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关键词： IL-33   ILC2 cells   ST2 receptor   allergic response   immunotoxin   ribotoxin α-sarcin  

摘要： Allergies constitute one of the major health problems worldwide, increasing their prevalence in developed countries. To overcome this multifactorial disease, immunotherapy and the use of immune molecules, such as immunotoxins, have arisen as promising therapeutic tools. We have designed, produced, and characterized a new immunotoxin called IL-33αS, encompassing the murine IL-33 (mIL-33) as the target domain and the ribotoxin α-sarcin as the toxic domain. IL-33 is a widely described alarmin that binds to the ST2 receptor of a variety of immune cells, including ILC2s, leading to Th2-derived inflammatory response, as occurs in allergic reactions. Both IL-33αS and mIL-33 were successfully produced in the methylotrophic yeast and purified to homogeneity through affinity chromatography for their characterization. Both IL-33αS and mIL-33 were able to specifically bind to ST2 Raw 264.7 cells, and IL-33αS kept the ribonucleolytic activity of α-sarcin, allowing IL-33αS to exhibit cytotoxic effects against ST2-targeted cells. In addition, IL-33αS induced significantly less secretion of the Th2-linked cytokine IL-13 in comparison to mIL-33, suggesting steric interference produced by the presence of the α-sarcin. These results assess the potential therapeutic effect of this new immunotoxin against allergies, causing ST2-targeted cytotoxicity while avoiding the Th2 cytokine secretion.

关键词： C反应蛋白   原核表达   分子伴侣   单克隆抗体  

摘要： [目的]本研究旨在制备猪C反应蛋白（C-reactive protein， CRP）的单克隆抗体，为建立用于监测猪血清中CRP蛋白浓度的血清学方法提供生物材料。[方法]通过优化CRP基因并克隆至原核表达载体pET28a，并将分子伴侣质粒pTF16与重组质粒共转化并表达重组CRP蛋白，将该蛋白作为免疫原免疫BALB/c雌鼠，通过细胞融合、间接ELISA筛选，得到分泌抗重组CRP蛋白的杂交瘤细胞，制备单克隆抗体，进行效价测定及亚型鉴定，采用Western blot检测其特异性，进行生物素标记制备生物素标记单克隆抗体。[结果]成功构建重组质粒及重组表达载体，经细胞融合和亚克隆成功获得2株分泌抗重组猪CRP蛋白单克隆抗体的杂交瘤细胞，分别命名为CRP-Mab1和CRP-Mab2，两株杂交瘤细胞上清效价分别为1：409 600和1：204 800。Western blot结果显示，2株单克隆抗体均能与重组猪CRP蛋白特异反应。亚型鉴定结果显示，CRP-Mab1抗体轻链属于κ型，重链属于IgG1型，CRP-Mab2单克隆抗体轻链属于κ型，重链属于IgM型。进行生物素标记并获得标记的单克隆抗体CRP-Mab1-Bio，其效价可达1：12 800。[结论]成功制备2株针对重组猪CRP蛋白的单克隆抗体，并进行生物素标记，为猪CRP蛋白浓度检测及炎性情况监测方法的开发提供了重要的生物材料。

关键词： 桑术养正饮   呼吸道感染   淋巴细胞   呼吸道菌群   免疫  

摘要： 目的 观察桑术养正饮对健康成年人免疫功能及上呼吸道菌群的影响。方法 采用随机、空白对照研究，纳入60名健康成年受试者，随机分为对照组、试验组。对照组不进行药物干预；试验组给予桑术养正饮口服14天。试验第0、14天采用FC500流式细胞仪检测血液中淋巴细胞亚群；采用16S rRNA技术检测上呼吸道微生态优势菌群，使用PICRUSt2软件预测菌群的功能，包括京都基因与基因组百科全书（KEGG）等代谢通路等功能的预测。结果 试验组29名受试者及对照组30名受试者完成研究并纳入统计分析。与本组干预前比较，试验组干预后总T淋巴细胞、总B淋巴细胞、辅助T淋巴细胞、细胞毒性T淋巴细胞、咽部菌群γ-变形杆菌纲（Gamma-Proteobacteria）、嗜血菌属（Haemophilus）相对丰度升高，KEGG功能预测分析显示，遗传信息通路上调（P<0.05）;Sobs、Ace、Chao指数、咽部菌群放线菌门（Actinobacteria）、互养菌门（Synergistetes）、放线菌纲（Actinobacteria）相对丰度降低，KEGG功能预测分析显示，人类疾病通路、细菌性传染性疾病通路、分解代谢通路下调（P<0.05,P<0.01）。与对照组同期比较，试验组干预后总T淋巴细胞、总B淋巴细胞、辅助T淋巴细胞、细胞毒性T淋巴细胞升高（P<0.05）。结论 桑术养正饮干预后淋巴细胞亚群水平升高及呼吸道微生态的有益重塑，提示桑术养正饮可能对某些呼吸道感染具有一定防治作用。

