应用巢式RT-PCR扩增HA和NA基因片段并测序鉴定甲型H1N1流感病毒

目的 建立一种利用巢式RT-PCR特异扩增HA和NA基因片段并测序鉴定甲型H1N1流感病毒的技术.方法 设计两套共7条特异引物,通过巢式RT-PCR分别扩增甲型H1N1流感病毒HA和NA基因片段并测序,所得序列与人感染甲型流感病毒主要HA和NA亚型序列进行进化树分析以对结果作进一步鉴定,蛋白序列比对后分析其特征.结果 4例甲型H1N1流感患者流感病毒HA和NA基因RT-PCR扩增均分别得到442 bp和543 bp片段产物.核苷酸序列进化分析表明,该4例患者HA和NA序列分别与2009年爆发的甲型H1N1流感病毒HA及NA序列聚集在一起,与季节性H1、H2、H3、人禽流感H5亚型及季节性N1、N2、人禽流感N1亚型特异分开.蛋白序列分析表明,4例患者流感病毒HA蛋白裂解位点附近氨基酸序列均为PSIQSR↓GLF,不具有高致病性流感病毒的特性,NA蛋白第275位氨基酸为His,未出现H275Y的耐药变异.结论 本方法能特异扩增甲型H1N1流感病毒HA和NA基因片段,测序后可用于甲型H1N1流感病毒的进一步鉴定;同时,得到的序列也可用于流感病毒致病力及耐药性的分析.     

河北省2009-2014年甲型H1N1流感流行特征分析

目的 分析河北省2009-2014年度甲型H1N1流感的流行特征.方法 采用描述性流行病学方法对2009年4月1日-2014年3月31日的流感监测结果进行分析.结果 采集的47949例流感样病例(influenza-like illness,ILI)标本经流感病毒核酸检测,8248例阳性(阳性率为17.20％),其中甲型H1N1 4168例,季节性H3 2006例,B型1820例,季节性H1等254例.各年度优势病原不同,除2010-2011年度外,其他年度均存在甲型H1N1流感病毒流行.对4168例甲型H1N1病例进一步分析显示:全省11个地市均有该病例检出,男女性别比1.09:1.自2009年甲流大流行后,2011年11月至2014年3月共出现3次流行,具有明显的季节性,且呈单峰分布.结论 河北省甲型N1N1流感的流行趋势已趋于季节性,无明显的地域性,除2010-2011年度外,其他年度都是优势病原之一,近年来感染以25-59岁年龄组为主.     

中国首例甲型H1N1流感二代病例的临床特征

目的了解2009年我国首例甲型H1N1流感二代病例的流行病学、临床、病原学检查特点及预后转归。方法对患者流行病学及临床资料进行回顾性分析,并采用实时荧光聚合酶链反应测定甲型H1N1流感病毒核酸。结果患者与甲型H1N1流感输入病例接触1天后发病。以发热、咽痛、咳嗽起病,白细胞及CD4＋T淋巴细胞计数降低,无肺炎等并发症。多级机构检测咽拭子甲型H1N1流感病毒核酸阳性确诊甲型H1N1流感。RT-PCR测序证实其病毒核苷酸序列与一代输入病例的一致,同源性为100%。经奥司他韦抗病毒及对症治疗痊愈出院。结论本病例的传染源明确,为我国首例报告的甲型H1N1流感二代确诊病例,其临床表现轻,病情恢复快。未发生院内感染,早隔离早诊断等防控措施有效。     

H1N1患者心肌标志物及常规生化检测的临床意义

目的:探讨心肌标志物及部分生化项目检测对于H1N1甲型流行性感冒患者(简称"甲流")的临床意义.方法:对54例H1N1甲流患者(包括4例重症甲流死亡患者)和50例正常对照者用电发光免疫分析、全自动临床化学分析检测H1N1甲流患者心肌标志物及常规生化项目.结果:与正常对照组相比,甲型H1N1感染者在治疗前血清肌酸激酶(C...     

2011-2017年河北省甲型H1N1流感流行特征分析

目的掌握河北省甲型H1N1流感的流行特征,为流感防控提供科学依据.方法通过中国流感监测信息系统收集河北省2011年4月至2017年3月流感及流感样病例(influenza-likeillness,ILI)监测数据进行统计分析.结果共检测ILI标本77 008份,甲型H1N1流感病毒核酸阳性2 699份,阳性率3.50％.对2 699例甲型H1N1流感病例进一步分析显示:全省11个地市均有该病例检出,男女比例为1.09:1,各年龄组均可发病,检出率最高的是25～59岁组,最低0～4岁组.2011年11月至2017年3月共出现4次流行,具有明显的季节性,且呈单峰性.各年度均有甲型H1N1流感病例检出,甲型H1N1流感分别是2012-2013、2013-2014和2016-2017年度的优势病原.结论河北省甲型N1N1流感流行具有明显的季节性,冬春季流行,近年来感染以5～14岁年龄组为主.     

