白细胞介素-1α与其Ⅰ型可溶性受体的相互作用及动力学特征

研究了基于亲和型生物传感器的白细胞介素-1α(IL-1α)与I型可溶性白细胞介素-1受体(sIL-1RⅠ)相互作用的实时监测,并应用FASTfit软件对数据进行拟合分析。采用共价交联法在样品池表面固定sIL-1RⅠ,其密度为2.13 ng/mm2,用于检测不同浓度IL-1α与sIL-1RⅠ的结合所引起响应信号变化。对浓度为2400 nmol/L IL-1α的响应信号为122.66弧度秒,并且响应信号同IL-1α浓度呈线性关系,相关系数为0.972。FASTfit拟合数据结果表明IL-1α与sIL-1RⅠ相互作用符合一级动力学方程,其误差值不超过0.5弧度秒,其解离平衡常数(KD)和结合平衡常数(KA)分别为4.15×10-6mol/L和2.41×105mol/L。验证了亲和型生物传感器研究IL-1α和sIL-1RⅡ相互作用及动力学分析的可行性,有助于阐明IL-1α和sIL-1RⅠ的重要生物学功能,并为快速筛选其拮抗剂研究提供了理论和实验数据。     

以9943 Tc-二乙三胺五乙酸-白细胞介素三元体系对白细胞介素-2进行标记和检测

白细胞介素-2(IL-2)是机体免疫网络系统起重要调节作用的淋巴因子,它能激活多种免疫细胞使其产生免疫活性.以9943 Tc标记IL-2,用9943 Tc-二乙三胺五乙酸(DTPA)-IL-2的三元体系产物作为标记物进行检测,然后将标记物用于体内的IL-2受体显像的研究,有助于认识某些疾病的发生机理,可用于监测某些疾病的动态变化和实验性治疗.     

新生儿感染性肺炎血清Gal-3、IL-17水平与肺超声评分关系及对继发呼吸窘迫综合征的预测价值

为探讨新生儿感染性肺炎(NIP)血清半乳糖凝集素-3(Gal-3)、白细胞介素-17(IL-17)水平与肺超声评分关系及对继发呼吸窘迫综合征(RDS)的预测价值,本研究选取NIP患儿98例作为研究组,根据是否继发RDS分为RDS患儿和无RDS患儿,选取同期健康新生儿98例作为对照组,均进行血清Gal-3、IL-17水平检测,研究组进行肺超声检查。结果显示研究组血清Gal-3、IL-17水平高于对照组,血清Gal-3、IL-17水平与肺超声评分呈负相关;Gal-3、IL-17是NIP继发RDS的独立危险因素,胎龄是NIP继发RDS的独立保护因素;血清Gal-3、IL-17水平联合预测NIP继发RDS的AUC值大于单一指标预测值,患者受益良好。由此可见,NIP患儿血清Gal-3、IL-17水平与肺超声评分关系密切,临床通过检测血清Gal-3、IL-17水平可评估NIP患儿继发RDS的风险。     

人嗜中性白细胞活化蛋白-1/白细胞介素-8基因的半化学半酶促合成及在大肠杆菌的表达

本文采用半化学半酶促法合成了人嗜中性白细胞活化蛋白-1/白细胞介素-8(NAP-1/IL-8)的全基因,并在大肠杆菌系统中利用P_RP_1串联启动子进行了温控型的高效表达,通过SDS聚丙烯酰胺凝胶电泳(SDS-PAGE)检测表明,在摇瓶培养的大肠杆菌中,NAP-1/IL-8的表达量可占菌体可溶性蛋白的18.5％,而在实验室规模的发酵试验中,其表达量则占可溶性蛋白的10.9％.经凝胶过滤层析和阳离子交换层析两个简单的纯化步骤,得到SDS-PAGE纯的重组NAP-1/IL-8,采用琼脂糖平板法进行测定,表明所得NAP-1/IL-8纯品在<10ng/ml的水平上显示出强烈的嗜中性白细胞趋化活性,家兔皮肤试验的结果显示,重组人NAP-1/IL-8可在家兔体内引起早发型超敏反应,采用Edman降解法测定重组人NAP-1/IL-8纯品N末端36个氨基酸的序列,与天然人NAP-1/IL-8的成熟分子完全符合。     