关键词： AISI   SCC-Ag   早期宫颈癌   淋巴结转移  

摘要： 目的:本研究旨在探讨术前全身炎症综合指数（aggregate index of systemic inflammation,AISI）以及其与鳞状细胞癌抗原（squamous cell carcinoma antigen,SCC-Ag）预测早期宫颈癌患者发生盆腔淋巴结转移的临床价值。方法:回顾性分析188例接受根治性手术治疗的早期宫颈癌患者临床资料。计算术前AISI。单因素及多因素分析影响淋巴结转移的危险因素。应用R软件绘制可视化列线图并进行验证。绘制AISI及SCC-Ag联合AISI预测淋巴结转移的受试者工作特征（ROC）曲线,得出诊断精度。结果:本研究中淋巴结转移率21.8%。单因素分析淋巴结转移与糖尿病、FIGO分期、肿瘤大小、肿瘤浸润深度、肿瘤脉管浸润及AISI有关,均P<0.05。绘制AISI预测淋巴结转移的ROC曲线。计算AISI曲线下面积AUC值为0.782（95%CI:0.736～0.838）,最佳截断值为128.885,灵敏度为0.878,特异度为0.66。SCC-Ag+AISI阳性定义为SCC>2.5,AISI>128.885。计算SCC-Ag+AISI曲线下面积AUC为0.836（95%CI:0.775～0.886）。通过DElong检验,SCC-Ag+AISI与AISI比较差异有统计学意义（Z=2.104,95%CI:0.004～0.104,P=0.035）。Logistic回归分析淋巴结转移与糖尿病、FIGO分期、脉管浸润、AISI及SCC-Ag+AISI有关,均P<0.05。Hosmer-Lemeshow检验结果χ2=1.408,P=0.994。使用Logistic回归分析结果构建早期宫颈癌患者淋巴结转移风险预测列线图模型。采用Bootstrap法将入组样本自助抽样1 000次对模型行内部验证,平均绝对误差为0.019。ROC曲线分析显示,列线图的最佳截断值为101.31分,C-index为0.779,AUC为0.936（95%CI:0.892～0.963）,灵敏度为0.853,特异度为0.925。结论:本研究发现术前AISI可预测早期宫颈癌患者发生淋巴结转移,与糖尿病、FIGO分期、脉管浸润都是淋巴结转移的独立危险因素。当AISI>128.885时,淋巴结转移风险增高。SCC-Ag和AISI的联合检验可以更好地预测淋巴结转移。通过列线图分析,当危险因素总分≥101.31提示淋巴结转移风险增高。

关键词： Ovarian cancer   Lobaric acid   Tocilizumab   STAT3   Immunotherapy  

摘要： BackgroundOvarian cancer is the most deadly gynecological malignancy, largely due to difficulties in early detection, therapeutic resistance and toxicity. The aim of this study was to evaluate the potential therapeutic effects of the small-molecule drug candidate lobaric acid (LA) as monotherapy and in combination with tocilizumab (TCZ) and atezolizumab (ATE) in ovarian *** and resultsIn this study, the anti-proliferative and immunotherapeutic effects of LA, TCZ, and ATE in different ovarian cancer cell lines were determined. In addition, the intracellular signalling effects of LA and its combination with TCZ were extensively investigated by silencing the STAT3 pathway and the results were confirmed. This study demonstrated for the first time that the combination of LA and TCZ exerts a significant immunotherapeutic effect on ovarian cancer cells. LA treatment, either as monotherapy or in combination with TCZ, was found to have therapeutic potential for ovarian cancer without cytotoxic effects on healthy *** findings emphasise that LA is a promising candidate for the treatment of ovarian cancer and indicate its potential for inclusion in advanced treatment protocols.

