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关键词： 人A组轮状病毒   VP6蛋白   原核表达   多克隆抗体  

摘要： 目的 本研究旨在表达和纯化人A组轮状病毒VP6蛋白，制备小鼠抗VP6蛋白多克隆抗体。方法 通过RT-PCR方法从人A组轮状病毒感染的粪便标本中扩增VP6基因，构建pET-32a-VP6原核表达质粒并转化至大肠杆菌BL21（DE3）感受态细胞；用异丙基-β-D-硫代吡喃半乳糖苷（IPTG）诱导VP6表达，采用镍柱（Ni-NTA）亲和层析纯化VP6蛋白，用SDS-PAGE和Westernblotting鉴定VP6蛋白；将VP6蛋白免疫BALB/c小鼠制备多克隆抗体，采用间接ELISA检测抗体滴度。结果 构建了pET-32a-VP6重组表达质粒，VP6蛋白主要以包涵体形式表达。经亲和层析法纯化了VP6蛋白，免疫小鼠后获得了多克隆抗体，抗体效价为1:32000，抗体可以和VP6蛋白发生特异性反应。结论 表达和纯化了人A组轮状病毒的VP6蛋白，制备了小鼠抗VP6蛋白的多克隆抗体，为人A组轮状病毒诊断抗原的制备和血清学检测方法的建立奠定了基础。

关键词： SLE   pDC   monocyte   interferon   TLR   cGAS-STING   cellular senescence  

摘要： Type I interferons (IFNs) play crucial roles in the pathogenesis of systemic lupus erythematosus (SLE). Plasmacytoid dendritic cells (pDCs) stimulated by Toll-like receptor (TLR) pathways have been thought to be the major producers of IFN alpha in patients with SLE. However, the responsiveness of pDCs from SLE patients to stimuli that produce IFN alpha differs depending on the type of TLR pathway involved. In addition to pDCs, monocytes from SLE patients were found to produce IFN alpha when responding to the cGAS-STING pathway. Here, we outline the major pathways that induce IFN alpha production by myeloid cells in SLE, and the possible mechanisms by which IFN alpha overproduction occurs by these cells. Finally, we discuss the current and future therapeutic strategies to regulate IFN alpha production in patients with SLE.

关键词： TNF alpha   molecular adjuvant   starry flounder  

摘要： Tumor necrosis factor alpha (TNF alpha) is a central pro-inflammatory cytokine that mediates host immune responses during infection. In this study, we identified and characterized the TNF alpha gene in the starry flounder (Platichthys stellatus) through transcriptomic analysis. The deduced protein contained a conserved TNF domain and transmembrane region, and phylogenetic analysis confirmed its homology with other teleost TNF alpha proteins. Tissue-specific expression profiling revealed high baseline expression in immune-related peripheral organs and a distinct temporal modulation in response to Streptococcus parauberis infection. Recombinant TNF alpha (rTNF alpha), produced using a cell-free expression system, significantly enhanced phagocytic activity in peripheral and kidney-derived leukocytes in a dose-dependent manner. Peak activity was observed at 150-200 mu g/mL, while a decline at higher concentrations suggested a threshold for immune stimulation. Importantly, hemolysis assays confirmed the safety of rTNF alpha even at the highest tested concentrations. These results demonstrate the immunomodulatory potential of TNF alpha as a molecular adjuvant in aquaculture vaccines and underscore its potential utility in immune-enhancing strategies for sustainable aquaculture.

关键词： immune monitoring   immunosuppression   kidney transplantation   translational nephrology   personalized medicine  

摘要： Kidney transplantation (KT) is the current treatment of choice in patients with end-stage kidney disease. Immunosuppression is required to prevent acute rejection but is associated with a high incidence of adverse events. The immunosuppressive burden substantially differs between individuals, necessitating new immune monitoring strategies to achieve personalization of immunosuppression. To compare the evolution of T cell profiles in correlation with immunosuppression and clinical outcomes, 87 kidney transplant recipients were followed for 12 months after KT. Flow cytometry along with assessment of T cell activation markers and clinical data was performed before KT and during study visits 10 days, 2 months and 12 months after KT. Longitudinal T cell phenotyping revealed a significant decrease of T cell activation markers HLA-DR, FCRL3, and CD147 in CD4(+) effector T cells after KT. The most pronounced reduction (75%) was found for the activation-proliferation marker HLA-DR, which persisted throughout the observational period. The decrease in HLA-DR expression reflected immunosuppressive burden through strong associations with tacrolimus trough-level exposure (coeff = -0.39, p < 0.01) and BK viremia incidence (coeff = -0.40, p < 0.01) in multivariable regression analysis. T cell activation marker HLA-DR emerges as a potential biomarker for tacrolimus-related immunosuppressive burden in association with BK viremia risk following KT.

