2015─2017年新疆青河县山区动物鼠疫血清学和病原学调查

目的通过鼠疫病原学和血清学的实验检测结果,了解新疆青河县山区存在鼠疫自然疫源地的潜在状况。方法采用鼠疫"四步检验"法分离青河县山区捕获的长尾黄鼠及灰旱獭脏器、自毙动物脏器及骨骼和寄生蚤中的鼠疫菌;间接血凝试验(HA)和酶联免疫吸附试验(ELISA)检测动物脏器和寄生蚤悬液中的鼠疫F1抗原;长尾黄鼠、灰旱獭以及牧犬血清中的鼠疫F1抗体。结果经鼠疫"四步检验"和反向间接血凝(RIHA)检验长尾黄鼠脏器552份、灰旱獭脏器4份、自毙长尾黄鼠和灰旱獭脏器及骨骼13份、长尾黄鼠体外寄生蚤583只,均未检出鼠疫菌及F1抗原;检测长尾黄鼠血清552份,IHA法检出阳性1份、滴度1∶2~9,ELISA法检出阳性3份、平均滴度1∶2~8;检测牧犬血清129份,IHA法检出阳性6份、平均滴度1∶2^(3.83),ELISA法检出阳性8份、平均滴度1:26.75;其中牧犬鼠疫F1抗体阳性标本中检出鼠疫IgM抗体阳性2份,平均滴度1∶2^(5.5)。结论新疆青河县山区啮齿类动物间有鼠疫流行迹象,流行区域为花海子地区,存在潜在鼠疫自然疫源地,建议对该区域开展进一步鼠疫疫源性调查。     

胶体金免疫层析法与双单克隆抗体夹心ELISA检测鼠疫F1抗原的比较

目的比较胶体金免疫层析法（GICA）和双单克隆抗体夹心酶联免疫吸附试验（DMcAbS-ELISA）检测鼠疫F1抗原的敏感性和特异性。方法用GICA试纸条和DMcAbS-ELISA平行检测308份强毒鼠疫耶尔森菌感染鼠脏器标本和327份对照鼠标本,用RIHA微量法同时检测作为参照。结果327份对照鼠鼠疫细菌学检验及GICA,DMcAbS-ELISA和RIHA的F1抗原检测均为阴性,GICA,DMcAbS-ELISA和RIHA在108cfu/mL水平上未发现与近缘假结核耶尔森菌有交叉反应;308只感染鼠样本细菌培养阳性284只,24只样本未分离到鼠疫菌;F1抗原检测GICA阳性287份,DM-cAbS-ELISA阳性280份,RIHA阳性276份,GICA阳性检出率最高93.19%,与DMcAbS-ELISA和RIHA比较,差异均有统计学意义（χ^2=5.14,9.09;P=0.016,0.001）;GICA与DMcAbS-ELISA符合率97.73%（Kappa=0.845）;GICA和DMcAbS-ELISA与RIHA符合率分别是96.43%和98.70%（Kappa=0.774,0.926）;284份细菌培养阳性鼠标本F1抗原检测GICA阳性率是98.59%,高于DMcAbS-ELISA和RIHA,差异有统计学意义（χ^2=4.17,5.14;P=0.031,0.016）;24份细菌培养阴性实验鼠标本F1抗原检测：GICA检出7份阳性,阳性率29.17%,DMcAbS-ELISA检出6份阳性,阳性率25%,RIHA检出3份阳性,阳性率12.5%,GICA高于DMcAbS-ELISA和RIHA,但差异无统计学意义（χ^2=2.25,0;P=0.125,1.000）。结论GICA检测鼠疫F1抗原敏感特异,快速简便,优于DMcAbS-ELISA和RIHA,是鼠疫快速诊断中有应用价值的检测技术。     

河北省鼠疫自然疫源地蒙系绵羊血清鼠疫F1抗体调查

目的调查河北省鼠疫疫源地绵羊鼠疫F1抗体的分布水平,探讨绵羊在鼠疫监测过程的意义。方法在河北历史人间鼠疫发生地和动物间鼠疫活动疫源地的1个镇1个行政村进行随机抽样,共采集绵羊血样394份。同时进行常规鼠疫间接血凝试验、鼠疫F1抗体胶体金试验和双抗原夹心ELISA检测F1抗体试验。结果鼠疫间接血凝试验阳性1份;双抗原夹心ELISA测F1抗体试验阳性1份;鼠疫F1抗体胶体金试验阳性2份。结论在河北省鼠疫自然疫源地有一定数量的鼠疫F1抗体阳性绵羊,具体原因尚不清楚,有待进一步调查和研究。     

