DEHP对大鼠睾丸支持细胞间隙连接蛋白43表达和间隙连接通讯功能的影响

目的 通过对原代培养的大鼠睾丸支持细胞进行邻苯二甲酸二乙基己酯(di-2-ethylhexyl phthalate,DEHP)染毒,探讨DEHP体外对大鼠睾丸支持细胞间隙连接蛋白43(connexin 43,Cx43)表达及间隙连接通讯(gap junction intercellular communication,GJIC)功能的影响.方法 体外分离和原代培养大鼠睾丸支持细胞,应用不同浓度的DEHP(10、50、100nmol/L)作用于支持细胞24h,应用荧光定量PCR法检测Cx43基因mRNA表达,用Weastern blot方法检测Cx43蛋白表达,划痕标记荧光染料示踪方法检测GJIC功能.结果 与对照组相比,DEHP各组支持细胞Cx43基因mRNA和蛋白表达均下降(P＜0.05),呈浓度依赖关系,同时支持细胞GJIC功能降低.结论 DEHP在体外可通过抑制大鼠睾丸支持细胞Cx43基因表达而抑制GJIC功能,进而影响支持细胞功能.     

乙醇对胰岛β细胞功能及胰岛素基因表达影响

目的 观察乙醇对原代培养的大鼠胰岛细胞及小鼠胰岛瘤细胞（NIT-1细胞）胰岛素分泌功能的影响并探讨其机制。方法 体外培养的大鼠胰岛细胞及NIT-1细胞，不同剂量乙醇作用不同时间后，用放免法测定胰岛素分泌情况，RT-PCR方法检测胰岛素mRNA的表达。结果 （1）乙醇对NIT-1细胞胰岛素分泌及胰岛素mRNA表达影响：作用6h，胰岛素分泌增加，各剂量组（除400mmol／L）胰岛素mRNA表达升高；作用12，24h，胰岛素分泌降低，胰岛素mRNA表达均降低。（2）乙醇对原代培养的大鼠胰岛细胞的胰岛素分泌及胰岛素mRNA表达影响：作用12h，低剂量组胰岛素分泌升高，胰岛素mRNA表达升高，高剂量组胰岛素分泌下降；胰岛素mRNA表达下降；随乙醇作用时间延长，胰岛素分泌先升高（6h），然后下降（24h），胰岛素mRNA表达短时间（6h）上升，作用24h表达下降。结论乙醇对胰岛β细胞胰岛素分泌及胰岛素mRNA表达的影响与作用时间、剂量有关，胰岛素mRNA表达水平的改变可能是乙醇影响胰岛β细胞胰岛素分泌的机制之一。     

肠内营养对阻塞性黄疸大鼠肝损伤及清蛋白基因转录的影响

目的:观察不同配方肠内营养对阻塞性黄胆(OJ)大鼠肝损伤及清蛋白基因转录的影响.方法:将40只雄性SD大鼠随机分为两大组:即假手术(SHAM)组和胆总管结扎(CBDL)组.后者进一步分为CBDL经口饮食对照组(对照组)、CBDL 能全素组和CBDL 能全力组,每组10只.接受持续5天的等氮、等热量的营养支持.总热量为630 kJ/(kg·d),氮量为1.2 g/(kg·d).结果:CBDL大鼠出现明显的肝功能损伤,能全素和能全力组大鼠肝功能损伤程度与对照组相似.能全素和能全力组大鼠体重下降程度均明显改善,血清总蛋白和清蛋白水平无显著性变化.SHAM组大鼠肝清蛋白信使核糖核酸(ALB mRNA)和D位点结合蛋白信使核糖核酸(DBP mRNA)表达明显高于CBDL各组.CBDL各组中,能全素和能全力组大鼠肝ALB mRNA略高于对照组,但无显著性差异.能全素和能全力组大鼠在体重、肝功能损伤、血清蛋白水平和清蛋白基因转录方面均无显著性差异.结论:短期肠内营养没有加重OJ大鼠肝功能损伤.能全素和能全力较经口饮食能更好地维持大鼠体重,但不能明显促进OJ大鼠肝清蛋白基因转录和提高血清蛋白水平.     

