP+-3T3单克隆细胞株.为后续开展LAG3与乙型肝炎等慢性病毒感染性疾病发展的关系以及LAG3干预治疗等实验研究打下基础.

关键词： comparative effectiveness   multidisciplinary care   perianal fistula   setons  

摘要： Background and Aims: We aimed to assess the impact of setons on perianal fistula outcomes in patients with perianal fistulising Crohn's disease (PFCD) treated with anti-TNF therapy. Methods: We included patients treated with their first anti-TNF therapy for PFCD after undergoing a pelvic MRI between 2005 and 2022 from 6 North American centres. Our exposure was one or more setons at the time of anti-TNF therapy. Our primary outcome was major adverse fistula outcome (MAFO), a composite of repeat local surgical intervention, hospitalisation, or faecal diversion for PFCD, and our secondary outcome was fistula remission defined clinically. We used 1:1 cardinality matching and propensity score weighting to control for fistula severity based on centrally read MRIs, luminal characteristics, and concomitant therapies. Results: Our analysis included 221 patients: 81 with setons and 140 without setons. After cardinality matching, our cohorts were balanced (standardised difference < 0.1 for all covariates). Patients with setons had similar rates of MAFO (HR 1.23;95% CI, 0.68-2.21) and fistula remission at 6 months (OR, 0.81;95% CI, 0.41-1.59) and 12 months (OR, 0.63;95% CI, 0.31-1.27) compared to patients without setons. Our results remained stable when analysed by propensity score weighting and in a sensitivity analysis of patients who underwent an exam under anaesthesia. In patients with abscesses, there were lower rates of MAFO (HR, 0.49;95% CI, 0.19-1.25) but not statistically significant in patients with setons. Conclusions: In this multicentre, setons were not associated with improved fistula outcomes. Future prospective controlled studies are warranted.

关键词： Breast Cancer   Immunosuppression   Major histocompatibility complex - MHC  

摘要： Background Breast cancer (BC) continues to be a major health concern with 250,000 new cases diagnosed annually in the USA, 75% of which are hormone receptor positive (HR+), expressing estrogen receptor alpha (ER) and/or the progesterone receptor (PR). Although ER-targeted therapies are available, 30% of patients will develop resistance, underscoring the need for new non-ER/estrogen-based treatments. Notably, HR+BCs exhibit poor lymphocyte infiltration and contain an immunosuppressive microenvironment, which contributes to the limited efficacy of immunotherapies in HR+BC. In this study, we demonstrate that PR/progesterone signaling reduces major histocompatibility complex (MHC) Class I expression, facilitating immune evasion and escape from immune-based clearance of PR+*** To determine the effect of PR/progesterone on MHC Class I expression, we treated human and mouse mammary tumor cell lines with progesterone and/or interferon (IFN) and measured expression of genes involved in antigen processing and presentation (APP), as well as surface MHC Class I expression. We used the OT-I/SIINFEKL model antigen system to measure the impact of progesterone on immune cell-mediated killing of modified tumor cells. We also analyzed two large BC clinical cohorts to determine how PR expression correlates with APP gene expression and MHC Class I expression in ER-positive *** In vitro, we show that PR/progesterone signaling reduces APP gene expression and MHC class I expression in human and breast mammary tumor cell lines. PR-mediated attenuation of APP/MHC Class I expression is more pronounced in the presence of IFN. In immune cell killing assays, PR-expressing mammary tumor cells treated with progesterone are protected from immune-mediated cytotoxicity. We demonstrate that PR expression in vivo prevents immune-mediated rejection of xenoantigen-modified mammary tumor cell lines through mechanisms involving MHC Class I expression and CD8 T cells. Data analysis of

关键词： antigen presentation/processing   MHC class I   dendritic cells   cytotoxic T cells   Tapasin  

