作者： Zhao, Feijun;Liu, Shuangquan;Zhang, Xiaohong;Yu, Jian;Zeng, Tiebing;...

期刊： Human Vaccines & Immunotherapeutics,2013年9(4):753-760 ISSN：2164-5515

通讯作者： Wu, Yimou

作者机构： [Wu, Yimou; Zhao, Feijun; Zeng, Tiebing] Univ South China, Pathogen Biol Inst, Hengyang City, Hunan, Peoples R China.;Centers Dis Control Hunan Prov, Changsha, Hunan, Peoples R China.;[Liu, Shuangquan] Cent S Univ, Dept Anat & Neurobiol, Xiangya Sch Med, Changsha, Hunan, Peoples R China.;[Liu, Shuangquan; Cao, Xunyu] Univ South China, Affiliated Hosp 1, Hengyang, Hunan, Peoples R China.;[Zhang, Xiaohong] Univ South China, Dept Histol & Embryol, Sch Med, Hengyang, Hunan, Peoples R China.

通讯机构： [Wu, Yimou] U;Univ South China, Pathogen Biol Inst, Hengyang City, Hunan, Peoples R China.

关键词： DNA vaccine;CpG ODN;membrane protein;Treponema pallidum;Tp Gpd;mucosal adjuvant;mucosal immune

摘要： Objectives: The protective response against Treponema pallidum (Tp) infection of a DNA vaccine enhanced by an adjuvant CpG ODN was investigated. Results: The mucosal adjuvant CpG ODN enhanced the production of higher levels of anti-TpGpd antibodies induced by pcD/Gpd-IL-2 in rabbits. It also resulted in higher levels of secretion of IL-2 and IFN-α, and facilitated T cell proliferation and differentiation (p < 0.05). No significant difference about testing index above-mentioned was found in the intranasal immunization group of pcD/Gpd-IL-2 vaccine adjuvanted by CpG ODN when compared with the immunization by pcD/ Gpd-IL-2 vaccine intramuscular injection alone (p > 0.05). Furthermore, CpG ODN stimulated the production of mucosaspecific anti-sIgA antibodies and resulted in the lowest Tp-positive rate (6.7%) for Tp-infection of skin lesions and the lowest rates (8.3%) of ulceration lesions, thus achieving better protective effects. Methods: New Zealand rabbits were immunized with the eukaryotic vector encoding recombinant pcD/Gpd-IL-2 using intramuscular multi-injection or together with mucosal enhancement via a nasal route. The effect of the mucosal adjuvant CpG ODN was examined. Conclusions: The CpG ODN adjuvant significantly enhances the humoral and cellular immune effects of the immunization by pcD/Gpd-IL-2 with mucosal enhancement via nasal route. It also stimulates strong mucosal immune effects, thus initiating more efficient immune-protective effects. © 2013 Landes Bioscience.

语种： 英文

作者： 曾焱华;何军;游晓星;唐双阳;朱翠明;...

期刊： 中华微生物学和免疫学杂志,2013年33(4):287-292 ISSN：0254-5101

通讯作者： Wu, Y.-M.

作者机构： [唐双阳; 游晓星; 余敏君; 朱翠明; 吴移谋; Zeng Y.-H.] Institute of Pathogenic Biology, University of South China, Hengyang 421001, China

通讯机构： [Wu, Y.-M.] I;Institute of Pathogenic Biology, University of South China, China

关键词： 生殖支原体;黏附蛋白;模拟表位;多抗原肽;免疫原性

摘要： 目的 制备基于生殖支原体黏附蛋白(MgPa)模拟表位的多抗原肽(MAP),检测其诱导小鼠产生特异性体液免疫和细胞免疫应答的水平,为研制安全、有效的基于模拟表位的Mg表位肽疫苗奠定实验基础.方法 以多聚赖氨酸为核心基质,人工合成3种含MgPa模拟表位的八分枝MAP,用反相高效液相色谱(RP-HPLC)和质谱分析对其进行纯化与鉴定.将其单独或混合免疫BALB/c小鼠.间接ELISA测定小鼠血清中的特异性IgG抗体及其亚类的水平；MTT比色法检测MAP刺激免疫小鼠脾淋巴细胞的增殖反应,并用ELISA检测其脾细胞培养上清中的IL-4和IFN-γ水平.结果 成功合成了较高纯度的含MgPa模拟表位的3种MAP,这3种MAP均能刺激小鼠产生IgG抗体,且以Th1型免疫应答产生的IgG2a型抗体为主；与单个MAP组相比,混合MAP免疫组小鼠产生了更多的IgG、IgG1和IgG2a抗体(P均＜0.01)；3种MAP均能特异性诱导免疫小鼠的脾淋巴细胞增殖,且能刺激其产生较多的IL-4和IFN-γ,混合MAP免疫组小鼠产生的IL-4和IFN-γ,与单个MAP组相比差异具有统计学意义(P均＜0.01).结论 基于MgPa模拟表位的3种MAP均能刺激小鼠产生较强的体液免疫和细胞免疫应答,且混合免疫组诱导的免疫应答效果比单个MAP组更好.

语种： 中文

作者： 邓红玉;李忠玉;吴移谋;周辉;马康康;...

期刊： 中华微生物学和免疫学杂志,2013年33(2):107-111 ISSN：0254-5101

通讯作者： Li, Z.-Y.