2010年广州市越秀区居民新甲型H1N1流感病毒血清抗体调查

目的了解广州市越秀区居民各年龄段人群新甲型H1N1流感病毒抗体水平,为完善新甲型H1N1流感疫情防控措施提供科学依据。方法采取多阶段分层随机抽样方法抽取研究对象309人,进行血清标本采集和问卷调查,应用红细胞血凝抑制（HI）方法检测新甲型H1N1流感病毒抗体。结果人群新甲型H1N1流感病毒抗体阳性率为20.06%,0～岁组、6～岁组、10～岁组、16～岁组、25～岁组和60岁以上组的新甲型H1N1流感病毒抗体阳性率分别为18.84%、33.33%、46.67%、30.00%、6.67%和5.00%。6～24岁组的新甲型H1N1流感病毒抗体阳性率较高,为35.00%,无症状抗体阳性率达10.43%。抗体阳性率在性别上差异无统计学意义（P〉0.05）。结论广州市越秀区居民新甲型H1N1流感病毒感染状况各年龄段不同,青少年抗体阳性率较高,老年人抗体阳性率相对较低。     

江门市居民甲型H1N1流感病毒血清抗体的调查分析

目的了解江门市各年龄段人群甲型H1N1流感病毒抗体水平,为完善甲型H1N1流感疫情防控措施提供科学依据。方法采取多阶段分层随机抽样方法抽取研究对象480人,进行血清标本采集和问卷调查,应用红细胞血凝抑制方法检测甲型H1N1流感病毒抗体。结果人群甲型H1N1流感病毒抗体阳性率为19.38%,0～、6～、16～、25～和60岁以上组的甲型H1N1流感病毒抗体阳性率分别为22.09%、27.36%、24.47%、14.58%和8.16%。6～24岁的甲型H1N1流感病毒抗体阳性率较高,为51.83%,无症状抗体阳性率达12.61%。抗体阳性率在不同年龄、性别和职业上差异均有统计学意义（P〈0.05）。结论江门市甲型H1N1流感病毒感染状况各年龄段不同,青少年抗体阳性率较高,老年人抗体阳性率相对较低,男性抗体阳性率高于女性。     

奥司他韦对新型甲型H1N1流感轻症病例治疗效果的回顾性分析

目的 通过回顾性分析系统评价奥司他韦(达菲)对新型甲型H1N1流感轻症病例的临床治疗效果.方法 收集2009年5-8月国内三家传染病医院的308例甲型H1N1流感住院患者的临床资料,了解甲型H1N1流感的年龄分布特征及主要临床特征,之后剔除不符合要求的研究对象,将289例甲型H1N1流感轻症患者分为对照组和达菲组,了解两组病例在发热持续时间以及病毒持续时间的差异.结果 甲型H1N1流感患者的发病年龄主要集中在10～29岁.发热是甲型H1N1流感的最主要症状,以低、中度发热(＜39℃)为主,咽喉疼痛、咳嗽、头痛、流涕也是常见症状,但呕吐及腹泻患者比例较少,分别为1.3％和3.9％.最后对照组和达菲组在发病-病毒转阴时间和治疗-病毒转阴时间上的差异均无统计学意义(P =0.32,0.93).结论 甲型H1N1流感发病以青少年为主,多数患者病情较轻,预后良好.对于甲型H1N1流感轻症患者而言,达菲并不能有效缩短发热持续时间及病毒持续时间,流感病毒可以迅速被机体自身免疫系统清除,对轻症患者使用达菲抗病毒治疗并无临床意义.     

