白细胞介素-18及其抗肿瘤作用研究进展

白细胞介素18(IL-18)具有多向免疫调节功能。它对T细胞向Th1或Th2分化起有独特的调节能力。最近的研究表明了IL-18的结构、活化形式和调节树突状细胞、NK细胞等方面的重要功能。研究也发现许多肿瘤患者血清IL-18水平显著升高及其在肿瘤发生发展中的作用。本文主要介绍了IL-18的结构、产生与调节、生物学作用及其在肿瘤方面的研究进展。     

白细胞介素18的生物学作用

白细胞介素 18(IL - 18)是新近发现的细胞因子 ,具有诱导 IFN-γ的产生 ,促进 T细胞的增殖 ,促进 Th1细胞的发育和增殖 ,增强 NK细胞及 Th1细胞的细胞毒性 ,增强粒细胞 -巨噬细胞集落刺激因子 (GM- CSF)的产生 ,抗肿瘤效应 ,抑制 Ig E的生成以及破骨细胞的形成等多种生物学活性和作用     

白细胞介素-18研究进展

白细胞介素 １８（ＩＬ １８）,是最近发现的一种新型免疫细胞调节因子,能显著刺激Ｔｈ１细胞产生ＩＦＮ γ,所以最初命名为ＩＧＩＦ（ＩＦＮ γｉｎｄｕｃｉｎｇｆａｃｔｏｒ）。它与ＩＬ １２有协同作用,却不互相依赖。此外,它还能增强ＮＫ细胞和ＦａｓＬ介导的细胞毒效应,能促进内毒素引发的肝损伤。在肿瘤、自身免疫性疾病及抗炎症治疗中可能有潜在的应用前景     

白细胞介素-2加强小鼠T淋巴细胞产生白细胞介素-3

本文报道了白细胞介素-2(IL-2)刺激ConA(5μg/ml)活化的小鼠T细胞产生的条件培养液(TCM)中含有CFU-GEMM诱导活性。这种CFU-GEMM诱导活性的生成在IL-2作用后48h达到高峰。特异性抗IL-3单克降抗体可以完全中和该条件培养液中的CFU-GEMM诱导活性。进一步证明,TCM可以刺激IL-3依赖细胞系FDC-P_1细胞的增殖;在IL-2作用于ConA活化的T细胞后可促进其细胞表达高水平的IL-3mRNA。这些结果表明IL-2可以加强小鼠T细胞产生IL-3。     

白细胞介素18

白细胞介素-18(IL-18)是新发现的细胞因子,具有多种生物学功能.IL-18能促进外周血单个核细胞产生干扰素-γ(IFN-γ)、IL-2、粒细胞巨噬细胞集落刺激因子(GM-CSF)等细胞因子,增强天然杀伤细胞(NK细胞)的细胞毒作用.IL-18结构上与IL-1相似,而功能更接近IL-12,IL-18与IL-12均能诱导Th1细胞产生IFN-γ,存在协同效应,但它们的作用途径不同.IL-18在抗感染抗肿瘤等方面有着潜在的应用前景,并与自身免疫性疾病的发病密切相关.     

人白细胞介素18的基因克隆与表达

人白细胞介素是新近发现的细胞因子之一。研究表明它参与Ｔ１辅助细胞介导的细胞免疫,利用ＲＴ－ＰＣＲ技术从人外周血细胞扩增得到了ＩＬ－１８的ｃＤＮＡ并测定其核酸序列。利用基因生组技术构建ＩＬ－１８的表达载体,并在大肠杆菌中进行了表达。     

白细胞介素—2及其受体与肿瘤基因治疗

作为最重要的细胞因子之一,白细胞介素－2（IL－2）抗瘤作用的研究一直引导着肿瘤生物免疫治疗的方向,本从分子生物学的和细胞生物学角度介绍IL－2的结构,激活信号系统,功能活性,各免疫细胞的IL－2分泌动力学特点,IL－2受体亚单位组成,表达调控,功能意义及在不同细胞中的表达状况,IL－2基因转导接种疗法的基本方法,作用原理,主要优点,近年获得的实验与临床研究数据,以及待解决的问题等。     

白细胞介素18在免疫应答中的作用

白细胞介素18(IL-18)是IL-1细胞因子超家族中的一员,是γ干扰素(IFN-γ)细胞因子产生的主要诱导因子之一[1],在调节机体的免疫反应中起着重要的作用,可调节天然免疫应答和获得性免疫应答,而且也是一个能够在不同的免疫环境中调节Th1或Th2类免疫应答的细胞因子[2].     