作者机构： [邓红玉; 李忠玉; 吴移谋; 周辉; 马康康; 陆春雪] 南华大学医学院微生物学与免疫学教研室;[钟光明] 美国德州大学圣安东尼奥健康科学中心

通讯机构： Institute of Pathogenic Biology, Medical College, University of South China, China

关键词： 沙眼衣原体;pORF5质粒蛋白;免疫损伤;细胞因子

摘要： 目的 研究沙眼衣原体(Chlamydia trachomatis,Ct)pORF5质粒蛋白对小鼠生殖道组织的免疫损伤作用,并初步探讨其损伤机制.方法 表达纯化GST-pORF5融合蛋白,融合蛋白经PreScission Protease酶切和去内毒素处理后分别在第1、3、6天接种于BALB/c小鼠阴道后穹窿,第7天处死小鼠,分离小鼠生殖道组织,肉眼大体观察并进行炎症病理评分；ELISA方法检测血清、阴道灌洗液和小鼠脾细胞培养上清中的TNF-α、IL-1β和IL-6细胞因子水平.结果 pORF5蛋白接种组小鼠输卵管壶腹部与峡部出现不同程度的肿胀,周围结缔组织发生黏连,并出现了不同程度的扭曲与积水,而PBS和GST(谷胱甘肽S-转移酶)蛋白对照组输卵管组织未出现明显改变；同时生殖道组织炎症病理积分明显高于PBS对照组(P＜0.01)和GST对照组(P＜0.01)；pORF5蛋白接种组小鼠脾细胞培养上清、阴道分泌物中TNF-α、IL-1β、IL-6水平均显著高于PBS和GST蛋白对照组(P＜0.05)；血清中TNF-α、IL-1β水平显著高于PBS和GST蛋白对照组(P＜0.01).结论 pORF5蛋白能引起BALB/c小鼠生殖道组织免疫损伤,该损伤机制可能与TNF-α、IL-1β、IL-6等炎症因子水平升高有关.

语种： 中文

作者： Jiang, Chuanhao;Zhao, Feijun;Xiao, Jinhong;Zeng, Tiebing;Yu, Jian;...

期刊： Clinical and Vaccine Immunology,2013年20(10):1563-1568 ISSN：1556-6811

通讯作者： Wu, Yimou

作者机构： [Jiang, Chuanhao; Wu, Yimou; Zhao, Feijun; Zeng, Tiebing; Wu, Haiying; Ma, Xiaohua] Univ South China, Coll Med, Pathogen Biol Inst, Hengyang, Peoples R China.;[Xiao, Jinhong] Hunan Prov Peoples Hosp, Clin Lab, Changsha, Hunan, Peoples R China.;[Yu, Jian] Univ South China, Coll Med, Dept Expt Zool, Hengyang, Peoples R China.

通讯机构： [Wu, Yimou] U;Univ South China, Coll Med, Pathogen Biol Inst, Hengyang, Peoples R China.

摘要： Syphilis is a chronic infection caused by Treponema pallidum subsp. pallidum, and diagnosis with sensitive and specific methods is a challenging process that is important for its prevention and treatment. In the present study, we established a recombinant protein TpF1-based indirect immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) and a Western blot assay for human and rabbit sera. The 20-kDa recombinant protein TpF1 was detected by Western blotting performed with sera from rabbits immunized with recombinant TpF1 and infected with the T. pallidum Nichols strain and T. pallidum clinical isolates but was not detected by Western blotting with sera from uninfected rabbits. The sensitivity of the recombinant protein was determined by screening sera from individuals with primary, secondary, latent, and congenital syphilis (n = 82). The specificity of the recombinant protein was determined by screening sera from uninfected controls (n = 30) and individuals with potentially cross-reactive infections, including Lyme disease (n = 30) and leptospirosis (n = 5). The sensitivities of TpF1-based ELISAs were 93.3%, 100%, 100%, and 100% for primary, secondary, latent, and congenital syphilis, respectively, and the specificities were all 100% for sera from uninfected controls and individuals with potentially cross-reactive infections. In Western blot assays, the sensitivities and specificities of TpF1 for human sera were all 100%. The reactivities of TpF1 with syphilitic sera were proportional to the titers of the T. pallidum particle agglutination (TPPA) assay. These data indicate that the recombinant protein TpF1 is a highly immunogenic protein in human and rabbit infections and a promising marker for the screening of syphilis. Copyright © 2013, American Society for Microbiology. All Rights Reserved.

语种： 英文

作者： 谢小平;刘双全;张秋桂;吴移谋

期刊： 中华皮肤科杂志,2013年46(5):305-308 ISSN：0412-4030

作者机构： [谢小平; 刘双全; 张秋桂] 南华大学第一附属医院检验科;[吴移谋] 南华大学病原生物学研究所

关键词： 密螺旋体;苍白;重组蛋白质类;免疫活性

摘要： 目的探讨梅毒螺旋体重组蛋白TP0993在梅毒血清学诊断中的应用价值。方法通过生物信息学分析,获取TP0993基因序列,构建原核载体进行诱导表达;Ni-NTA亲合层析柱纯化重组蛋白,Western印迹检测其免疫抗原性,用重组蛋白直接注射免疫新西兰家兔,评价其免疫原性。用纯化的TP0993重组蛋白包被微孔板,建立间接酶联免疫吸附试验(ELISA)方法,检测临床梅毒患者血清和健康体检者正常血清,同时与梅毒螺旋体明胶凝集试验(TPPA)进行比较,根据重组蛋白与梅毒阴阳性血清的反应情况,评价重组抗原在梅毒血清学诊断中的应用价值。结果成功构建PET-28a(+)-0993原核表达载体,经表达、纯化后获得相对分子质量约为34 000的重组蛋白;Western印迹检测显示该重组蛋白能与梅毒患者阳性血清发生特异性反应;利用纯化的TP0993重组蛋白免疫新西兰兔,能诱导新西兰兔产生特异性免疫应答。以重组蛋白为包被抗原建立间接ELISA法,对TPPA法检测的480份临床血清进行检测,与TPPA法比较ELISA法的灵敏度为88.3%,特异度为85.8%,ELISA法与TPPA法符合率为86.5%。结论重组表达的TP0993蛋白具有较好的免疫活性,可作为梅毒血清学诊断的候选抗原之一。

语种： 中文

作者： Zhou Hui;Huang QiuLin;Li ZhongYu*;Wu YiMou;Xie XiaoBing;...