靶向血管紧张素Ⅱ受体基因干扰及有效shRNA的筛选(英文)

目的RNA干扰是基因技术的重要进展,人们可以在转录后水平减少基因的表达。对siRNA是否有效而言,基因靶序列的选择是至关重要的因素。我们研究了哺乳细胞中shRNA调节大鼠血管紧张素Ⅱ1型受体基因表达的有效性并探索有效shRNA的特征与shRNA功能性之间的关系。方法设计并构建四个靶向大鼠AT1受体基因的shRNA表达载体。4个shRNA靶向同一基因的不同区域。合成shRNA寡核苷酸片段的正义链和反义链,退火形成双链并克隆进入载体pGenesil-1。用构建完成的pGenesil-1-shRNA质粒和重新洗牌的对照质粒转染大鼠胶质瘤细胞,并在不同时相收集转染的细胞,进行RT-PCR和Western blot检测。结果转染24小时之后,各处理组均未见AT1mRNA水平有明显减少。与对照组相比,Pb组在转染后48小时血管紧张素Ⅱ受体mRNA基因水平下降到55.7±7.6%,72小时后达到最低点(43.7±8.2%)。其它组AT1受体mRNA表达无明显抑制(P>0.05)。抑制大鼠血管紧张素Ⅱ受体mRNA基因及其蛋白的结果类似。与对照组相比,Pb组血管紧张素Ⅱ受体蛋白在24小时和48小时分别减少至46.9±4.2%和36.98±3.7%,最大减少在转染后72小时(28.1±4%)。结论成功地构建了编码大鼠血管紧张素Ⅱ受体mRNA的短发夹RNA真核表达载体后,在体外转染哺乳细胞中表达shRNA可有效地抑制靶基因的表达,并导致其蛋白的翻译受到抑制,达到基因干扰的目的。在选择有效的shRNA序列时,除了一般的原则之外,我们认为靶位mRNA的环状结构以及siRNA是否能到达靶位对基因干扰实验能否成功具有重要作用。     

胶质细胞源性神经营养因子与肿瘤坏死因子α在星形胶质细胸活化后的分泌特点研究

目的观察星形胶质细胞活化后胶质细胞源性神经营养因子(glial cell derived neurotrophic factor,GDNF)、肿瘤坏死因子α(tumor necrosis factor,TNF)α两种不同作用因子的分泌特点,探讨星形胶质细胞活化在神经退行性疾病后的神经再生和功能恢复的作用。方法体外分离培养星形胶质细胞,分为对照组、活化组、抑制组。通过光学显微镜及免疫荧光化学观察各组细胞的形态变化;应用半定量RT-PCR方法分析各组细胞间胶原纤维酸性蛋白(glial fibrillary acidic protein,GFAP)mRNA及GDNF mRNA、TNF mRNA表达变化;用ELISA法检测各组细胞上清液中不同时间点(6、24、48、72 h)GDNF、TNF的含量。计量资料用x珋±s表示,采用t检验,多组数据间的比较采用单因素方差分析,P<0.05为差异有统计学意义。结果活化组与对照组比较,细胞数量增多,胞体变大,突起变长增多,交织成网状,GFAP荧光增强;RT-PCR示GFAP mRNA、GDNF mRNA、TNF mRNA表达均明显增高,与对照组比较差异有统计学意义(P<0.01);ELISA法检测示星形胶质细胞活化后GDNF分泌量在活化后48 h达到高峰,与对照组比较差异有统计学意义(P<0.01);活化后24 hTNF的含量达到高峰,与对照组比较差异有统计学意义(P<0.01);应用抑制剂Olomoucine干预后,与活化组相比,抑制组细胞胞体变小,数量减少,星形胶质细胞活化被抑制,GFAP mRNA表达下降,GDNF mRNA、TNF mMRA表达亦下降,与活化组比较差异有统计学意义(P<0.01)。结论星形胶质细胞活化后GDNF、TNF表达均上调,但TNF表达高峰时间早于GDNF表达时间;Olomoucine抑制星形胶质细胞活化,可使GDNF、TNF表达下调。表明星形胶质活化后TNF可能有促进GDNF分泌从而起到一定的神经功能恢复作用。     