乳果糖对严重烫伤大鼠白细胞介素-18介导的脏器功能的影响

目的：探讨乳果糖对烫伤大鼠白细胞介素-18 mRNA（IL-18 mRNA）介导的肺、肝、肾脏功能的影响。方法：将大鼠致30%体表面积Ⅲ度烫伤,随机分成2组（n=40）：对照组（C组）和乳果糖预防组（L组）。用改良的鲎试验基质显色法定量分析循环内毒素-脂多糖（LPS）含量;采用逆转录PCR技术检测肺、肝、肾组织中IL-18 mRNA水平;用改良的方法检测肺组织髓过氧化物酶（MPO）活性：使用7170型全自动生化分析仪测定血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）、血清尿素氮（BUN）、血清肌酐（Cr）。结果：L组LPS含量伤后22、4 h显著低于C组（P〈0.05）;伤后C组各时点肺、肝、肾组织中IL-18 mRNA表达较伤前增强（P〈0.01）,伤后L组各脏器组织IL-18 mRNA表达明显低于C组（P〈0.05~0.01）;L组伤后81、6 h MPO活性显著低于C组（P〈0.05~0.01）;L组伤后2 h8、h1、6 h,ALT、AST、BUN、Cr明显低于C组（P〈0.05~0.01）。结论：烧伤早期循环LPS含量居高不下,肺、肝、肾等脏器IL-18 mRNA表达明显过度,其介导所致脏器的功能遭受严重损害,而经过乳果糖预处理可以在一定程度上保护肺、肝、肾等脏器功能。     

携带白介素-18基因的人脐带间充质干细胞的构建及其意义(英文)

目的:构建携带白介素-18(IL-18)基因人脐带间充质干细胞(hUMSCs),为肿瘤靶向性基因治疗研究提供一种工具。方法:体外分离培养hUMSCs,流式细胞仪(FACS)检测hUMSCs的细胞免疫表型。应用基因重组技术将表达IL-18基因的慢病毒转染至hUMSCs,利用RT-PCR及Western blot法检测IL-18的蛋白及mRNA表达水平。结果:成功在体外分离和培养了hUMSCs,流式细胞仪检测结果显示hUMSCs表达CD29、CD44和CD105,而不表达CD34和CD45,符合hUMSCs的表型。成功构建携带IL-18基因的hUMSCs,RT-PCR及Western blot法检测结果提示IL-18基因转染至hUMSCs并能稳定表达。结论:构建携带IL-18基因的hUMSCs并稳定表达IL-18,为肿瘤靶向性基因治疗实验性研究提供了一种新实验工具。     

内江猪IL-18基因的克隆及原核表达

以ConA刺激的内江猪外周血单核淋巴细胞(PBMC)为模板,采用RT-PCR技术扩增出猪IL-18全长基因,克隆其成熟蛋白的编码基因(471bp)至pMD18-T克隆载体,经双酶切和测序获得阳性克隆。通过EcoRⅠ/XhoⅠ双酶切及连接反应,构建了pET-32a( )-IL-18原核表达质粒。经双酶切和DNA测序证实重组质粒构建正确,将阳性重组质粒转化大肠杆菌BL21(DE3),IPTG诱导表达,SDS-PAGE,Western-blot证实表达出35kD左右的融合蛋白。经实验确定重组内江猪IL-18蛋白诱导表达的最佳条件为IPTG 0.2mmol/L,30℃诱导培养5h。在最佳表达条件下,pIL-18的相对含量为59.6%。内江猪IL-18的原核表达为重组IL-18的纯化及其生物学功能的进一步研究提供了基础。     

突变型人IL-18的肿瘤靶向性改造及真核表达

为了构建肿瘤靶向性的人突变型IL-18新基因并进行真核表达,以重组PCR技术构建EGF-IL-18融合基因,利用 Bac-to-Bac杆状病毒表达系统和Sf 9昆虫细胞株（来自秋天草地夜蛾）表达融合基因,纯化表达产物,并以IFN-γ诱导实验和EGFR竞争结合实验初步评价融合蛋白的生物活性。测序证明构建的融合基因为原设计EGF-IL-18融合基因。SDS-PAGE和Western blot证明EGF-IL-18融合基因在昆虫细胞中获得表达,表达的融合蛋白的Mr约为20 000,与理论值相符,纯化后融合蛋白具有特异的IL-18单抗结合活性。IFN-γ诱导实验和EGFR竞争结合实验显示,该融合蛋白具有肿瘤导向性和抗肿瘤活性。表明对突变型IL-18成功地进行了肿瘤导向性改造并使其在真核细胞获得表达。     