期刊： 中国科学:生命科学(英文版),2013年56(5):460-466 ISSN：1674-7305

通讯作者： Li ZhongYu

作者机构： [Wu YiMou; Cao WenJuan; Zhou Zhou; Lu ChunXue; Zhou Hui; Li ZhongYu; Ma KangKang] Univ South China, Pathogen Biol Inst, Hengyang 421001, Peoples R China.;[Xie XiaoBing; Zhou Hui] Hunan Univ Chinese Med, Hosp 1, Dept Lab Med, Changsha 410007, Hunan, Peoples R China.;[Huang QiuLin] Univ South China, Affiliated Hosp 1, Dept Gen Surg, Hengyang 421001, Peoples R China.;[Zhong GuangMing] Univ Texas Hlth Sci Ctr San Antonio, Dept Microbiol & Immunol, San Antonio, TX 78229 USA.

通讯机构： [Li ZhongYu] U;Univ South China, Pathogen Biol Inst, Hengyang 421001, Peoples R China.

关键词： Chlamydia trachomatis;pORF5 plasmid protein;mitogen-activated protein kinase;proinflammatory cytokines;TLR2

摘要： Infection with Chlamydia trachomatis induces inflammatory pathologies in the urogenital tract that can lead to infertility and ectopic pregnancy. Pathogenesis of infection has been mostly attributed to excessive cytokine production. However, precise mechanisms on how C. trachomatis triggers this production, and which protein(s) stimulate inflammatory cytokines remains unknown. In the present study, the C. trachomatis pORF5 protein induced tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta (IL-1 beta) and interleukin-8 (IL-8) in dose- and time-dependent manners in the THP-1 human monocyte cell line. We found that intracellular p38/mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK)/MAPK signaling pathways were required for the induction of TNF-alpha, IL-1 beta and IL-8. Blockade of toll-like receptor 2 (TLR2) signaling reduced induction levels of TNF-alpha, IL-8 and IL-1 beta. We concluded that the C. trachomatis pORF5 protein might contribute to the inflammatory processes associated with chlamydial infections.

语种： 英文

作者： 曾怀才;孙诗博;李武;吴移谋

作者机构： [曾怀才; 孙诗博; 李武; 吴移谋] 南华大学基础医学博士后流动站,湖南衡阳421001;[曾怀才] 南华大学公共卫生学院,湖南衡阳421001

会议名称： 中国毒理学会第六届全国毒理学大会

会议时间： 2013-11-12

会议地点： 广州

会议主办单位： 中国毒理学会

会议论文集名称： 中国毒理学会第六届全国毒理学大会论文集

关键词： 全氟辛烷磺酸盐;孕期暴露;心脏;凋亡

摘要： 目的探讨孕期全氟辛烷磺酸盐（PFOS）暴露对子鼠心脏细胞的凋亡效应及线粒体凋亡通路的作用。方法 40只SD孕鼠随机分成对照组、PFOS 0.1,0.6和2.0 mg·kg-1·d-1组,从受孕第2天开始（GD2）天到GD21结束,每天上午8.00灌胃一次。在出生后21 d（PND21）处死幼鼠,并迅速分离心脏组织,提取mRNA,经逆转录形成cDNA,-80℃保存备用,或者4%多聚甲醛固

语种： 中文

作者： Lu, Chunxue;Peng, Bo;Li, Zhihong;Lei, Lei;Li, Zhongyu;...

期刊： Microbes and Infection,2013年15(4):329-338 ISSN：1286-4579

通讯作者： Wu, Yimou

作者机构： [Li, Zhongyu; Lu, Chunxue; He, Qingzhi; Wu, Yimou; Chen, Li; Zhong, Guangming] Univ South China, Dept Microbiol & Immunol, Hengyang 421001, Hunan, Peoples R China.;[Peng, Bo] Univ South China, Canc Res Inst, Hengyang 421001, Hunan, Peoples R China.;[Li, Zhihong] Cent S Univ, Xiangya Hosp 2, Dept Surg, Changsha 410000, Hunan, Peoples R China.;[Lei, Lei; Zhong, Guangming] Univ Texas Hlth Sci Ctr San Antonio, Dept Microbiol & Immunol, San Antonio, TX 78229 USA.;[Wu, Yimou] Univ South China, Dept Microbiol & Immunol, 28 West Changsheng Rd, Hengyang 421001, Hunan, Peoples R China.

通讯机构： [Wu, Yimou] U;Univ South China, Dept Microbiol & Immunol, 28 West Changsheng Rd, Hengyang 421001, Hunan, Peoples R China.