关键词： 萆薢前列安丸   慢性非细菌性前列腺炎   细胞因子   TLR4/NF-κB/NLRP3信号通路   细胞焦亡  

摘要： 目的：基于TLR4/NF-κB/NLRP3信号通路介导的细胞焦亡探讨萆薢前列安丸治疗CNP的作用机制。方法：雄性SD大鼠适应性喂养1周，随机抽取空白组和假手术组各10只，40只大鼠制备CNP大鼠模型（1%交叉菜胶），1周判定模型，造模成功后随机分为模型组、萆薢前列安丸低、中、高剂量组，每组各10只，药物组分别给予浓度为2.7g/kg·d、5.4g/kg·d、10.8g/kg·d的药物混悬液灌胃，其余组用无菌水灌胃，每天1次，灌胃30天，末次给药后取腹主动脉血及前列腺组织（双侧），计算前列腺指数，ELISA法检测血清细胞因子IL-1β、IL-18、TNF-α浓度，HE染色检测前列腺组织炎性浸润程度，免疫组化染色检测前列腺组织中NLRP3的分布和表达，免疫荧光检测GSDMD荧光强度，Western Blot检测前列腺组织TLR4、MyD88、NF-κBp65、p-NF-κBp65、NLRP3及Caspase-1、GSDMD蛋白的表达情况。结果：萆薢前列安丸中剂量组前列腺指数显著下降(p<0.05)，萆薢前列安丸低、中、高剂量组大鼠前列腺组织炎性细胞浸润程度减低，血清IL-1β、IL-18、TNF-α水平降低(p<0.05)，免疫组化定量分析NLRP3分子表达显著减少(p<0.05)，免疫荧光定量分析GSDMD荧光强度显著降低(p<0.05)，前列腺组织TLR4、MyD88、NF-κBp65、p-NF-κBp65、NLRP3、Caspase-1、GSDMD蛋白表达降低(p<0.05)。结论：萆薢前列安丸可降低CNP大鼠促炎细胞因子水平，改善前列腺组织炎症，其作用机制可能与激活TLR4/NF-κB/NLRP3信号通路从而抑制细胞焦亡有关。

关键词： type 2 inflammation   T2-low asthma   T2-high asthma   exacerbation   eosinophil   FeNO   lung function   glucocorticoid   population-based   adults   biomarker   asthma   ACT   Asthma Control Test   ATS   American Thoracic Society   BD   Bronchodilator   BEC   Blood eosinophil count   BMI   Body mass indexCI Confidence interval   COPD   Chronic obstructive pulmonary disease   COVID-19   Coronavirus disease 2019   ER   Emergency room   ERS   European Respiratory Society   FeNO   Fractional exhaled nitric oxide   FEV   1   Forced expiratory volume in 1 second   FVC   Forced vital capacity   GINA   Global Initiative for Asthma   ICS   Inhaled corticosteroids   IL   Interleukin   ILC2   Type 2 innate lymphoid cells   IQR   Interquartile range   KCO   Carbon monoxide transfer coefficient   kUA/L   Kilo unit allergen/liter   L   Liters   LABA   Long-acting β   2   agonist   mcg   microgram   mMRC   Modified Medical Research Council   OCS   oral corticosteroid   ppb   parts per billion   SABA   short-acting β   2   agonist   sIgE   Specific immunoglobulin E   SPSS   Statistical Package of Social Sciences   SPT   Skin prick test   T2   Type 2   T   H   2   T helper type 2   TLCO   Transfer factor for carbon monoxide   WSAS   West Sweden Asthma Study   χ   2   Chi square  

摘要： Background The ‘type 2 (T2)-high’ phenotype has a relatively well-defined pathophysiology compared to ‘T2-low’ asthma. T2-low asthma has been mostly studied among severe asthma cases. Objective To estimate the proportion and assess the clinical features of asthma with low T2 markers in a general asthma population, including all severities. Methods Participants with current asthma from the population-based adult West Sweden Asthma Study (WSAS) were included. High T2 markers were defined as blood eosinophil count (BEC) ≥0.15×10 9 /liter (L) or fractional exhaled nitric oxide (FeNO)≥ 20 parts per billion (ppb) or asthma being allergen driven. If none of these were present, it was considered as asthma with low T2 markers. Results In total, 896 participants were included, of which 14.3% had low T2 markers. The low T2 markers group was predominantly females and had later asthma onset age than the high T2 markers group. Participants with low T2 markers had better spirometry results in terms of airflow obstruction parameters but had more asthma-related emergency visits in the past 12 months than those with asthma with high T2 markers. Conclusion In a general adult asthma population, one in seven have low T2 markers according to the criteria used in this study. Importantly, these individuals had better lung function than individuals with high T2 markers, but a significantly increased risk of asthma exacerbations. Thus, having asthma with low T2 markers is an independent risk factor for exacerbations, which should be considered when assessing asthma patients clinically.