IL-1β在体内代谢调节机理探讨

本文利用抗人重组白细胞介素1β(rHIL-1β)单克隆抗体,对患者胸腔积液和脑脊液中IL-1β抗原、抗IL-1β抗体及含IL-1β特异免疫复合物三项指标进行动态检测。结果发现,标本中检出IL-1β抗原后,随着治疗和病程发展,其抗原含量伴随着检出抗IL-1β抗体和特异性免疫复合物而逐渐消失。文中认为,IL-1β抗体的出现,对IL-1β的生物学活性起反馈调节作用,并通过形成免疫复合物加速IL-1β的代谢。     

超声参数联合血清OPN、IL-1β对乳腺癌的临床诊断价值研究

选取我院收治的乳腺癌(BC)患者82例设为研究组,同期乳腺良性肿瘤患者82例设为对照组,探讨了超声参数联合血清骨桥蛋白(OPN)、白细胞介素-1β(IL-1β)对BC的诊断价值。结果显示,研究组患者病灶处的RI值、PI值和血清OPN、IL-1β水平均高于对照组(P<0.05);PI值、RI值、血清OPN、IL-1β水平与临床分期、肿瘤直径、远处转移存在正相关关系(P<0.05);预后良好者PI、RI、OPN、IL-1β低于预后不良者(P<0.05);超声参数联合血清OPN、IL-1β诊断BC的AUC值为0.900,预测预后的AUC值为0.875。本研究证实BC患者存在超声参数及血清OPN、IL-1β水平异常情况,且异常程度与疾病病理特征关系密切,通过超声参数及相关血清因子水平可对乳腺病变情况进行鉴别,并指导临床制定治疗方案。     

聚合物基质高效液相色谱填料在白细胞介素-2分离中的应用

聚合物型苯乙烯-二乙烯基苯(PS-DVB)微球装填的麦科菲反相高效液相色谱柱(MKF-RP-HPLC),用于白细胞介素-2(IL-2)分离的最佳色谱条件:流动相A:0.1%三氟乙酸,流动相B:80%乙腈 0.1%三氟乙酸.B液在30 min内从0%线性增大至100%,流速1.0 mL/min;检测波长:280 nm.在该色谱条件下进行系统性实验,结果表明:MKF-RP色谱柱分离IL-2的柱效、分离度、重复性和拖尾因子均能达到药典要求,且柱效和分离度与SOURCE色谱柱相当;MKF-RP色谱柱的pH适用范围为1～14;柱压与SOURCE色谱柱相当,且低于Hypersil C8色谱柱和Polaris C18色谱柱的柱压,可在更高流速下操作;MKF-RP色谱柱的非极性与SOURCE色谱柱相当;MKF-RP色谱柱的超载性能优于SOURCE色谱柱,其对IL-2进样液的最大载样量为300 μg.     

在昆虫细胞中克隆与表达人白细胞介素6

本文利用DNA合成及PCR技术对人白细胞介素6(hIL-6)cDNA进行转译优化与扩增,并同杆状病毒载体pVL 1393重组构建了pVL·IL-6载体;通过磷酸钙共沉淀法将pVL·IL-6 DNA与野生型苜蓿银纹夜蛾核型多角体病毒(wtAcMNPV)DNA共转染昆虫细胞Sf9,筛选出表达人IL-6的重组病毒rAc·IL-6,经ELISA定量测定rhIL-6表达水平约为1μg/ml;rhIL-6生物活性经IL-6依赖细胞系7TD1测定为10~6u/ml。     

纯化基因重组白细胞介素-2的初步研究

 采用羟基磷灰石 (HAP)作为填料 ,以pH 6 8的磷酸盐缓冲液为流动相 ,用制备型高效液相色谱来分离提纯基因重组白细胞介素 2 (IL 2 ) ,通过SDS 聚丙烯酰胺凝胶 (SDS PAGE)电泳测定IL 2的纯度 ,测定其活性为1× 1 0 6 U/mg。该法对IL 2的纯化、含量测定及活性测定等效果良好。     