关键词： Chlamydia muridarum;Protective immunity;Macrophage infectivity potentiator;Vaccine

摘要： We previously reported that 5 Chlamydia muridarum antigens reacted with antisera from >90% mice urogenitally infected with C. muridarum and they are TC0660 (ABC transporter or ArtJ), TC0727 (outer membrane complex protein B or OmcB), TC0828 (macrophage infectivity potentiator or MIP), TC0726 (inclusion membrane protein or Inc) & TC0268 (hypothetical protein or HP). The orthologs of these antigens in Chlamydia trachomatis were also highly reactive with antisera from women urogenitally infected with C. trachomatis. In the current study, we evaluated these C. muridarum antigens for their ability to induce protection against a C. muridarum intravaginal challenge infection in mice. We found that only MIP induced the most pronounced protection against C. muridarum infection. The protection correlated well with robust C. muridarum MIP-specific antibody and Th1-dominant T cell responses. The MIP-immunized mice displayed significantly reduced live organism shedding from the lower genital tract and highly attenuated inflammatory pathologies in the upper genital tissues. These results demonstrate that MIP, an immunodominant antigen identified by both human and mouse antisera, may be considered a component of a multi-subunit chlamydial vaccine for inducing protective immunity.

语种： 英文

作者： Yu, Qin;Su, Yunkai;Lu, Bing;Ma, Yan;Zhao, Xiuqin;...

期刊： PLOS ONE,2013年8(5):e57660- ISSN：1932-6203

通讯作者： Wan, Kanglin

作者机构： [Zhao, Xiuqin; Wan, Li; Lu, Bing; Yu, Qin; Lian, Lulu; Wan, Kanglin; Dong, Haiyan] Chinese Ctr Dis Control & Prevention, Natl Inst Communicable Dis Control & Prevent, State Key Lab Infect Dis Prevent & Control, Beijing, Peoples R China.;[Wu, Yimou; Yu, Qin; Lian, Lulu; Wan, Kanglin] Univ South China, Pathogen Biol Inst, Hengyang, Hunan, Peoples R China.;[Liu, Yao; Su, Yunkai; Yang, Xiaomin; Ma, Yan] Inner Mongolia Inst TB Control & Prevent, Hohhot, Peoples R China.

通讯机构： [Wan, Kanglin] C;Chinese Ctr Dis Control & Prevention, Natl Inst Communicable Dis Control & Prevent, State Key Lab Infect Dis Prevent & Control, Beijing, Peoples R China.

关键词： Mycobacterium tuberculosis;China;Tuberculosis;Genotyping;Polymerase chain reaction;Molecular epidemiology;Tandem repeats;Fungal genetics

摘要： Background: Tuberculosis (TB) is a serious public health problem in China, and within China, Inner Mongolia has a high prevalence area of TB. Though studies on the genetic diversity of Mycobacterium tuberculosis (MTB) have been reported in many provinces, there are no such studies to date in Inner Mongolia. In this study, we investigated the genetic diversity of MTB in Inner Mongolia. Methodology/Principal Findings: In this study, we analyzed 372 clinical MTB isolates with 22-loci mycobacterial interspersed repetitive unit and variable-number tandem repeats (MIRU-VNTR), spoligotyping, large sequence polymorphism (LSP), and NTF region analysis to understand the TB genotypes prevalent in Inner Mongolia. We found that the Beijing family was the most prevalent genotype (85.48%, 318/372), and the "modern'' sublineage accounted for 76.73% (244/318) of the isolates. Our data also showed that there was no statistically significant association between the two major nationalities and the Beijing genotype (chi(2) = 3.612, P = 0.057; P > 0.05). Conclusion/Significance: The Beijing genotype is the most prevalent family of M. tuberculosis in Inner Mongolia, and we do not find any correlation between the Beijing genotype and the major nationalities.

语种： 英文

作者： 陈列松;游晓星;刘良专;曾焱华;吴移谋

期刊： 当代医学,2013年(29):23-24,160 ISSN：1009-4393

作者机构： 湖南421001南华大学病原生物学研究所

关键词： 肺炎支原体;融合蛋白

摘要： 目的克隆表达与纯化肺炎支原体mpn 668蛋白。方法以Mp 129株基因组DNA为模板,PCR法扩增目的基因mpn 668,将其亚克隆至pGEX-6 P-1载体中。经鉴定后将其转化至表达菌E.coli BL 21中进行诱导表达。采用SDS-PAGE和蛋白质印迹等方法对表达产物进行分析鉴定并纯化GST融合蛋白。结果成功扩增出总长度为423 bp的mpn 668基因,所构建的原核重组质粒经PCR、双酶切以及测序鉴定与预期目的基因相符。SDS-PAGE显示,IPTG可诱导一分子量约为41 kD的可溶性GST融合蛋白,经GST·BindTMPurification Kit纯化后,纯度可达95%以上。结论本研究成功克隆表达出mpn 668融合蛋白,为下一步研究其功能奠定了基础。

语种： 中文

作者： 刘小军;蒋传好;曾铁兵;赵飞骏;余坚;...