2009甲型H1N1流感大流行期间北京儿童的流感监测

目的 了解2009年甲型H1N1流感大流行期间北京地区儿童中流感流行的情况.方法 采用WHO推荐的实时荧光定量RT-PCR和国家流感中心推荐的分型方法,对2009年甲型H1N1流感大流行期间因流感样症状来首都儿科研究所附属儿童医院就诊患儿的咽拭子标本进行流感病毒核酸检测.结果 2009年6月1日至2010年2月28日期间共检测了4363份咽拭子标本,其中623例为甲型H1N1阳性,阳性率为14.3%,657例为其他甲型流感病毒阳性(15.1%),所有甲型流感病毒的总阳性率为29.3%.623例中有23例为危重症病例(占阳性患者的3.7%),其中5例死亡.618例信息完整的甲型H1N1病例中,患儿年龄为14天～16岁,性别比例为男比女为1.3:1.1～3岁儿童占25.2%,3～6岁学龄前儿童和6～12岁学龄儿童所占比例相近,各约占30%.在监测期间,仅呈现了一个甲型H1N1的流行波.2009年11月达到最高峰,随后减弱,2010年2月快速下降至2.7%.对监测期间每周20～30份临床标本同时进行季节性流感的监测显示,季节性H3N2、甲型H1N1和乙型流感交替流行.呼吸道合胞病毒(RSV)在甲型H1N1流行趋势减缓后逐渐流行成为流行优势株.结论 2009年6月至2010年2月北京地区儿童中出现甲型H1N1的流行,主要累及学龄前和学龄儿童.季节性流感和RSV与甲型H1N1交替流行.     

流感病毒H1N1感染后宿主细胞MTH1基因表达的研究

目的观察H1N1型流感病毒诱导的MDCK细胞抗氧化相关修复基因MTH1表达量的变化。方法 MDCK细胞培养后,H1N1型流感病毒感染0、1、3、6、12、24、48h,在各时间点经RT-PCR反应检测细胞内MTH1基因的相对表达量。结果甲型H1N1流感病毒感染后0、6、12h,MTH1基因的表达量与正常对照组差异无统计学意义;感染后1、3hMTH1基因的表达量显著高于正常对照组;感染后24、48h表达量显著低于正常对照组。结论 H1N1流感病毒感染细胞的MTH1基因表达的增加,可能增加细胞对DNA氧化损伤的修复能力,在流感发生和防御机制中起到作用。     

HLA-DRB1, -DQA1, -DQB1, -DPA1 and -DPB1 genes in Japanese multiple sclerosis patients

Japanese MS patients and controls were examined for the distribution of HLA-DRB1, -DQA1, -DQB1, -DPA1 and -DPB1 alleles using in vitro amplification of genomic DNA and probing with sequence-specific oligonucleotides. No significant difference in frequency of the examined alleles was observed among the two groups. This is in contrast to Norwegian MS patients, where an association to a combination of certain DQA1 and DQB1 alleles has previously been demonstrated.     

NDE1 and NDEL1: Multimerisation,alternate splicing and DISC1 interaction

Nuclear Distribution Factor E Homolog 1 (NDE1) and NDE-Like 1 (NDEL1) are highly homologous mammalian proteins. However, whereas NDEL1 is well studied, there is remarkably little known about NDE1. We demonstrate the presence of multiple isoforms of both NDE1 and NDEL1 in the brain, showing that NDE1 binds directly to multiple isoforms of Disrupted in Schizophrenia 1 (DISC1), and to itself. We also show that NDE1 can complex with NDEL1. Together these results predict a high degree of complexity of DISC1-mediated regulation of neuronal activity.     

韩国人CYP450 1A1、CYP450 2E1和GSTM1、GSTT1、GSTP1基因座遗传多态性分析

目的 调查代谢相关的CYP4501A1、CYP4502E1和GSTM1、GSIT1、GSTP1基因座在韩国人群中的遗传多态性分布状况。方法 采用多重聚合酶链式反应、聚合酶链式反应-限制性片段长度多态性技术，分析300名韩国健康大学生的CYP1A1基因3′端限制性内切酶Msp Ⅰ位点、CYP2E1基因5′端转录调节区Pst Ⅰ位点和GSTM1、GSTT1缺失与存在、GSTP1基因第5外显子BsmA Ⅰ位点的基因型，计算基因型和基因频率。结果 CYP1A1基因型频率为ml／ml型39．7％、ml／m2型49．7％、m2／m2型10．7％，基因频率为ml 0．645、m2 0．355。CYP2E1基因型频率为cl／cl型66．7％、cl／c2型30％、c2／c2型3．3％，基因频率为C1 0．818、C2 0．182。GSTM1基因缺失型频率为53．3％。GSTT1基因缺失型频率为54．7％。GSTP1基因型频率为Ile／Ile型62％、Ile／Val型34．3％、VaL／Val型3．7％，基因频率为Ile 0．792、Val 0．208。基因分布符合Hardy-Weirtberg平衡定律。结论 韩国人CYP1A1、CYP2E1、GSTM1、GSTT1基因分布与我国人群较为相近，半数以上人缺乏GSTM1和GSTT1基因，纯合缺失型频率超过印度人的3倍。     