急性肝衰竭大鼠微囊化肝细胞移植后HMGB1的动态变化

目的 本实验将微囊化肝细胞移植人急性肝衰竭大鼠腹腔内,动态观察大鼠肝功能和肝组织中高迁移率族蛋白B1(HMGB1)的表达情况和意义. 方法 用D-氨基半乳糖制作大鼠急性肝衰竭模型,随机分成3组:Ⅰ组为生理盐水组,Ⅱ组为裸肝细胞移植组,Ⅲ组为微囊化肝细胞移植组.在6、12、24、48、72、120和168 h,检测各组血生化,用RT-PCR方法检测肝组织HMGB1 mRNA的表达.另取6只正常大鼠样本作为参考值. 结果 各模型组ALT、AST和TBil在6 h就开始上升,24 h均已显著升高(P＜0.05),Ⅲ组ALT、AST和TBil显著下降,与Ⅰ、Ⅱ组比较差异有统计学意义(P＜0.05或P＜0.01).各模型组HMGB1mRNA在6 h显著升高(P＜0.01),12 h达高峰,Ⅱ组、Ⅲ组在24 h后表达量较Ⅰ组显著下降,72 h、120 h时Ⅲ组与Ⅱ组比较差异有统计学意义(P＜0.01). 结论 HMGB1可能参与了急性肝衰竭的病理生理过程,微囊化肝细胞移植组明显减少了肝衰竭大鼠肝组织中HMGB1 mRNA的表达,从而减轻肝组织炎症反应并改善肝衰竭大鼠的预后.     

青蒿琥酯对高温复合内毒素致细胞凋亡保护作用

目的 探讨高温复合内毒素(LPS)作用下,青蒿琥酯(AR)对RAW264.7细胞热休克蛋白70(HSP70)合成及细胞凋亡的影响.方法采用体外培养的巨噬细胞系RAW264.7细胞.分为空白对照组、高温复合内毒素组、AR组(又分为4个亚组),于40℃、95%O2、5%CO2条件下共同孵育24h.用RT-PCR技术检测细胞内HSP70mRNA及其蛋白的表达量,用流式细胞术检测细胞凋亡率.结果 空白组和高温复合内毒素组HSP70m RNA的表达率分别为10%,23%,而AR组可显著提高HSP70mRNA的表达率,为54%,并与高温内毒素组差异有统计学意义(P＜0.05).HSP70蛋白的表达情况与HSPT0mRNA一致,而细胞凋亡率的变化则呈相反趋势.结果 AR进一步诱导高温复合内毒素作用下RAW264.7细胞HSP70m RNA和HSP70蛋白的表达,且可明显降低细胞的凋亡率,可能是其抗损伤保护作用的机制之一.     

酒精对小鼠NIT-1细胞胰岛素分泌及COXⅡ基因表达的影响

目的:观察酒精对小鼠NIT-1细胞胰岛素分泌功能的影响并初步探讨其机制。方法:体外培养的小鼠NIT-1细胞,经不同浓度酒精(0、50、100、200、400mmol/L)作用6、12、24h后,用放免法测定NIT-1细胞分泌胰岛素情况,半定量RT-PCR方法检测细胞色素氧化酶Ⅱ(cytochromeoxidaseⅡ,COXⅡ)基因mRNA的表达。结果:酒精作用6h,NIT-1细胞葡萄糖刺激的胰岛素分泌增加;作用12h、24h,葡萄糖刺激的胰岛素分泌降低。酒精作用6h,COXⅡmRNA表达升高,12h表达开始降低,24h时高浓度组表达呈显著性降低。结论:酒精可增加或降低NIT-1细胞胰岛素分泌,与酒精作用时间、浓度有关。酒精导致COXⅡmRNA表达水平的改变很可能是酒精影响NIT-1细胞胰岛素分泌的机制之一。     

氢醌诱导下肝细胞XPV基因mRNA表达规律的研究

目的研究氢醌(HQ)对L-02肝细胞XPV基因mRNA表达的影响及其规律,探讨XPV基因在HQ所致肝细胞毒性过程中分子作用机制。方法体外培养的L-02肝细胞经40μmol/L的HQ诱导后,采用实时荧光定量PCR法检测不同时间点(6、12、24和48h)XPV基因mRNA的表达;并检测不同剂量(0、5、10、20、40、80和160μmol/L)HQ诱导24h后,各浓度组XPV基因mRNA的表达。结果在24h的时间范围内,XPV基因在mRNA水平上的表达无论是染毒组(40μmol/L)还是正常对照组,均具有随时间的延长而增加的趋势;到24h时,相对表达量达到最大值,当培养时间达到48h时,该基因的表达则有所下降;此外,在各时间处理组(6h、12h、24h和48h)中,染毒组L-02肝细胞内XPV基因在mRNA水平上的表达均高于正常对照组。各剂量HQ作用24h后,L-02肝细胞内XPV基因mRNA的表达在0～80μmol/L范围内随着剂量的增加而增加,以正常对照组XPV基因的相对表达量为1,各染毒剂量组内XPV基因mRNA的相对表达量分别增至1.20、2.02、2.37、2.67、4.40和2.32倍;80μmol/L时其相对表达量达到峰值,160μmol/L的HQ作用后,其表达却有所下降,但仍高于正常对照组。结论HQ可以诱导XPV基因mRNA的表达变化,并且呈现一定的时间和剂量依赖性,提示XPV基因可能参与了肝细胞对HQ的应答过程。     