期刊： 医学信息,2013年(14):266-267 ISSN：1006-1959

作者机构： 南华大学附属第一医院血液科，湖南 衡阳421001;南华大学医学院病原生物学研究所，湖南 衡阳421001;南华大学医学院病原生物学研究所,湖南 衡阳,421001

关键词： 梅毒螺旋体;重组抗原;抗原性

摘要： 目的表达梅毒螺旋体(Treponema pal idum, Tp)重组蛋白Tp1038(rTp1038)，鉴定其抗原性，为进一步探讨其在梅毒血清学诊断中的价值奠定基础。方法PCR扩增Tp1038全长基因，构建含6个组氨酸(His)标签的原核表达重组体pET-28a(+)/Tp1038，转化宿主菌 E. coli诱导表达 rTp1038，以 Ni-NTA亲和层析柱纯化蛋白，SDS-PAGE分析蛋白表达形式与纯度，Western blot鉴定rTp1038的抗原性。结果成功构建了表达重组体pET-28a/Tp1038，经诱导宿主菌高效表达了一分子量大约为20 KDa的重组蛋白，以可溶性表达形式为主，纯化蛋白的纯度>98%；Western blot结果显示该纯化蛋白仅能被抗 His单抗和梅毒患者混合血清所特异性识别。结论高效表达了可溶性rTp1038，该重组蛋白有良好的抗原性和特异性。

语种： 中文

作者： 陈玉玉;吴移谋;张秋桂;刘双全;黄再平

期刊： 现代预防医学,2013年(04):715-717 ISSN：1003-8507

作者机构： 1. 南华大学附属第一医院;2. 南华大学病原研究所

关键词： 呼吸道感染;革兰阴性杆菌;抗菌药物;敏感性

摘要： 目的分析某医院呼吸道感染患者的病原菌群及其对常用抗菌药物的敏感性,以指导呼吸道感染性疾病的抗菌药物的使用。方法对2009年1月～2010年12月医院感染患者的6848例痰和咽拭子标本经常规细菌培养分离到的细菌进行病原菌种类和药敏分析。结果本地区呼吸道感染患者共检出细菌2801例,检出率为40.9%,其中革兰阴性杆菌占69.0%,其次为真菌,占28.5%,革兰阳性球菌占3.5%。占前5位的依次是铜绿假单胞菌（PAE）、鲍曼不动杆菌（A-BA,）、产酸克雷伯菌（KOX）、肺炎克雷伯菌（KPN）、大肠埃希氏菌（ECO）,分别占15.9%、13.2%、7.1%、6.8%、6.2%。这5种革兰阴性杆菌对头孢呋辛、庆大霉素、复方磺胺甲噁唑、环丙沙星等的敏感性均﹤60%,而对亚胺培南、美罗培南较为敏感;革兰阳性球菌对万古霉素、米诺环素、呋喃妥因较敏感。结论医院呼吸道感染以革兰阴性杆菌为主,并存在比较严重的耐药情况,亚胺培南、美罗培南是较好的治疗药物。临床应加强进行痰细菌培养及药敏试验,合理使用抗菌药物,以早期诊断和控制呼吸道感染。

语种： 中文

作者： 朱翠明;余敏君;高顺利;曾焱华;游晓星;...

期刊： 细胞与分子免疫学杂志,2013年29(6):585-588,592 ISSN：1007-8738

通讯作者： Zhu, C.

作者机构： [朱翠明; 余敏君; 曾焱华; 游晓星; 吴移谋] 南华大学医学微生物学与免疫学教研室;[高顺利] 南华大学附属第一医院

关键词： 肺炎支原体;P1基因;基因疫苗

摘要： 目的研究肺炎支原体(Mp)P1C-IL-2融合基因疫苗经鼻饲免疫小鼠后的免疫应答水平和免疫保护作用,了解IL-2对P1C核酸疫苗的免疫佐剂效应。方法将构建的P1C-IL-2核酸疫苗鼻饲免疫BALB/c鼠,ELISA检测免疫小鼠血清IgG滴度、IgG亚类和支气管肺泡灌洗液中IgA及IFN-γ、IL-4的水平;建立小鼠Mp感染模型,观察Mp攻击后小鼠肺组织炎症情况和支气管肺泡灌洗液中Mp菌落数的变化。结果P1C-IL-2双基因疫苗组小鼠血清中的总IgG、IgGl、IgG2a亚类和支气管肺泡灌洗液中IFN-γ和IL-4水平均较P1C疫苗组小鼠显著增高(P<0.05),但两组支气管肺泡灌洗液IgA差异无显著性(P>0.05)。用Mp滴鼻感染免疫小鼠,第1、3、6天P1C-IL-2双基因融合疫苗组小鼠肺组织炎症病理评分显著高于P1C单基因疫苗免疫组小鼠,两组小鼠支气管肺泡灌洗液中的Mp菌落数差异无显著性(P>0.05)。结论IL-2能显著增强PlC疫苗的免疫应答水平,但在感染早期也激发了较强的肺组织炎症。

语种： 中文

作者： 周洲;陈丽丽;刘良专;李忠玉;游晓星;...