Maternal genetic polymorphisms in CYP1A1, GSTM1 and GSTT1 and the risk of hypospadias

Hypospadias is one of the most common congenital anomalies. Increased exposure to environmental factors (endocrine-disrupting chemicals and smoking) or maternal endogenous estrogen may cause hypospadias because male sexual differentiation is dependent on normal androgen homeostasis. Moreover, interactions between genetic factors and cigarette smoking and other chemicals have been suggested. It has been demonstrated that the CYP1A1 metabolizes not only environmental chemicals but also estrogens, and glutathione-S-transferases (GSTs) are detoxification enzymes that protect cells from toxicants by conjugation with glutathione. In this study, to investigate the association of CYP1A1 (MspI), GSTM1 and GSTT1 polymorphisms with hypospadias, a case-control study of 31 case mothers who had boys with hypospadias and 64 control mothers was performed in Japan. These polymorphisms were investigated by PCR-based methods using DNA from peripheral lymphocytes. We found that the heterozygous CYP1A1 and heterozygous and homozygous CYP1A1 were less frequent in the case mothers than in the control mothers [adjusted odds ratio (OR)=0.17, 95% confidence interval (CI)=0.04-0.74, OR = 0.28, 95% CI = 0.08-0.97, respectively]. We found no effect of maternal smoking on the hypospadias risks among the gene polymorphisms. The results suggest that mothers with the CYP1A1 MspI variant allele may have a decreased risk for hypospadias.     

Rb1cc1 is critical for myoblast differentiation through Rb1 regulation

Rb1-inducible coiled-coil 1 (Rb1cc1) expressed at high levels is associated with the maturation of human embryonic musculoskeletal cells. To clarify the molecular role of Rb1cc1 in muscular differentiation, we investigated the expression of Rb1cc1 and other genes that regulate differentiation in murine embryonic tissues and in C2C12 myoblasts. We also evaluated the effects of RNA interference (RNAi)-mediated Rb1cc1 knockdown on C2C12 myoblast differentiation. After Rb1cc1, Rb1 and myosin heavy chain (Myhc) were expressed in mouse embryonic muscles. The synchronous expression of Rb1cc1 and Rb1 predicted Myhc expression during C2C12 myoblast differentiation. RNAi-mediated knockdown of Rb1cc1 led to Rb1 suppression, and C2C12 myoblasts failed to differentiate. These results indicated that Rb1cc1 is a potent regulator of the Rb1 pathway and a novel mediator that plays a crucial role in muscular differentiation. Rb1cc1 expression is, thus, a prerequisite for myogenic differentiation.     

Circulatinglong non-coding RNA FEZF1-AS1 and AFAP1-AS1 serve as potential diagnostic biomarkers for gastric cancer

BackgroundGrowing evidence indicates that two long non-coding RNAs (lncRNAs), FEZ family zinc finger 1 antisense RNA 1 (FEZF1-AS1) and Actin filament associated protein 1 antisenseRNA1 (AFAP1-AS1), are highly expressed in different cancers, including gastric cancer (GC). However, the expression pattern and clinical utility of these two lncRNAs are still unknown.MethodsSerum expression levels of FEZF1-AS1 andAFAP1-AS1 were measured by quantitative real-time polymerase chain reaction (qRT-PCR). CEA and CA19-9 were detected by ARCHITET I2000 SR. Analyses were all performed using SPSS software version 20.0 (SPSS Inc., Chicago, USA). P < 0.05 was considered statistically significant.ResultsDetection of serum FEZF1-AS1 and AFAP1-AS1 showed both of them were up-regulated in GC patients compared with the normal controls (p < 0.0001), and high serum expression levels were correlated with tumor size, tumor-node-metastasis (TNM) stage and lymph node metastasis. Besides, the area under the ROC curve (AUC) demonstrated the two lncRNAs had higher diagnostic utility than CEA and CA19-9. Furthermore, when combined the two lncRNAs as a model, it yielded an AUC of 0.866, and the combination of the model, CEA and CA19-9 could observably improve diagnostic sensitivity to 95.5 %. What’s more, circulating FEZF1-AS1 and AFAP1-AS1 were significantly decreased after the GC patients underwent the operation (both p < 0.001).ConclusionOur study indicated that serum FEZF1-AS1 and AFAP1-AS1 had better sensitivity and efficiency for the diagnosis of GC and the combination of the two lncRNAs might be used as a potential prognostic indicator in GC.     