小剂量顺铂增加宫颈癌细胞凋亡抑制蛋白Survivin表达的研究

目的:研究化疗药物顺铂对宫颈癌细胞内凋亡抑制蛋白Survivin的诱导作用, 探讨Survivin在宫颈癌化疗抵抗中的作用机制。方法: 培养人宫颈癌细胞株Hela, 加入低浓度的化疗药物顺铂, 分别在加药前、加药后24h、48h、72h和96h采用逆转录聚合酶链反应(RT-PCR) 和免疫印迹技术(western-blot) 检测SurvivinmRNA和蛋白的表达。结果: 给予低浓度的顺铂可诱导SurvivinmRNA和蛋白的表达发生变化, 其中mRNA在加药后24h、48h、72h和96h分别较加药前提高了1 .5倍、2 .6倍、4 .9倍和1. 6倍, 蛋白在加药后24h、48h、72h和96h分别较加药前提高了0 .6倍、1. 7倍、2 .9倍和4 .7倍。结论:小剂量顺铂可诱导宫颈癌细胞内Survivin的表达增加, 抵抗化疗药物诱导的细胞凋亡。     

Calcium intake in the elderly: validation of a dietary questionnaire

Epidemiological studies of calcium and osteoporosis have been hampered by the lack of a suitable tool for assessing calcium intake. This report describes a new frequency and amount questionnaire for measuring present and past calcium intake in the elderly. The validity of the questionnaire was tested against two commonly used standards of dietary assessment, five-day duplicate diets and seven-day weighed dietary inventories. The resulting correlation coefficients were, respectively, r = 0.76 and r = 0.69, while that for repeatability was r = 0.84. Furthermore, the questionnaire categorized subjects into thirds of the distribution of intake with almost no gross misclassification. It is suggested that the present findings may be extended to the majority of normal, healthy elderly subjects, implying wide application for the questionnaire in the assessment of calcium intake in the elderly.     

Self-reported health problems among Swedish miners one year after unemployment

Unemployment is considered to be a public health concern sincedeterioration in the health of the unemployed is often anticipated.However, for some groups, such as miners, unemployment mightimprove health due to a cessation of potentially harmful occupationalexposures. This study evaluates the health of 79 miners in oneSwedish iron-ore mine, and 226 age-matched controls from thegeneral population, during one year after the closure of themine. The participants received a questionnaire regarding medicalhistory and subjective symptoms at the beginning of the studyperiod, and after one year. Statistically significant negativeeffects on self-reported health attributable to unemploymentwere not found, although neuropsychiatric symptoms were morecommon among the unemployed miners. The miners reported a statisticallysignificant improvement in grip force (p=0.031). They had asignificantly higher prevalence of symptoms associated withmining related exposures when compared with the population controls;pain in the upper extremities [relative risk (RR)=2.27, 95%confidence interval (Cl)=1.44–3.59), back pain (RR=1.84;Cl=1.237–2.75), vasospastic disease of the fingers (RR=2.05;Cl=1.18–3.57) and obstructive respiratory symptoms (attacksof dyspnea and wheezing: RR=3.67; Cl=1.167–11.6).     

Excès de cas humains et animaux de tularémie en France au cours de l’hiver 2007–2008 : émergence ou phénomène isolé ?

Context

Tularemia is a zoonosis affecting humans and hares in France. We describe the results of surveillance in both species, in 2007 and 2008.

Methods

Human tularemia cases are mandatorily notifiable in France since 2003. In hares, surveillance relies on volunteer hunter associations in all districts of the country. Data from mandatory reports and volunteer surveillance in 2007/2008 were analyzed and compared with previous results.

Results

In 2007/2008, 144 cases were reported in humans and 117 cases in hares. This was a 100% increase compared to previous years. Human cases differed from those of previous years only by the frequency of contact with breeding animals. Human cases without any documented risk exposure were also more frequent.