期刊： 免疫学杂志,2013年29(2):99-103 ISSN：1000-8861

作者机构： [周洲; 陈丽丽; 刘良专; 李忠玉; 游晓星; 吴移谋] 南华大学微生物与免疫学教研室;[陈曦] 湖南省疾病预防控制中心

关键词： 衣原体;炎症反应;白细胞介素

摘要： 目的探讨肺炎嗜衣原体(Chlamydophila pneumoniae,Cpn)热休克蛋白(heat shock protein,HSP)10(CHSP10)诱导人单核细胞分泌炎症因子的作用及Toll样受体(Toll-like receptor,TLR)2、TLR4与此作用的相关性。方法以去内毒素活性的不同质量浓度CHSP10(0.5、1、5、10、20、30μg/ml)刺激THP-1细胞0、6、12、24、36、48、60 h,检测蛋白未处理组、加热处理组、去蛋白处理组中IL-1β及IL-6水平;用CHSP10刺激C3H系野生型(C3H/HeN)和TLR4缺陷型(C3H/HeJ)小鼠腹腔巨噬细胞,分别检测IL-1β及IL-6水平;用CHSP10刺激被抗TLR2/TLR4抗体处理的THP-1细胞,检测IL-1β、IL-6的变化。结果CHSP10可诱导THP-1细胞产生炎症因子IL-1β、IL-6;CHSP10诱生C3H系野生型小鼠细胞分泌的炎症因子明显高于TLR4缺陷型小鼠细胞;用TLR2和/或TLR4抗体封闭后,CHSP10诱生的IL-1β、IL-6不同程度减少。结论CHSP10可能作为炎症相关蛋白参与了Cpn对宿主细胞的致炎作用;并且TLR2及TLR4在该炎症刺激信号的传递过程中发挥一定的作用。

语种： 中文

作者： Zhao FeiJun;Zhang XiaoHong;Liu ShuangQuan;Zeng TieBing;Yu Jian;...

期刊： 中国科学:生命科学(英文版),2013年56(2):174-180 ISSN：1674-7305

通讯作者： Wu YiMou

作者机构： [Wu YiMou; Zhao FeiJun; Zeng TieBing; Zhang YueJun] Univ S China, Sch Med, Pathogen Biol Inst, Hengyang 421001, Peoples R China.;[Zhang XiaoHong] Univ S China, Sch Med, Dept Histol & Embryol, Hengyang 421001, Peoples R China.;[Liu ShuangQuan] Univ S China, Affiliated Hosp 1, Hengyang 421001, Peoples R China.;[Yu Jian] Univ S China, Sch Med, Dept Expt Zool, Hengyang 421001, Peoples R China.;[Gu WeiMing] Skin Dis & STD Hosp, Shanghai 200050, Peoples R China.

通讯机构： [Wu YiMou] U;Univ S China, Sch Med, Pathogen Biol Inst, Hengyang 421001, Peoples R China.

关键词： Treponema pallidum (Tp);membrane protein;Gpd;CS;IL-2;nucleic acid vaccine

摘要： Syphilis is a multistage, sexually transmitted disease caused by the spirochete, Treponema pallidum (Tp). A significantly high incidence of syphilis has been reported in several countries, including China, and there is an urgent need for the development of efficacious vaccines against syphilis. DNA vaccines are a major breakthrough in the field of vaccination with several advantages over traditional vaccines. Animal model studies of Tp DNA vaccines have not been reported elsewhere but our previous reports describe the development of a single-gene Tp DNA vaccine and preclinical immunization study. In this study, chitosan (CS) nanoparticles were used as a vector and an interleukin-2 expression plasmid (pIL-2) as an adjuvant to enhance a TpGpd DNA vaccine candidate (pTpGpd) in a rabbit Tp skin challenge model. At week 8 after the first immunization, three rabbits from each group were used to determine cytokine measurements and spleen lymphocyte proliferation assay. pTpGpd in combination with pIL-2 wrapped with CS led to the greatest enhancement of anti-TpGpd antibodies and T-cell proliferation. During infection, levels of anti-TpGpd antibodies and T-cell proliferation were measured. Both the serum special IgG and IL-2, interferon-γ were significantly increased by the co-injection of the IL-2 plasmid compared with the injection of TpGpd DNA alone (P< 0.05). Furthermore, IL-2 plasmid coinjection efficiently enhanced the antigen-specific lymphocyte proliferation response. Additionally, the ratios of positive skin lesions and ulcer lesions in groups immunized with pTpGpd were significantly lower than those of the pIL-2, CS or pIL-2 mixed with CS control groups (P<0. 001). CS vectored and pIL-2 adjuvanted pTpGpd immunized animals exhibited the lowest rates of positive skin tests (8. 33%) and ulcer lesions (4. 17%) and the fastest recovery (42 d). These experiments indicate that co-injection of a pIL-2 plasmid with pTpGpd DNA vaccine wrapped with CS can significantly strengthen the long-term stability of immune response during infection, efficiently improve the protective effect against T. pallidum spirochetes infection and attenuate syphilitic lesion development. © 2013 The Author(s).

语种： 英文

作者： Li Zhong Yu;Huang Qiu Lin;Su Sheng Mei;Zhong Guang Ming;Wu Yi Mou*

期刊： 生物医学与环境科学：英文版,2013年26(3):169-175 ISSN：0895-3988

通讯作者： Wu Yi Mou

作者机构： [Su Sheng Mei; Wu Yi Mou; Li Zhong Yu] Univ S China, Sch Med, Dept Microbiol & Immunol, Hengyang 421001, Hunan, Peoples R China.;[Huang Qiu Lin] Univ S China, Dept Gen Surg, Affiliated Hosp 1, Hengyang 421001, Hunan, Peoples R China.;[Zhong Guang Ming] Univ Texas Hlth Sci Ctr San Antonio, Dept Microbiol & Immunol, San Antonio, TX 78229 USA.

通讯机构： [Wu Yi Mou] U;Univ S China, Sch Med, Dept Microbiol & Immunol, Hengyang 421001, Hunan, Peoples R China.