螺旋CT成像联合血清copeptin、IL-18水平检测在急性脑梗死患者神经功能及预后评估中的应用价值

本文选取74例急性脑梗死(ACI)患者作为研究对象,入院时根据美国国立卫生院卒中量表(NIHSS)评分分为重度组(NIHSS评分>15分,n=21)、中度组(NIHSS评分5~15分,n=24)、轻度组(NIHSS评分<5分,n=29),均接受血清copeptin和IL-18水平检测及螺旋CT成像检查。结果发现,随ACI神经功能缺损加重,血清copeptin、白细胞介素18(IL-18)水平及峰值时间升高,CBV、CBF异常下降。经ROC曲线显示,血清IL-18联合CBF预测ACI患者预后不良的曲线下面积(AUC)最大。由上述结果可见螺旋CT成像参数、血清copeptin、IL-18水平与ACI患者神经功能存在一定相关性,三者联合为临床实现ACI患者个体化治疗和改善预后提供了可能。     

An optical biosensor for kinetic analysis of soluble Interleukin-1 receptor I binding to immobilized Interleukin-1alpha

We report here the development of an optical biosensor based on the resonant mirror for kinetic analysis of soluble Interleukin-1 receptor I (sIL-1R I) in solution binding to immobilized Interleukin-1α (IL-1α). IL-1α was immobilized through its surface amine groups via amide bonds with the carboxyl groups of the carboxymethyl dextran (CMD) on cuvette surface. The interaction of sIL-1R I and IL-1α was monitored in real time. Evaluation of the binding curves allowed the analysis of the binding kinetics. The linear range of sIL-1R I in solution was over a range of 100-1600 nM (R = 0.9962). Equilibrium dissociation constant (K_D) was derived by Scatchard plot analysis for sIL-1R I binding to immobilized IL-1α. For this assay, the K_D was 2.6 × 10⁻⁶ M. The CMD cuvette modified by IL-1α was successfully regenerated using 10 mM HCl, and the same sensing surface was used repeatedly for the interaction analysis.     

Flow-injection immuno-bioassay for interleukin-6 in humans based on gold nanoparticles modified screen-printed graphite electrodes

A flow-injection electrochemical immunoassay system based on a disposable immunosensor for the determination of interleukin-6 (IL-6) was proposed. The immunosensor was prepared by entrapping horseradish peroxidase (HRP)-labeled IL-6 antibody into gold nanoparticles-modified composite membrane at a screen-printed graphite electrode. With a non-competitive immunoassay format, the immunosensor was inserted in the flow system with an injection of sample, and the injected sample containing IL-6 antigen was produced transparent immunoaffinity reaction with the immobilized HRP-labeled IL-6 antibody. The formed antigen–antibody complex inhibited partly the active center of HRP, and decreased the immobilized HRP to H₂O₂ reduction. The performance and factors influencing the performance of the immunosensor were investigated. Under optimal conditions, the current change obtained from the labeled HRP relative to thionine–H₂O₂ system was proportional to the IL-6 concentration in the range of 5–100 ng L⁻¹ with a detection limit of 1.0 ng L⁻¹ (at 3δ). The flow-injection immunoassay system could automatically control the incubation, washing and measurement steps with acceptable reproducibility and good stability. Moreover, the proposed immunosensors were used to analyze IL-6 in human serum specimens. Analytical results of clinical samples show the developed immunoassay has a promising alternative approach for detecting IL-6 in the clinical diagnosis.     

Molybdenum trioxide hybridized kaempferol: double-powered nanosystem for salvaging oxidative stress and electrochemical immunoprobing of interleukin-6

Intracellular reactive oxygen species (iROS) are the culprit in inflammation-linked diseases. Excessive radical generation triggers an inflammation cascade involving interleukin-6 (IL-6) and other cytokines release, causing oxidative stress to cells. Developing healthcare materials with dual-functionality controlling iROS and diagnosing IL-6 would be extremely beneficial for chronic inflammatory disease management. Herein, molybdenum trioxide hybridized kaempferol nanoparticles (MoHK NPs) have been synthesized with iROS scavenging and in situ electrochemical redox property for immunoassay of IL-6. Physicochemical integrity of nanosystem comprising MoHK NPs is characterized by X-ray absorption/photoelectron, Raman, and fourier transform infrared (FT-IR) spectroscopy as well as scanning transmission electron microscopy–high-angle annular dark field microscopic analysis. In vitro radical scavenging mechanism of MoHK NPs was studied by electron paramagnetic spectroscopy. Distinctly, these MoHK NPs exhibit a clinically significant antioxidant function and cytocompatibility with RAW 264.7 macrophage cell line. Bioaffinity layer–assisted monoclonal antibodies of IL-6 immobilized on MoHK electrode enable superior selectivity, electrochemical signal transduction (sensitivity 0.63 μA/fM/cm²), and rapid analytical response time even at ultralow IL-6 concentrations (detection limit 0.91 fM). This work demonstrates that hybridizing redox-active and antioxidant-rich phytochemical on metal oxide nanosystem can be a promising strategy for multifunctional theranostics.     