关键词： span>major histocompatibility complex (MHC)membrane recyclingtraffickingkineticsplasma membranebeta2-microglobulin dissociationcell surface retentionfree heavy chains (fH)in silico modelingcross-presentation  

关键词： Tumor necrosis factor (TNF-alpha)   IL-6 receptor   Rheumatoid arthritis   Bispecific antibody  

摘要： Tumor necrosis factor-alpha and interleukin-6 are proinflammatory cytokines involved in several autoimmune diseases, including rheumatoid arthritis. Although monoclonal antibodies targeting these cytokines or their receptors are successful treatment approaches for autoimmune diseases, approximately 30% of treated patients fail to respond. Therefore, the designation of more effective and versatile therapeutics for simultaneously targeting multiple inflammatory pathways is pivotal. This study aimed to design a recombinant bispecific antibody (BisAb) targeting TNF-alpha and IL-6 receptor based on the complementarity-determining regions (CDRs) of adalimumab and tocilizumab antibodies. The physicochemical properties and tertiary structure of the modeled bispecific antibody proteins were studied through bioinformatics. The interaction of the designed bispecific antibody with human TNF-alpha and IL-6R was computed by molecular docking and molecular dynamics. The recombinant BisAb (60kDa) was expressed in Escherichia coli. The ELISA results demonstrated that the affinity of BisAb and its Ada-scFv moiety to TNF-alpha was quite similar with Kaff values of 7.7 and 9.4 x 10-13 M, respectively. The recombinant BisAb and Toci-scFv also represented similar affinities towards IL-6R (Kaff values of 1.65 and 1.87 x 10-12 M), closely comparable with the parental tocilizumab antibody. A dose-dependent neutralization of TNF-mediated cytotoxicity was also observed on L929 cells. Decreased phosphorylation of signal transducer and activator of transcription 3 (STAT3) was determined through western blotting with specific antibodies, indicating the blockade of IL-6 receptors before the preincubation with IL-6. The designed BisAb, a minimized biotherapeutic molecule, could successfully target two main ligands (IL-6R and TNF-alpha) involved in rheumatoid arthritis and many inflammatory/infectious *** points center dot A new bispecific antibody was designed to target TNF-alpha and IL-6

关键词： HLA alleles   Immune factors   Rheumatoid arthritis  

摘要： Objective Rheumatoid arthritis (RA) is a chronic, systemic autoimmune disorder characterized by persistent inflammation and joint pain. However, the pathogenesis of RA remains incompletely understood. The aim of this study was to investigate the interaction between HLA alleles and immune factors in relation to the risk of RA. Methods Linear regression model was used to assess the association between 133 HLA alleles (with gene frequencies > 1%) and RA. Additionally, we evaluated the potential effects of interactions between HLA alleles and 26 simple immune indicators, as well as four composite immune indicators, on the risk of RA. All analyses were grouped by gender to further explore the potential impact of gender on HLA alleles and immune indicators in RA risk. Results The results showed 37 HLA alleles significantly associated with RA. In the subgroups, 17 and 29 HLA alleles were significantly associated with RA in males and in females, respectively. We identified 44 significant interactions between human leukocyte antigen (HLA) alleles and immune indicators on RA. Further, sex-stratified analyses identified 37 and 45 associations between HLA alleles x immune indicators and RA in males and in females, respectively. Conclusion This study identified interactions between various HLA alleles and immune indicators that may increase or decrease the risk of RA. These findings may further advance the study of immune genetic factors in RA and contribute to the clinical assessment of RA by HLA alleles and various immune indicators.

关键词： PKC delta   Tight junction   ZO-1   proliferative retinopathy   vascular leakage  

摘要： Proliferative retinopathies are associated with abnormal angiogenesis that can result in visual impairment or vision loss. The tight junction complex regulates blood-retinal barrier integrity; however, its role in proliferative retinopathies is still at an early stage. Here, we employed human retinal endothelial cells (HRMVECs), and a mouse model of oxygen-induced retinopathy (OIR) to investigate the impact of IL-33 signaling on tight junction disintegration and pathological angiogenesis. Our experimental findings demonstrate that IL-33 induces ZO-1 serine/threonine phosphorylation and tight junction disruption in HRMVECs. In addition, mass-spectroscopy (MS) analysis revealed that treating HRMVECs with IL-33 induces ZO-1 phosphorylation at Thr861 residue. Furthermore, we observed that NOX1-PKC-δ signaling modulates IL-33-induced ZO-1 phosphorylation and tight junction integrity in HRMVECs. We also observed that IL-33 depletion significantly reduces OIR-induced NOX1-PKCδ-ZO-1 signaling, vascular leakage in the ischemic retina. We also observed that the NOX1-specific inhibitor, fluoflavine (ML-090), attenuated OIR-induced NADPH oxidase activity and pathological retinal neovascularization in the ischemic retina. Thus, we infer that IL-33-mediated NOX1-PKCδ-ZO-1 signaling regulates ischemia-induced retinal endothelial cell tight junction disruption and retinal neovascularization.