关键词： Chlamydia trachomatis;Monoclonal antibody;Polyclonal antibody;pORF5;DAS-ELISA

摘要： Objective To prepare antibodies against pORF5 plasmid protein of Chlamydia trachomatis and develop double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) for the detection of genital C. trachomatis infections. Methods The pORF5 protein was expressed in Escherichia coli and used to immunize BALB/c mice and New Zealand rabbits to produce monoclonal antibodies (mAbs) and polyclonal antibody (pAb) for DAS-ELISAs. Clinical samples from 186 urogenital infection patients (groups I) and 62 healthy donors (groups II) were detected in parallel by the DAS-ELISAs developed in this study and by IDEIA PCE commercial ELISA. Results Two hybridoma cell lines, named 2H4 and 4E6, stably secreting specific mAbs against pORF5 were obtained. The mAb 2H4 was recognized by 32 (17.20%, positive recognition rate) and 25 (13.44%), mAb 2H4 by 0 (0%) and 2 (3.22%) samples from groups I and II, respectively. The sensitivities of mAbs 2H4 and 4E6 were 92.11% and 77.78% and the specificities were 100% and 96.88%, respectively in relation to the IDEIA PCE commercial ELISA. The sensitivities of detection for the DAS-ELISAs were 10 ng/mL (based on 2H4) and 18 ng/mL (based on 4E6). Conclusion Two DAS-ELISAs were developed in this study that provided a feasible and effective assay that could be considered alternative tools for the serodiagnosis of C. trachomatis infection. © 2013 The Editorial Board of Biomedical and Environmental Sciences.

语种： 英文

作者： Zeng, Yanhua;You, Xiaoxing;Liu, Liangzhuan;He, Jun;Zhu, Cuiming;...

期刊： CANADIAN JOURNAL OF MICROBIOLOGY,2013年59(7):479-484 ISSN：0008-4166

通讯作者： Wu, Yimou

作者机构： [Wu, Yimou; Liu, Liangzhuan; Zhu, Cuiming; You, Xiaoxing; Yu, Minjun; Zeng, Yanhua; Ma, Xiaohua] Univ South China, Inst Pathogen Biol, Hengyang 421001, Peoples R China.;[He, Jun] Univ South China, Affiliated Nanhua Hosp, Hengyang 421001, Peoples R China.

通讯机构： [Wu, Yimou] U;Univ South China, Inst Pathogen Biol, Hengyang 421001, Peoples R China.

关键词： Mycoplasma genitalium;adhesin protein;mimic epitope;multiple antigen peptide;immunogenicity;Mycoplasma genitalium;protéine adhésine;épitope mimétique;peptide à antigènes multiples;immunogénicité

摘要： The purpose of this study was to investigate the humoral and cellular immune responses stimulated by multiple antigen peptides (MAPs) containing the mimic epitopes of Mycoplasma genitalium adhesion protein (MgPa). Three MAPs containing the mimic epitopes of MgPa were synthesized on a branched polylysine matrix. After purification and characterization, these MAPs were used to immunize BALB/c mice. The immunoglobulin G (IgG) antibody and the subtype of IgG antibody in the serum of the immunized mice were detected by indirect ELISA (enzyme-linked immunosorbent assay). The proliferation of the spleen lymphocyte was detected using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay. The gamma interferon (IFN-γ) and interleukin-4 (IL-4) levels in the cultured supernatant of spleen lymphocytes were measured by ELISA. The 3 different MAPs were prepared with high purity. Levels of IgG, IgG1, and IgG2a antibodies were elevated in the mice serum immunized by all 3 MAPs. The major antibody isotype was IgG2a. Importantly, mice immunized with a mixture of the 3 MAPs produced significantly more antibodies than those immunized with a single MAP (p < 0.05). Moreover, these MAPs could stimulate the proliferation of spleen lymphocytes of immunized mice and induce the production of IFN-γ and IL-4. The IFN-γ and IL-4 levels stimulated by the mixed MAPs were significantly higher than those stimulated by a single MAP (p < 0.01). The 3 different MAPs could induce strong cellular and humoral immune responses. The immunoreactivity of the mixed MAPs was stronger than that of the single MAP.

语种： 英文

作者： Liu Zhi Yun;Hao Qin;Hou Xue Xia;Jiang Yi;Geng Zhen;...

期刊： 生物医学与环境科学：英文版,2013年26(3):190-200 ISSN：0895-3988

通讯作者： Wan Kang Lin

作者机构： [Liu Zhi Yun; Wan Kang Lin; Hou Xue Xia; Jiang Yi; Hao Qin; Geng Zhen] Chinese Ctr Dis Control & Prevent, Natl Inst Communicable Dis Control & Prevent, State Key Lab Infect Dis Prevent & Control, Beijing 102206, Peoples R China.;[Liu Zhi Yun; Wu Yi Mou] Univ S China, Inst Pathogen Biol, Hengyang 421001, Hunan, Peoples R China.;[Liu Zhi Yun] YuShui Dist Ctr Dis Control & Prevent, Xinyu 338000, Jiangxi, Peoples R China.

通讯机构： [Wan Kang Lin] C;Chinese Ctr Dis Control & Prevent, Natl Inst Communicable Dis Control & Prevent, State Key Lab Infect Dis Prevent & Control, Beijing 102206, Peoples R China.