四川省德格县旱獭鼠疫流行病学调查

目的证实喜马拉雅旱獭(旱獭)鼠疫在四川是否存在,为鼠疫防治提供依据。方法应用现场流行病学调查和实验室检测相结合。结果通过鼠疫间接血凝试验(IHA)检出犬阳性血清2份,从33份旱獭标本中检出鼠疫F1抗原阳性材料10份,17份旱獭检材中检获鼠疫菌4株。结论确定四川德格县为喜马拉雅旱獭鼠疫自然疫源地。     

腹泻患者中新型志贺菌(福氏4X)的检出

2005年6月份,在腹泻病人的粪便中检出1株福氏4型志贺菌,群抗原只有福氏菌群7,经多次传代和回复试验,均未能检出其它抗原,该抗原结构为首次发现。现将结果报告如下。     

鼠疫菌F1抗原与假结核菌和钩端螺旋体抗原的交叉性研究

鼠疫菌F_1抗原作为特异性抗原已广泛应用于鼠疫诊断中,WHO推荐的F_1抗原IHA和RIP试验,已作为常规诊断方法,从1973年我们用IHA方法在雷州半岛历史鼠疫区反复从鼠类血清中检出阳性,RIP方法检出2份低滴度阳性血清,但始终没检出鼠疫菌,为了排除F_1抗原与其它细菌有无相关抗原与相应抗体存在交叉反应,我们从1984年～1992年用IHA、RIP、MAT做了多次研究,现将结果报告如下。     

PCR检测鼠疫菌种中F1抗原基因

目的应用PCR检测试验用EV菌株是否存在caf1基因。方法应用含内部对照PCR技术,以鼠疫菌特异基因caf1合成一对引物,进行PCR实验。结果该试验用EV菌株经PCR扩增,呈现一条与caf1基因分子量相吻合的清晰目的扩增带。结论该试验用EV菌株未缺失产生F1抗原的基因,可以用于提取F1抗原等试验。     

元江县一起鼠疫暴发流行的调查分析

1992年1～3月,云南省元江县鼠疫自然疫源地间歇69年后在县城爆发了人类鼠疫流行。经病原学和血清学调查,确诊腺鼠疫患者9人,从黄胸鼠体内分离到鼠疫菌4株,查出鼠类脏器、骨髓各1份为鼠疫F1抗原阳性,检出11只褐家鼠为F1抗体阳性。     

胶体金免疫层析法快速诊断鼠疫的实验研究

目的建立一种快速、简易的检测鼠疫F1抗原的胶体金免疫层析法(GICA).方法采用直径为20nm胶体金颗粒,标记F1单克隆抗体,并将标记物以6μL/cm喷于玻璃纤维;将另1株F1单克隆抗体与羊抗鼠IgG以1μL/cm喷线固定于硝酸纤维素膜上,装配后以4mm/条切割制成免疫层析检测试条.分别用粗制F1抗原和EV菌对该试纸条的敏感性进行测定,同时用44株鼠疫近缘菌及昆明小鼠、黄胸鼠、褐家鼠、大足鼠的正常血清等标本进行特异性检测.结果本试纸条可在15min之内完成实验,检测粗制F1抗原的浓度可达0.5ng/mL,最小检出EV菌菌量为10万个;本方法的特异性为100%.结论 GICA检测鼠疫F1抗原特异性强、灵敏度高、简便快速,无需特殊仪器设备,有广泛应用价值.     

鼠疫F1单克隆抗体的胶体金标记及斑点渗滤法检测F1抗原方法的建立

目的建立斑点渗滤法快速检测鼠疫F1抗原的方法。方法粗提鼠疫F1单克隆抗体,在pH8.2下对其进行胶体金标记,以兔抗F1抗体与金标记的单克隆抗体建立斑点渗滤快速检测鼠疫F1抗原的方法,并对该方法进行特异性与敏感性试验。结果成功建立斑点渗滤法快速检测鼠疫F1抗原的方法,该检测可在15min内完成,敏感性为3.12mg/L,对假结核、小肠结肠炎等近缘菌的特异性实验未发现交叉反应。结论该方法可作为鼠疫快速诊断的手段,但实验敏感性有待进一步提高。     

Timely immunization subverts the development of peripheral nonresponsiveness and suppresses tumor development in simian virus 40 tumor antigen-transgenic mice.