Conclusion

An increase of tularemia cases occurred in 2007/2008 in both species. Complementary studies are needed to identify the species reservoir in France to understand the causes of this peak of cases.     

Occupational health hazards in mining: an overview

This review article outlines the physical, chemical, biological, ergonomic and psychosocial occupational health hazards of mining and associated metallurgical processes. Mining remains an important industrial sector in many parts of the world and although substantial progress has been made in the control of occupational health hazards, there remains room for further risk reduction. This applies particularly to traumatic injury hazards, ergonomic hazards and noise. Vigilance is also required to ensure exposures to coal dust and crystalline silica remain effectively controlled.     

Manufacture and use of nanomaterials: current status in the UK and global trends

This paper provides an overview of the production and use of nanomaterials (NMs), particularly in the UK. Currently, relatively few companies in the UK are identifiable as NM manufacturers, the main emphasis being the bulk markets in metals and metal oxides, and some niche markets such as carbon nanotubes and quantum dots. NM manufacturing in the UK does not reflect the global emphasis on fullerenes, nanotubes and fibres. Some assumptions have been made about the types of NM that are likely to be imported into the UK, which currently include fullerenes, modified fullerenes and other carbon-based NMs including nanotubes. Many university departments, spin-offs and private companies have developed processes for the manufacture of NMs but may only be producing small quantities for research and development (R&D) purposes. However, some have the potential to scale up to produce large quantities. The nanotechnology industry in the UK has strong R&D backup from universities and related institutions. This review has covered R&D trends at such institutions, and appropriate information has been added to a searchable database. While several companies are including NMs in their products, only a few (e.g. manufacturers of paints, coatings, cosmetics, catalysts, polymer composites) are using nanoparticles (NPs) in any significant quantities. However, this situation is likely to change rapidly. There is a need to collect more information about exposure to NPs in both manufacturing and user scenarios. As the market grows, and as manufacturers switch from the micro- to the nanoscale, the potential for exposure will increase. More research is required to quantify any risks to workers and consumers.     

深圳公立医院管理体制改革制度设计

深圳公立医院管理体制改革实行政事分开、管办分开,在理事会架构下按法定机构模式组建市医管中心,落实公立医院运营管理自主权.作者从当前公立医院管理体制的弊端入手,介绍了深圳市进行公立医院管理体制改革的基本思路及改革方案设计的主要举措,深入剖析了的改革方案的特点,并对改革效果进行了预测.     

Effects of nitrogen dioxide on red blood cells of rats: Changes in components of red cell membranes during in vivo exposure to NO2

Red cell membranes, prepared from red blood cells of rats exposed to 4, 10, or 20 ppm nitrogen dioxide (NO₂) for 1 to 10 days, were examined for evidence of changes in membrane components. Appreciable changes were not found in contents of phospholipid and cholesterol during exposure to 10 ppm NO₂. By contrast, protein content altered with the time of exposure. Moreover, changes in protein composition were observed by employing sodium dodecyl sulfate — polyacrylamide gel electrophoresis. Twenty-four-hour exposure to NO₂ at the concentration above 10 ppm resulted in a marked increase in the percentage of lysophosphatidylethanolamine (LysoPE) to the total phospholipids. The prolonged exposure to 10 ppm NO₂ gave rise to a further increase in LysoPE, whereas the percentage of phosphatidylethanolamine (PE) showed a gradual decrease. A 1-day exposure to 4.0 ppm NO₂ also caused an increase in sialic acid content and decreases in those of PE and hexose. In addition to contents of these components the percentage of LysoPE increased 5 days after exposure and the elevated values were maintained up to the end of exposure period. These results demonstrate that red blood cells in circulation exhibit different membrane properties in terms of lipid and carbohydrate composition during 10 days of exposure to 4.0 ppm NO₂.     

Investigating clusters in the workplace and beyond

Clusters of disease are common and occur in the workplace and in the general community. They often arouse considerable concern among the population. Investigations have sometimes lead to exciting new knowledge, but in general the investigation of clusters is difficult and often unrewarding, especially for community clusters. In the workplace, investigations are more likely to find associations and even new causes, but still many clusters remain enigmatic. Despite this, there are many reasons for investigating clusters, including allaying community concern and identifying uncontrolled exposures. A structure for investigating clusters in the workplace is suggested.