关键词： Lyme disease;Western blot;Diagnostic method;Borrelia afzelii

摘要： Objective To study the technique of Western blot for the diagnosis of Lyme disease caused by Borrelia afzelii in China and to establish the standard criteria by operational procedure. Methods FP1, which is the representative strain of B. afzelii in China, was analyzed by SDS-PAGE, electro transfer and immunoblotting assays. The molecular weights of the protein bands of FP1 were analyzed by Gel-Pro analysis software. In a study using 451 serum samples (159 patients with Lyme disease and 292 controls), all observed bands were recorded. The accuracy of the WB as a diagnostic test was established by using the ROC curve and Youden index. Results Criteria for a positive diagnosis of Lyme disease were established as at least one band of P83/100, P58, P39, OspB, OspA, P30, P28, OspC, P17, and P14 in the IgG test and at least one band of P83/100, P58, P39, OspA, P30, P28, OspC, P17, and P41 in the IgM test. For IgG criteria, the sensitivity, specificity and Youden index were 69.8%, 98.3%, and 0.681, respectively; for IgM criteria, the sensitivity, specificity and Youden index were 47%, 94.2%, and 0.412, respectively. Conclusion Establishment of WB criteria for B. afzelii is important in validating the diagnostic assays for Lyme disease in China. © 2013 The Editorial Board of Biomedical and Environmental Sciences.

语种： 英文

作者： 黎知青;刘良专;伍海英;彭菁;陈丽丽;...

期刊： 中华微生物学和免疫学杂志,2013年33(12):933-937 ISSN：0254-5101

通讯作者： Wu, Y.-M.

作者机构： [黎知青; 刘良专; 贺庆芝; 彭菁; 吴移谋; 陈丽丽; 伍海英] Institute of Pathogenic Biology, University of South China, Hengyang 421001, China

通讯机构： [Wu, Y.-M.] I;Institute of Pathogenic Biology, University of South China, China

关键词： 鹦鹉热嗜衣原体;细胞定位;表达时相;促炎症因子

摘要： 目的:研究鹦鹉热嗜衣原体( Chlamydophila psittaci,Cps) CPSIT_0271蛋白的定位和表达时相,及其重组蛋白对人单核细胞( THP-1)产生促炎症因子的诱生作用。方法原核表达、纯化GST-CPSIT_0271重组蛋白,免疫BALB/c小鼠制备多克隆抗体,ELISA分析血清抗体滴度。间接免疫荧光法初步观察内源性CPSIT_0271的定位,并通过Western blot 分析CPSIT_0271在Cps感染HeLa细胞8 h、18 h、24 h、36 h和48 h后的表达情况。用不同浓度GST-CPSIT_0271刺激THP-1细胞,ELISA检测IL-6、IL-1β和TNF-α水平。结果在Cps感染的HeLa细胞中,CPSIT_0271分布于衣原体包涵体内。 CPSIT_0271在Cps感染HeLa细胞后36 h开始表达,在48 h时表达量增加。 GST-CPSIT_0271免疫小鼠,产生较强的免疫应答,小鼠血清特异性抗体效价为1∶16000。当GST-CPSIT_0271蛋白浓度为2~5μg/ml时,刺激THP-1细胞产生IL-6、IL-1β和TNF-α的水平与CPSIT_0271呈明显的剂量依赖关系。5μg/ml的GST-CPSIT_0271蛋白刺激THP-1细胞,TNF-α和IL-1β在24 h时达到高峰,IL-6在48 h时达到高峰。结论 CPSIT_0271蛋白分布于包涵体内,CPSIT_0271基因可能为Cps晚期表达基因;GST-CPSIT_0271具有较好的免疫原性,并能促进THP-1细胞分泌促炎症因子IL-6、TNF-α和IL-1β。

语种： 中文

作者： Liu, Liangzhuan;You, Xiaoxing;Chen, Liesong;Zeng, Yanhua;Tang, Shuangyang;...

期刊： MOLECULAR MEDICINE REPORTS,2012年6(6):1239-1242 ISSN：1791-2997

通讯作者： Wu, Yimou

作者机构： [Wu, Yimou; Liu, Liangzhuan; Chen, Liesong; You, Xiaoxing; Tang, Shuangyang; Yu, Minjun; Zeng, Yanhua] Univ S China, Pathogen Biol Inst, Hengyang 421001, Peoples R China.;[Xhen, Xi] Hunan Prov Ctr Dis Prevent & Control, Changsha 410005, Hunan, Peoples R China.

通讯机构： [Wu, Yimou] U;Univ S China, Pathogen Biol Inst, Hengyang 421001, Peoples R China.

关键词： Chlamydophila pneumoniae;Cpn0425;cell localization;immunogenicity

摘要： The present study aimed to determine the intracellular localization of Cpn0425 in Chlamydophila pneumoniae-infected cells. The recombinant plasmid pGEX-6p/Cpn0425 was transformed into E.coli Bl21 cells to express the fusion protein. Following purification with glutathione S-transferase (GST) resin chromatography, the Cpn0425 fusion protein was used to induce immunity in mice to develop monoclonal and polyclonal antibodies, which were subsequently used to localize the endogenous Cpn0425 protein by indirect immunofluorescence assay (IFA). ELISA was used to determine the immunogenicity of the Cpn0425 plasmid protein by recognizing the pool sera of patients infected with Chlamydia trachomatis and the pool sera of mice immunized with the Cpn0425 fusion protein. The Cpn0425 gene was expressed as the GST-Cpn0425 fusion protein in E. coli and its antibody was prepared by immunizing mice with the fusion protein. An anti-GST-Cpn0425 antibody was used to localize the protein in cells infected with Chlamydophila pneumoniae AR-39 using an IFA. The anti-GST-CT058 antibody detected an inclusion signal in the IFA. Cpn0425 protein strongly reacted with antiserum. Although Cpn0425 protein is not a secreted protein, it has good immunogenicity. Therefore, this protein may be useful for developing vaccines against Chlamydophila pneumoniae infection.

语种： 英文