Tolerance to tumor cell-expressed molecules and selection of cells that evade immune surveillance during tumor progression create effective barriers to immunotherapy. We investigated the cytotoxic T-lymphocyte response to simian virus 40 (SV40) tumor (T/t) antigen in two lineages of transgenic mice bearing the same rat insulin promoter-SV40 T/t antigen (RIP Tag) hybrid gene. RIP1-Tag2 mice, which express Tag as embryos, are tolerant to Tag, whereas RIP1-Tag4 mice, which express the transgene in pancreatic islet beta cells several weeks after birth and develop insulinomas, can be immunized to generate active Tag-specific cytotoxic T lymphocytes as determined by in vitro assays. Indeed, RIP1-Tag4 mice immunized with Tag by SV40 infection prior to the time of endogenous transgene expression also mount an effective in vivo cellular immune response to the Tag-expressing pancreatic beta cells, and Tag-induced tumor growth is significantly delayed (up to 1 year). However, after the transgene is expressed, RIP1-Tag4 mice are unable to mount a tumor-inhibiting response upon immunization, although Tag-specific cytotoxic T cells can still be demonstrated in vitro. Our data suggest that Tag-specific T cells are rendered unresponsive in vivo in RIP1-Tag4 mice and that the establishment of this unresponsiveness to Tag can be prevented by SV40 immunization only before the onset of the transgene expression. In the older, successfully immunized mouse, decreased immune surveillance and selection of cells with down-regulation of major histocompatibility complex class I expression most likely set the stage for insulinoma development.     

放射免疫沉淀试验检测鼠疫FI抗体的研究——Ⅰ.特异性与敏感性

试验证明,用放射免疫沉淀试验(RIP)检测鼠疫抗体时,将被试血清与相应的阴性对照血清的结合比定为≥3作为反应标准是适宜的;用RIP检测假结核,小肠结肠炎菌等抗血清30份,这些抗血清在1:10到1:100的各种稀释度中与~(125)I-鼠疫FI抗原均无交叉反应,从非鼠疫疫区收集的2196份哺乳动物血清,亦未发现阳性反应,说明该法特异性强。由于RIP是用分离剂沉淀鼠疫抗原-抗体复合物,故能检出不完全抗体,检出率不仅高于常规的间接血球凝集试验(IHA)和酶联免疫吸附试验(ELISA),且滴度往往高十几倍、几十倍,乃至几百倍。所以用RIP检测潜在性鼠疫疫源地、考核灭源效果以及在科研中用于检测微量鼠疫抗体具有重要意义。加之该法自动化程度高,测试信息以数字显示,因而指标客观,容易分析,适于大规模现场调查。     

鼠疫放射免疫沉淀试验在鼠疫监测中的应用及效果评价

目的：建立敏感性高、特异性强的鼠疫血清学诊断方法，应用于鼠疫血清学监测。方法：用标记的鼠疫菌FI抗原（^123I-FI）检测阳性血清、免疫黄鼠血清、实验区各种动物血清及正常血清。结果:建立鼠疫放射免疫沉淀试验（RIP），并确定其诊断标准，研制出了鼠疫抗体放免测定盒及其使用方法。RIP在鼠疫监测中的应用。结论：RIP的敏感性和特异性均明显高于IHA，是目前建立的鼠疫血清学方法中最为敏感特异的方法。     

Antibodies to recombinant-derived glycoprotein B after natural human cytomegalovirus infection correlate with neutralizing activity.

Glycoprotein B (gB) of human cytomegalovirus (HCMV) was partially purified by lentil-lectin column chromatography from cells infected with an adenovirus-gB recombinant. This antigen, which contained specifically reactive proteins of approximately 130 and 55 kDa, was used to investigate gB antibody levels after natural HCMV infection in 48 individuals. All sera had IgG antibody to gB as detected by radioimmunoprecipitation (RIP) assays. Quantitative RIP showed a strong correlation between gB antibody and neutralizing activity (r = .74, P less than .001) but a weak correlation between gB antibody and total HCMV-specific IgG (r = .36, P less than .02). When gB antibody was specifically absorbed from 20 serum specimens, neutralizing antibody titer was reduced a median of 48% (range, 0-98%). These data confirmed that antibodies to gB are a large component of the neutralizing antibody response to HCMV and support a role for this protein in the development of subunit vaccines.     