Lipopolysaccharide-stimulated interleukin-6 production for radioimmunoassay

Prepration of liquid phase radioimmunoassay (RIA) to assessment the interleukin-6 (IL-6) that produced in-house by microbial stimulation was the aim of the present study. The production of IL-6 through mouse and macrophage cell infections were caused by Escherichia coli lipopolysaccharide (LPS). Optimum dose of lipopolysaccharide was determined. Then, both ¹²⁵I- IL-6 tracer and polyclonal anti-IL-6 antibody were prepared. These reagents were used in detection of produced interleukin-6. Optimization and validation tests of local RIA were carried out. The results showed that the interleukin-6 produced by microbial macrophage or mouse infections gave positive qualitative detection using enzyme linked immunoassay sorbent assay. The local RIA was used in quantitative determination of local purified interleukin-6.     

Dual inhibitors of Interleukin-6 and acetylcholinesterase for treatment of Alzheimer's disease: Design,docking, synthesis and biological evaluation

Multitarget compounds intercept two or more functionally complementary pathways simultaneously, and are therefore considered to have potential in effectively treating complex multifactorial diseases like Alzheimer's disease (AD). In the present study, novel molecules are designed by coupling a chromone and a N,N-disubstituted carbamoyl amine as pharmacophore for interleukin-6 (IL-6) and acetylcholinesterase (AChE) inhibition, respectively. Four series (Y1–Y4) of 40 compounds are designed by using alkyl linkers of different lengths (1–4 carbon atoms) for the coupling of the two selected pharmacophore. Docking of all designed compounds in AChE leads to the identification of twelve best fit compounds (Docking score >8.3). The data suggests that a 1- or 2-carbon atom linker is the most conducive to orient the pharmacophore for optimum binding with AChE active site. The predicted ADME properties of the 12 selected compounds suggest that these can cross the blood brain barrier (BBB) with good oral bioavailability. These compounds are synthesised and evaluated for anti-AChE activity. Five compounds, showing >45% inhibition of AChE, are further evaluated for IL-6 inhibitory activity. Compound Y1f is found to be the most potent inhibitor of both AChE and IL-6 (IC₅₀ 0.7 and 0.8 ​μM, respectively). It suggests that a chromone moiety connected to a piperidine ring through a 1-carbon atom linker may provide a useful template to medical chemists for the development of new chemical entities effective against AD.     

维生素B6在纳米金/石墨烯修饰电极上的电化学行为

将金纳米粒子电沉积在石墨烯修饰的玻碳电极表面,研究了维生素B6(VB6)在该修饰电极上的电化学行为。扫描电镜用于该修饰电极组装过程的形貌表征。实验结果表明:VB6在此修饰电极上出现一个良好的氧化峰,在最佳实验条件下,其氧化峰电流与VB6浓度在5.0×10-8～2.0×10-5 mol/L范围内呈线性关系,其线性回归方程为I(μA)=0.5697c(μmol/L)+0.06275,R=0.9992,检出限为2.0×10-8 mol/L(S/N=3)。一些常见的干扰物质如抗坏血酸不干扰VB6的检测。方法已用于片剂中VB6的含量的检测。     

维生素B_6在纳米Au-N,P/石墨烯修饰电极上的电化学行为

将金纳米粒子(AuNPs)电沉积在N,P/石墨烯(N,P/Graphene)修饰的玻碳电极表面,研究了维生素B_6(VB_6)在该修饰电极上的电化学行为。实验结果表明:VB_6在该修饰电极上出现一个良好的氧化峰,在最佳实验条件下,其氧化峰电流与VB_6的浓度在2.0×10~(-5)~4.0×10~(-4) mol/L范围内呈线性关系,相关系数R=0.998,检出限为9.2×10~(-6) mol/L。一些常见的物质如K~+、Na~+、Zn~(2+)、葡萄糖(Glu)不干扰VB_6的检测。此方法已用于片剂中VB_6含量的检测,获得较好结果。     

Development of molecularly imprinted polymer films used for detection of profenofos based on a quartz crystal microbalance sensor