关键词： Myeloproliferative neoplasms   PEG-IFN alpha 2a   Shortage   Outcomes  

摘要： Between March to December 2024, we observed a PEG-IFN alpha 2a shortage in France for MPN patients. We identified and followed 90 patients from our center and observed 80% of difficulties obtaining the drug, and 61 patients among all (67.8%) did not receive their prescribed drug on time (impacted group). Twenty-one patients need to reduce the dosage of PEG-IFN alpha 2a and 27 stopped the drug. Ten clinical events were identified with nine in the impacted group (p=0.04). No hematological or only partial responses were more frequent in the impacted group (p=0.01). The allelic burden variations were also studied during this period;17 patients obtained a VAF <10%. Eight patients stopped the drug because of VAF <4% and remained with molecular response. Three-quarters of the patients experimented with psychological and financial impacts of shortage. This study is the first one to explore a shortage of cytoreductive drugs happened in MPN. Much informations will be helpful for clinicians.

关键词： bispecific antibody   cytokine mimetic   IL-18   immunotherapy   nanobody  

摘要： Cytokines are promising in cancer immunotherapy, but their pleiotropic effects limit specificity and clinical utility. Through binding to IL-18Rα and IL-18Rβ, interleukin-18 (IL-18) stimulates innate lymphocytes and effector T cells for antitumor immunity. However, clinical trials of recombinant IL-18 have been hampered by IL-18 binding protein (IL-18BP), a secreted high-affinity decoy receptor. Here, we developed decoy-resistant bispecific nanobodies that maintain IL-18 signaling potential. Based on agonistic nanobodies targeting IL-18Rα and IL-18Rβ, bispecific nanobody A4B2-mdFc effectively enhanced CD8 + T cells responses with distinct transcriptomic profiles. Systemic delivery of A4B2-mdFc boosted CD8 + T cells infiltration and activation, demonstrating dose-dependent antitumor efficacy in cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) models. Interestingly, PDCD1 and CTLA-4 expressions were drastically increased on CD8 + T cells when treated with A4B2-mdFc. Injection of A4B2-mdFc significantly improved the antitumor efficacy of immune checkpoint inhibitors (ICIs) targeting both PD-1 and CTLA-4. Our findings demonstrated that nanobody-based bispecific IL-18 mimetics elicited superior antitumor activity via CD8 + T cells activation and IL-18BP resistance, providing the potential application of cytokine-targeting bispecific nanobody monotherapy or in combination with ICIs for cancer immunotherapy.

关键词： Akkermansia muciniphila   antiobesity   hypoacylated rough-type LPS   TLR4-IL-23-IL-22 immune axis  

摘要： Lipopolysaccharides (LPS) derived from intestinal symbionts plays a critical role in modulating and maintaining mucosal immunity. In this study, we investigated the chemical characteristics and antiobesity properties of Akkermansia muciniphila HW07 LPS (ALPS). ALPS was identified as hypo-acylated, mono/bis-phosphorylated, rough-type LPS. Compared to Escherichia coli LPS (ELPS), ALPS functions as a weak agonist of TLR4/TLR2. Intraperitoneal administration of ALPS in diet-induced obese (DIO) mice suppressed weight gain, improved metabolic parameters, restored gut barrier integrity, and modulated the gut microbiota. Notably, ALPS treatment significantly increased plasma interleukin (IL)-22 levels. Furthermore, neutralizing IL-22 with an antibody eliminated the antiobesity effects of ALPS in DIO mice. Mechanistically, ALPS upregulated the expression of both IL-22 and its upstream cytokine IL-23 in a TLR4-dependent manner. These findings confirm that activation of the TLR4-IL-23-IL-22 immune axis is a key mechanism underlying the antiobesity effect of ALPS. In acute toxicity assessment, no fatalities were observed in ALPS-treated mice, whereas ELPS treatment led to a 40% mortality rate. Collectively, our results demonstrate that hypo-acylated LPS from A. muciniphila functions as a metabolically beneficial immune modulator that exerts immunomodulatory effects through the TLR4-IL-22 axis and suggests ALPS as a promising novel therapeutic strategy for metabolic disorders.