神经系统RNA结合蛋白Musashi1 RRM1的初步结晶

目的 对Musashi1发挥功能的 RRM1结构域进行结晶,得到可用来衍射的蛋白晶体,为之后的结构解析打基础。方法 通过构建Musashi1RRM1的原核表达载体,并在BL21中表达、纯化高纯度的蛋白质,通过筛选结晶体条件得到蛋白晶体。结果 通过系统筛选和优化晶体生长条件得到了蛋白晶体。结论 Musashi1 RRM1的蛋白晶体质量较好,满足蛋白晶体衍射和数据收集的要求。     

HIV-1 Nef stabilizes AP-1 on membranes without inducing ARF1-independent de novo attachment

HIV-1 Nef affects the trafficking of numerous cellular proteins to optimize viral replication and evade host defenses. The adaptor protein (AP) complexes, which form part of the cytoplasmic coat of endosomal vesicles, are key cellular co-factors for Nef. Nef binds these complexes and alters their physiologic cycle of attachment and release from membranes. Specifically, while AP-1 normally becomes cytosolic when attachment events are blocked by inhibition of the GTPase cycle of ADP-ribosylation factor-1 (ARF1), the complex remains membrane-associated in Nef-expressing cells. To investigate the mechanism of this effect, we used a permeabilized cell system to detect the de novo attachment of exogenous AP-1 to endosomal membranes. Nef did not mediate de novo attachment independently of ARF1, despite its ability to maintain the association of AP-1 with endosomal membranes when the activity of ARF1 was blocked. We conclude that Nef stabilizes AP complexes on endosomal membranes after ARF1-dependent attachment. This stabilization may facilitate coat formation and stimulate the trafficking of multiple cellular proteins.     

IL—1受体相关激酶—1和—2协同调控IL—1诱导的AP—1的活化

目的研究白介素 - 1受体相关激酶 - 1(IRAK- 1)和 IRAK- 2在白介素 - 1(IL - 1)诱导 AP- 1活化中的作用。方法L ipofectin介导反义 IRAK- 1寡核苷酸和反义 IRAK- 2寡核苷酸转染 Hep G2细胞。用逆转录 PCR法检测 IRAK - 1和 IRAK- 2m RNA表达水平 ;Western blot分析 IRAK- 1和 IRAK - 2蛋白表达水平。以 Sandwich EL ISA法检测 AP- 1的活化。结果反义IRAK- 1寡核苷酸和反义 IRAK- 2寡核苷酸通过抑制各自靶基因 m RNA和蛋白表达抑制 IL- 1诱导的 AP- 1活化 ;反义 IRAK-1寡核苷酸与反义 IRAK- 2寡核苷酸共转染 Hep G2细胞对 AP- 1的抑制作用较两者单独转染明显增强。结论 IRAK- 1和 I-RAK- 2在调控白介素 - 1诱导的 AP- 1活化时协同作用。     

Ah receptor, CYP1A1, CYP1A2 and CYP1B1 gene polymorphisms are not involved in the risk of recurrent pregnancy loss

The etiology of recurrent pregnancy loss (RPL) remains unclear, but it may be related to a possible genetic predisposition together with involvement of environmental factors. We examined the relation between RPL and polymorphisms in four genes, human aryl hydrocarbon (Ah) receptor, cytochrome P450 (CYP) 1A1, CYP1A2 and CYP1B1, which are involved in the metabolism of a wide range of environmental toxins and carcinogens. All cases and controls were women resident in Sapporo, Japan and the surrounding area. The Ah receptor, CYP1A1, CYP1A2 and CYP1B1 genotypes were assessed in 113 Japanese women with recurrent pregnancy loss (RPL) and 203 ethnically matched women experiencing at least one live birth and no spontaneous abortion (control). No significant differences in Ah receptor, CYP1A1, CYP1A2 and CYP1B1 genotype frequencies were found between the women with RPL and the controls [Ah receptor: Arg/Arg (reference); Arg/Lys and Lys/Lys, odds ratio (OR)=0.67; 95% confidence interval (CI)=0.40-1.11, CYP1A1: m1m1 (reference); m1m2 and m2m2, OR = 0.86; 95% CI = 0.53-1.40, CYP1A2: C/C and C/A (reference); A/A, OR = 1.16; 95% CI = 0.71-1.88, CYP1B1: Leu/Leu (reference); Leu/Val and Val/Val, OR = 1.18; 95% CI = 0.68-2.02]. The present study suggests that the Ah receptor, CYP1A1, CYP1A2 and CYP1B1 gene polymorphisms are not major genetic regulators in RPL.