基于RIP1/RIP3/NLRP3信号通路探讨祛瘀生新方治疗溃疡性结肠炎小鼠的作用机制

目的:研究祛瘀生新方对溃疡性结肠炎(UC)小鼠的治疗效果以及对RIP1/RIP3/NLRP3信号通路的调节作用.方法:将36只小鼠随机分为实验组(祛瘀生新组)、模型组、阳性对照组(美莎拉嗪组)、空白组、中药拆方1组(生新组)、中药拆方2组(祛瘀组),6只/组,采用3.5％葡聚糖硫酸钠盐(DSS)溶液自由饮水法制备UC小...     

Human thymus antigen: Characterization and its expression on human leukemias

Specific anti-human thymus xenoantiserum (ATS) was utilized for characterizing a human thymus antigen (HTA) preferentially expressed on human thymocytes. Binding of ATS with different cell types was studied by immunofluorescence and immunoperoxidase techniques, as well as by radioimmunoprecipitation (RIP) followed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). ATS reacted selectively with the majority (87%) of human thymocytes among normal lymphoid cell populations tested. In the thymus, HTA was expressed largely on cortical thymocytes but not on those cells located in the thymic medulla. When 20 cases of different types of lymphatic leukemias were studied with ATS, HTA was found on acute lymphoblastic leukemia cells rosetting with sheep erythrocytes (T-ALL) but not on other lymphatic leukemia cases, including chronic lymphocytic leukemia cells rosetting with sheep red blood cells (T-CLL). SDS-PAGE analysis of immunoprecipitates formed between ATS and radiolabeled cell-surface components of human thymocytes demonstrated that ATS bound a 48,000-molecular-weight component that could be adsorbed to Sepharose 4B coupled with Lens culinaris hemagglutinin and could be labeled by periodate-tritiated borohydride, indicating that HTA is a sialoglycoprotein. Similar antigen molecules were also precipitated by ATS from T-ALL cells but not from other lymphatic leukemia cells. Five T-ALL-derived cell lines were tested with ATS by immunofluorescence and by RIP and SDS-PAGE, but only two cell lines (MOLT-3 and P12/Ich) possessed HTA molecules on their cell membrane.     

The design of RIP belts impacts the reliability and quality of the measured respiratory signals

Purpose

Evaluate the effect of respiratory inductance plethysmography (RIP) belt design on the reliability and quality of respiratory signals. A comparison of cannula flow to disposable cut-to-fit, semi-disposable folding and disposable RIP belts was performed in clinical home sleep apnea testing (HSAT) studies.

Methods

This was a retrospective study using clinical HSAT studies. The signal reliability of cannula, thorax, and abdomen RIP belts was determined by automatically identifying periods during which the signals did not represent respiratory airflow and breathing movements. Results were verified by manual scoring. RIP flow quality was determined by examining the correlation between the RIP flow and cannula flow when both signals were considered reliable.

Results

Of 767 clinical HSAT studies, mean signal reliability of the cut-to-fit, semi-disposable, and disposable thorax RIP belts was 83.0?±?26.2%, 76.1?±?24.4%, and 98.5?±?9.3%, respectively. The signal reliability of the cannula was 92.5?±?16.1%, 87.0?±?23.3%, and 85.5?±?24.5%, respectively. The automatic assessment of signal reliability for the RIP belts and cannula flow had a sensitivity of 50% and a specificity of 99% compared with manual assessment. The mean correlation of cannula flow to RIP flow from the cut-to-fit, semi-disposable, and disposable RIP belts was 0.79?±?0.24, 0.52?±?0.20, and 0.86?±?0.18, respectively.

Conclusion

The design of RIP belts affects the reliability and quality of respiratory signals. The disposable RIP belts that had integrated contacts and did not fold on top of themselves performed the best. The cut-to-fit RIP belts were most likely to be unreliable, and the semi-disposable folding belts produced the lowest-quality RIP flow signals compared to the cannula flow signal.

     

Necrostatin-1 Protects Against Paraquat-Induced Cardiac Contractile Dysfunction via RIP1-RIP3-MLKL-Dependent Necroptosis Pathway