Role of IL‐18 in atopic asthma is determined by balance of IL‐18/IL‐18BP/IL‐18R

It is recognized that IL‐18 is related to development of asthma, but role of IL‐18 in asthma remains controversial and confusing. This is largely due to lack of information on expression of IL‐18 binding protein (BP) and IL‐18 receptor (R) in asthma. In this study, we found that plasma levels of IL‐18 and IL‐18BP were elevated in asthma. The ratio between plasma concentrations of IL‐18 and IL‐18BP was 1:12.8 in asthma patients. We demonstrated that 13‐fold more monocytes, 17.5‐fold more neutrophils and 4.1‐fold more B cells express IL‐18BP than IL‐18 in asthmatic blood, suggesting that there is excessive amount of IL‐18BP to abolish actions of IL‐18 in asthma. We also discovered that more IL‐18R+ monocytes, neutrophils and B cells are located in asthmatic blood. Once injected, IL‐18 eliminated IL‐18R+ monocytes in blood, but up‐regulated expression of IL‐18R in lung macrophages of OVA‐sensitized mice. Our data clearly indicate that the role of IL‐18 in asthma is very likely to be determined by balance of IL‐18/IL‐18BP/IL‐18R expression in inflammatory cells. Therefore, IL‐18R blocking or IL‐18BP activity enhancing therapies may be useful for treatment of asthma.     

Microbubbles Enhance the Antitumor Effects of Sinoporphyrin Sodium Mediated Sonodynamic Therapy both In Vitro and In Vivo

Objectives: To evaluate the anti-cancer effect of sonodynamic therapy combined with microbubbles both in vitro and in vivo.Methods: Cell viability was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide and guava viacount assays. Annexin V-FITC/PI staining was adopted to analyze cell apoptosis rate. FD500 uptake assay was performed to assess cell membrane permeability changes. Tumor weight, mice weight and the visual image of tumor size were used to reflect the anti-tumor effect of this combined method. Histological change of tumor tissue after different treatments was measured through hematoxylin and eosin (H&E) staining.Results: Microbubbles can significantly enhance the cytotoxicity and necrocytosis rate induced by SDT treatment. Increased cell membrane permeability and more uptake of DVDMS were founded in SDT combined with microbubbles group. For in vivo experiments, SDT with microbubbles can significantly reduce tumor weight and size with pimping difference of mice weight compare with other treatment groups. In addition, microbubbles notably improved tumor tissue destruction caused by ultrasound and SDT treatment.Conclusion: The results suggest that microbubbles can markedly improve the anti-cancer effect of DVDMS mediate sonodynamic therapy both in vitro and in vivo.     

猪IL-18 在杆状病毒/昆虫细胞中的表达

IL-18作为一种多效性细胞因子,在机体免疫应答及各种生理功能中发挥着重要的调节作用,根据猪IL-18基因序列设计1对引物,将编码猪IL-18的成熟蛋白基因亚克隆到杆状病毒转移载体pFastBacDual中,并在C端融合6个组氨酸标签以利于纯化,然后转化到含穿梭载体Bacmid的感受态细胞DH10Bac中,发生转座作用。将重组质粒转染昆虫细胞,SDS-PAGE可检测到分子量为18 kDa左右的重组蛋白,Western blotting证实该重组蛋白可与兔抗猪IL-18抗体发生特异性反应。纯化后蛋白能明显促进猪T淋巴细胞转化,表明所表达的IL-18具有较高的生物活性。此研究为进一步开发研制新型免疫佐剂奠定了基础。     

小鼠白细胞介素18在大肠杆菌中的表达、纯化及抗肿瘤作用研究

用RT PCR从小鼠肝脏细胞中扩增出小鼠白细胞介素 18(mIL 18)的cDNA ,克隆到表达载体pJW2中 ,经热诱导后在大肠杆菌JM109中表达。表达的重组mIL-18(rmIL-18)主要以包涵体形式存在 ,占菌体总蛋白质的 18%。包涵体用 5mol L尿素溶解后 ,经SephadexG-100柱纯化。纯化的蛋白质经透析复性 ,在 0.5mg LConA存在的条件下 ,能够对小鼠脾细胞具有剂量依赖的IFN γ诱生作用。以 1× 105 H22 肝癌细胞腹腔注射昆明小鼠 ,构建小鼠H2 2 肝癌腹水瘤模型。分别在肿瘤细胞接种后 1、4和 8d ,用 10 μg纯化产物对荷瘤小鼠进行腹腔注射。结果显示rmIL 18注射组小鼠死亡速率延缓 ,生存率显著提高 (P <0 .0 1) ,达到 62.5 %。首次证明mIL-18对小鼠H2 2 肝癌具有显著的抑制作用 ,经rmIL 18作用后存活小鼠具有对H22 肝癌细胞的免疫记忆性     