A quartz crystal microbalance (QCM) sensor based on molecularly imprinted ultra-thin films was developed for detecting profenofos in real samples. Films prepared by physical entrapment (MIP-A) and in situ self-assembly (MIP-B) were compared. The results indicated that the best sensing signal was obtained through the in situ self-assembly method. The QCM sensor chip was pretreated with 11-mercaptoundecanoic acid (MUA) to form a self-assembled monolayer (SAM), and then polymer films were immobilized directly on the SAM using surface-initiated radical polymerization. In this paper, all detection experiments were taken in air. The reaction was processed in solution, and the electrode was washed with deionized water and dried with N(2) before QCM measurement. The film was characterized by a scanning electron microscope (SEM), AC impedance and cyclic voltammetry. Analysis of the QCM response in the presence of different concentrations of profenofos showed a good linear correlation during 1.0 × 10(-8) to 1.0 × 10(-5) mg mL(-1) (y = 5log x + 42.5, R = 0.9960) and 1.0 × 10(-5) to 1.0 × 10(-3) mg mL(-1) (y = 25.86log x + 146, R = 0.9959), respectively. The MIP-QCM sensor was used to detect profenofos in tap water, and showed good recovery and repeatability.     

A novel dual-impedance-analysis EQCM system--investigation of bovine serum albumin adsorption on gold and platinum electrode surfaces

Both quartz crystal micro-balance (QCM) impedance and electrochemical impedance spectroscopy (EIS) methods are widely used in interface studies. This paper presents details about a new strategy for simultaneous, mutual-interference-free and accurate measurements of QCM impedance and EI, through connecting a suitable capacitance in series with the piezoelectric quartz crystal (PQC) between QCM impedance and EIS measurement instruments. Combined and individual measurements of QCM impedance and EIS during silver deposition gave results comparable with each other, demonstrating the reliability of the proposed method. Bovine serum albumin (BSA) adsorption on gold and platinum electrodes in Britton-Robinson (B-R) buffers was investigated, and the Fe(CN)6(3-)/Fe(CN)6(4-) couple was used as an electrochemical probe to characterize BSA adsorption. While the reversibility of Fe(CN)6(3-)/Fe(CN)6(4-) couple on bare Au and Pt electrodes changed very slightly with decreasing solution pH from pH approximately 7 to pH approximately 2, the standard rate constant (ks) of this couple increased abruptly with solution pH below pH approximately 4.5 at a BSA-modified Au electrode, but decreased with solution pH at a BSA-modified Pt electrode. By analyzing the QCM impedance data with a modified BVD equivalent circuit and the EI data with a modified Randle's equivalent circuit, inflexion changes at pH approximately 4.5 were all found at pH-dependent responses of the resonant frequency, the double-layer capacitance, the capacitance of the adsorbed BSA layer, the peak-absorbance values of BSA solutions at 277.5 and 224.5 nm, and so on. It was also found that a BSA adsorption layer can effectively inhibit gold corrosion during ferrocyanide oxidation in a ferrocyanide-containing BR solution. Some preliminary explanations of these findings have been given. The proposed method is highly recommended for wider applications in surface science.     

Adsorption behavior of microbes on a QCM chip modified with an artificial siderophore-Fe3+ complex

Three hydroxamate-type artificial siderophores with terminal NH(2) groups, tris[2-{3-(N-acyl-N-hydroxamino)propylamido}propyl]aminomethane (1-3, acyl-R group = Me, Et, and Ph, respectively), and their Fe(3+) complexes, 4-6, were prepared. The stability constant (log β) of 4 was estimated to be about 31 by its EDTA titration. The biological activities of 4-6 for Microbacterium flavescens, which is a hydroxamate-type siderophore, auxotrophic gram-positive microbe, clearly indicated that they permeated the cell membrane depending on their terminal bulky acyl-R groups. These artificial siderophore complexes, 4-6, were modified on Au electrode surfaces with the terminal NH(2) group (4-6/Au). The surface modification of 4-6 was confirmed by several electrochemical measurements. The quartz crystal microbalance (QCM) chips were also modified with 4-6. Microbe adsorption measurements using these modified QCM chips for M. flavescens, Pseudomonas putida, and Eschrichia coli were performed. The QCM chips have the ability to adsorb microbes selectively as a result of the differences in the interactions between the structures of Fe(3+)-artificial siderophore complexes and their receptors or binding proteins within the cell membrane.