摘要： Endogenous and exogenous antigen processing and presentation through the MHC class I peptide-loading complex (PLC) are essential for initiating cytotoxic T lymphocyte responses against pathogens and tumors. Tapasin, a key component of the PLC, is produced in multiple isoforms through alternative splicing, each isoform influencing the assembly and stability of MHC class I molecules differently. While the canonical Tapasin isoform plays a critical role in stabilizing MHC class I by facilitating optimal peptide loading in the endoplasmic reticulum (ER), the other isoforms function in distinct ways that impact immune regulation. This study aimed to investigate the role of Tapasin isoforms, particularly soluble isoform 3, in modulating antigen presentation and immune responses, focusing on their effects on MHC class I peptide loading and surface expression. Our findings show that isoforms 1 and 2 stabilize TAP and facilitate efficient peptide loading onto MHC class I in the ER, promoting optimal antigen presentation. In contrast, isoform 3, which lacks both the ER retention signal and the transmembrane domain, is secreted and acts as a negative regulator. Isoform 3 inhibits the loading of exogenous peptides onto MHC class I molecules at the cell surface, thereby playing a critical role in the spatial and temporal regulation of MHC class I antigen presentation. The secreted Tapasin isoform 3 likely regulates immune responses by preventing inappropriate T cell activation and cytotoxicity, which could otherwise lead to immune-mediated tissue damage and contribute to autoimmune disorders. Understanding the distinct functions of Tapasin isoforms provides insights into immune regulation and highlights the importance of fine-tuning peptide-loading processes to ensure proper immune responses and prevent immune-related pathologies.

关键词： hypomorphic variants   IL2RG gene   common cytokine receptor gamma chain   flow cytometry standardization   STAT3 and STAT5 phosphorylation   T and B cell proliferation   plasmablast differentiation   NK degranulation  

摘要： Background Over the last decade, the identification of hypomorphic variants in patients previously diagnosed with Common Variable Immunodeficiency (CVID) has led to the association of milder phenotypes with variants of the IL2RG gene that are usually related to severe combined immunodeficiency. Indeed, several revertant mosaicisms have been described in cases with hypomorphic variants in that gene. Our main objective herein was the functional characterization of p. (Pro58Thr) variant in the IL2RG gene in an adult patient with antibody deficiency and moderate CD4+ T cell *** Evaluation of the patient included a clinical examination and a complete analysis of the peripheral blood phenotype. To further explore IL2RG functionality we selected downstream signaling readouts, namely STAT3 and STAT5 phosphorylation, NK degranulation and B- and T-cell proliferation capacity in vitro, which can be measured by flow cytometry, that reflect the strength of homeostatic signaling pathways in resting cells and after *** The patient presented reduced CD132 expression and conserved T- and B-cell proliferation capacity in vitro. However, we found that intracellular signaling downstream of IL2 gamma c is affected, with reduced STAT3 phosphorylation after IL-21 stimulation in B cells and CD4 T cells. In addition, CD4+ T cells showed a reduced STAT5 phosphorylation in response to IL-2, which was not so evident in CD8+ T cells. NK degranulation was impaired upon PHA and IL-2 as well as plasmablast differentiation in *** We conclude that p. (Pro58Thr) in the IL2RG gene is functionally a hypomorphic variant, as reported previously. Although the functionality of CD8+ is less impaired than the rest of the lymphocyte subsets, we did not detect a reversion of the variant in isolated CD8+, CD4+, CD19+ or NK cells.

关键词： Interleukin-2 (IL-2)   Interleukin-2 receptor (IL-2R)   T cell growth factor   Regulatory T cells (Tregs)   autoimmunity   cancerimmunology   transplantation  

摘要： BackgroundThe discovery of interleukin-2 (IL-2) and its receptor (IL-2R) almost 50 years ago revolutionized immunology, marking a pivotal moment in understanding T cell biology and immune regulation. Initially identified as a T cell growth factor, IL-2 unveiled critical insights into cytokine-mediated immune cell proliferation and *** review highlighted the characterization of IL-2R as a multi-chain receptor complex set a precedent for decoding cytokine receptor signaling. The unique interplay between IL-2 and its high-affinity receptor component, IL-2R alpha, epitomizes the principle of specificity and efficiency in cytokine signaling, enabling precise immune modulation. Regulatory T cells (Tregs) exploit IL-2R alpha high affinity to outcompete effector T cells for IL-2, ensuring immune tolerance and preventing *** its foundational role in immune homeostasis, leveraging IL-2 for therapeutic purposes has proven ***-2-based therapies hold transformative potential in autoimmunity, cancer immunology, and transplantation, yet they remain elusive due to the complex balance between immunostimulatory and immunosuppressive effects. This review explores the milestones in IL-2 biology, its dualistic functions, and the ongoing quest to harness its therapeutic promise.