Interleukin-18 enhances HIV-1 production in a human chronically-infected T cell line (H9-V)

Interleukin-18 (IL-18) is a recently identified immunoregulatory cytokine expressed by activated macrophages, that induces production of interferon-gamma (IFN-gamma) and Th-1 development. Recently some investigators reported controversial in vitro data on IL-18 stimulation of HIV-1 replication in several cell lines. In the present study the effect of IL-18 on HIV replication in a human chronically HIV-1-infected lymphocytic T cell line (H9-V) was investigated. HIV-1 replication was determined by an immunoassay method in order to evaluate the content of p24 antigen in the cell culture supernatants. Stimulation of H9-V cells with IL-18 resulted in increased production of p24, especially at concentrations of 0.01 microg ml(-1) and 0.10 microg ml(-1). Moreover a significant and persistent IL-18 stimulation of HIV-1 replication was observed at a concentration of 0.01 microg ml(-1) during a 7-day period. Pre-treatment of IL-18 with a specific neutralizing monoclonal antibody significantly reduced HIV-1 replication. These experiments show that IL-18 promotes the increase of HIV-1 replication in human chronically-infected lymphocytic T cells and confirm the role of IL-18 as a proimflammatory cytokine in stimulating and maintaining HIV-1 replication during the course of the disease. In a successive set of experiments, since one of the main activities of IL-18 is the induction of IFN-gamma, we evaluated the effect of this biological modifier on H9-V cells. In particular, IFN-gamma shows a significant effect on cell replication and on reduction of CD4 and CD71 surface expression.     

Expression of bovine (Bos indicus) interleukin‐18 in Escherichia coli and its biological activity

IL‐18 modulates immune functions by inducing IFN‐γ production and promoting Th1 immune responses. In the present study, we amplified and cloned the sequence (582 bp) encoding full‐length bovine IL‐18 from PBMC stimulated with PHA. The nucleotide and the deduced amino acid sequence of Bos indicus IL‐18 showed an identity of 86–98% compared with IL‐18 sequences of other ruminants. The insert was subcloned into a pET 32a vector and expressed in Escherichia coli as a fusion protein and the matured protein was obtained by caspase I treatment. The specificity of these proteins was confirmed by western blotting. The biological activity of the purified protein was analyzed by its ability to induce IFN‐γ production in PBMC measured by ELISA and qPCR.     

猪肌肉素基因的cDNA克隆与表达

从人肌肉素基因出发, 在dbEST数据库中进行同源性搜索, 找到七个有较高同源性的Expressed Sequence Tag(DY426490, CF787546, AJ660979, AJ664670, AJ663820, AJ680159, DN106254)。通过拼接和进一步RT-PCR实验验证, 获得猪肌肉素基因全长cDNA序列, 其全长651 bp, 开放阅读框为54~452 bp, 编码有132个氨基酸。同源性分析结果表明, 与人、小鼠和大鼠的肌肉素基因cDNA编码区(CDS)同源性分别为87.2%、77.6%和77.9%。利用克隆出的猪肌肉素cDNA, 构建表达载体pGEX-4T-1-musclin, 并在BL21大肠杆菌中成功表达和纯化了分子量为38.59 kD的融合蛋白GST-Musclin, 并运用蛋白印迹技术进行鉴定。     

Glial Expression of Interleukin-18 and its Receptor After Excitotoxic Damage in the Mouse Hippocampus

Interleukin (IL)-18, a member of the IL-1 cytokine family, is an important mediator of peripheral inflammation and host defence responses. However, although IL-1 is a key proinflammatory cytokine in the brain, little is known about IL-18 changes in glial cells under excitotoxic neurodegeneration. In this study, we characterized the expressions of IL-18 and IL-18 receptor (IL-18R) in kainic acid (KA)-induced excitotoxicity in mouse hippocampus by immunohistochemistry and Western blotting. IL-18 immunoreactivity was found in microglia whereas IL-18R immunoreactivity was observed in astrocytes. Levels of IL-18 and IL-18R in hippocampus homogenates increased progressively from day 1 post-KA and peaked at 3 days. This study demonstrates the cellular sources of IL-18 and IL-18R, and their temporal correlations after KA-insult, and suggests roles for IL-18 and IL-18R in glial cells in response to excitotoxic damage in the hippocampus.