Paraquat is a highly toxic prooxidant that triggers oxidative stress and multi-organ failure including that of the heart. To date, effective treatment of paraquat toxicity is still not established. Necroptosis, a newly discovered form of programmed cell death, was recently shown to be strongly associated with cardiovascular disease. Receptor interaction proteins 1 (RIP1), receptor interaction proteins 3 (RIP3), and mixed lineage kinase domain like (MLKL) are key proteins in the necroptosis pathway. Necrostatin-1 (Nec-1) is a specific inhibitor of necroptosis which acts by blocking the interaction between RIP1 and RIP3. In the present study, we studied the effect of Nec-1 on paraquat-induced cardiac contractile dysfunction and reactive oxygen species (ROS) production in the heart tissues using a mouse model. Our results revealed impaired contractile function, deranged intracellular Ca²⁺ handling and echocardiographic abnormalities in mice challenged with paraquat. We further found enhanced expressions of RIP1, RIP3, and MLKL along with overproduction of ROS in mice heart tissues. Nec-1 pre-treatment prevented cardiac contractile dysfunction in paraquat-challenged mice. Furthermore, Nec-1 reduced RIP1–RIP3 interaction, down-regulated the RIP1–RIP3–MLKL signal pathway, and dramatically inhibited the production of ROS. Collectively, these findings suggest that Nec-1 alleviated paraquat-induced myocardial contractile dysfunction through inhibition of necroptosis, an effect which was likely mediated via the RIP1–RIP3–MLKL signaling cascade. Further, ROS appeared to play an important role in this process. Thus, this process may represent a novel therapeutic strategy for the treatment of paraquat-induced cardiac contractile dysfunction.     

NOD1/RIP2信号通路对巨噬细胞炎性活化的作用

目的以人单核细胞株THP-1为基础,探讨NOD1/受体相互作用蛋白2(RIP2)信号通路对巨噬细胞炎性活化及表型的影响。方法用160 nmo L/L的佛波酯(PMA)将THP-1单核细胞诱导分化成巨噬细胞,分别用10、25和50 mg/L浓度的氧化型低密度脂蛋白(ox-LDL)刺激THP-1源性巨噬细胞24 h,以空白组为对照组。应用RT-PCR法检测NOD1和RIP2的mRNA表达情况;应用Western blot法检测NOD1和RIP2的蛋白表达情况;应用ELISA法检测细胞培养液中单核细胞趋化蛋白1(MCP-1)和巨噬细胞移动抑制因子(MIF)的分泌;应用流式细胞学检测巨噬细胞表面抗原CD16、CD68表达。结果 ox-LDL能以剂量依赖的方式激活THP-1源性巨噬细胞中NOD1/RIP2信号通路,随着ox-LDL刺激浓度的增加,NOD1、RIP2的mRNA和蛋白表达水平升高(P0.05)。NOD1/RIP2信号通路激活后能使细胞培养物上清液中炎症因子MIF和MCP-1的表达增加,随着ox-LDL刺激浓度的增加,MCP-1和MIF的分泌增多(P0.05)。NOD1/RIP2信号通路激活后能改变巨噬细胞表面抗原CD16、CD68的表达,随着ox-LDL刺激浓度的变化,巨噬细胞表面抗原CD16/CD68的平均荧光强度发生变化,其中50 mg/L组能显著下调CD16/CD68的表达(P0.01)。结论巨噬细胞中NOD1/RIP2信号通路能被ox-LDL以剂量依赖的方式激活,NOD1/RIP2信号通路激活后能导致巨噬细胞的炎性活化及其表型变化,这可能是其参与动脉粥样硬化形成和发展过程的主要机制。     

Detection of inspiratory flow limitation during sleep by computer assisted respiratory inductive plethysmography.

The potential of respiratory inductive plethysmography (RIP) to detect inspiratory flow limitation during sleep was investigated. Sixteen sleep apnoea patients underwent polysomnography. Airflow by a flowmeter attached to a nasal mask, oesophageal and mask pressure were recorded along with calibrated RIP. Presence of inspiratory flow limitation was defined by constant or decreasing flow without pressure dependence throughout significant portions of inspiration, its absence by a linear or mildly alinear pressure:airflow relationship. Based on this standard, three of various computerized RIP derived parameters, with highest performance to detect flow limitation, were identified. They were combined to an inspiratory flow limitation, (IFL)-Index(RIP), which was validated prospectively in another 10 sleep apnoea patients. RIP derived fractional inspiratory time, peak to mean inspiratory flow ratio, and ribcage contribution to tidal volume had the highest accuracy to detect flow limitation (area under the receiver operating characteristic (ROC) curves 0.81, 0.76, 0.76, respectively, 160 comparisons). Prospective validation revealed an area under the ROC curve for the IFL-Index(RIP) to detect flow limitation of 0.89 (95% confidence interval 0.85 to 0.93, 200 comparisons) with sensitivity and specificity at the point of equality of 80%. It is concluded that inspiratory flow limitation may be assessed by computer assisted analysis of respiratory inductive plethysmography derived breathing patterns with clinically acceptable accuracy.