德都红花-7味散原药方与优化方对肝纤维化大鼠Ⅰ,Ⅲ型胶原mRNA表达的影响

目的:探讨德都红花-7味散原药方及优化方对肝纤维化大鼠Ⅰ,Ⅲ型胶原mRNA表达的影响。方法:40只Wistar大鼠分为正常组、模型组、阳性药组、原药组、优化组。大鼠ip 30%CCl4橄榄油溶液建立肝纤维化模型。同时1次/日ig给药,阳性药组给予秋水仙碱片0.4 mg·kg-1;原药组给予德都红花-7味散0.6 g·kg-1;优化组给予德都红花-7味散优化方0.6 g·kg-1。连续40 d后处死大鼠。取肝脏天狼星红染色,观察肝组织纤维化程度和Ⅰ,Ⅲ型胶原分型。酶联免疫吸附法(ELISA)检测肝组织匀浆Ⅲ型前胶原和Ⅳ型胶原,层粘连蛋白(LN),透明质酸(HA)含量。实时荧光定量PCR法检测Ⅰ型胶原,Ⅲ型胶原mRNA表达变化。结果:天狼星红染色模型组Ⅰ,Ⅲ型胶原纤维较正常组增多,阳性药组、原药组、优化组Ⅰ,Ⅲ型胶原纤维较模型组减少。与正常组比较,酶联免疫法检测模型组Ⅳ型胶原,HA含量升高(P0.05)。与模型组比较,阳性药组、原药组、优化组Ⅲ型前胶原、Ⅳ型胶原、HA含量明显降低(P0.05)。实时荧光定量PCR法检测模型组Ⅰ型胶原mRNA和Ⅲ型胶原mRNA表达较正常组上调(P0.01)。阳性药组、原药组、优化组Ⅰ型胶原mRNA和Ⅲ型胶原mRNA表达较模型组下调(P0.01)。原药组、优化组Ⅰ型胶原mRNA表达较阳性药组下调(P0.05)。优化组Ⅰ型胶原mRNA表达较原药组下调(P0.05)。结论:德都红花-7味散原药方与优化方是通过抑制Ⅰ型胶原mRNA和Ⅲ型胶原mRNA的转录,达到阻断或延缓肝纤维化的发生发展,德都红花-7味散优化方优于原药方。     

Efficacy and complications of covered Wallstents in malignant distal biliary obstruction

BACKGROUND: This study evaluated the efficacy and the complications associated with the use of the covered Wallstent in the setting of unresectable malignant biliary obstruction. METHODS: Between March 2001 and January 2003, all patients with distal malignant biliary obstruction that required drainage were treated with a covered Wallstent. Every 2 months, the patients were evaluated clinically and biochemical tests of liver function were obtained. Data were recorded for the following variables: early complications (within 30 days of stent placement), early and late stent occlusion, duration of stent patency, need for subsequent biliary intervention, and patient survival. RESULTS: A total of 88 covered Wallstents were inserted in 80 patients. Stent patency rates at 3, 6, and 12 months were 90%, 82%, and 78%, respectively. Complications included stent migration (5), stent occlusion (12), episodes of cholecystitis (3), and episodes of post-ERCP pancreatitis (5). Biliary intervention was required in 9 patients subsequent to placement of the initial covered Wallstent. CONCLUSIONS: Deployment of a covered Wallstent is safe and relatively easy. It achieves biliary drainage with an acceptable risk to benefit ratio in the majority of patients with distal malignant biliary obstruction.     

中孕期非整倍体染色体异常血清学筛查不同方案比较

目的：对中孕期非整倍体染色体异常血清学筛查不同方案的检出率进行探讨。方法选取513名2009年9月~2013年3月在攀枝花市妇幼保健院产科进行常规产前筛查的孕妇血清样本,分别采用化学发光法和时间分辨荧光免疫法对孕妇血清进行二联、三联、四联检查,比较阳性率及假阳性率。结果二联化学发光法检测唐氏综合征(DS)高危假阳性率为9.10豫,三联为7.36豫,四联为6.28豫,假阳性率呈递减趋势(字2=5.119,P约0.05);二联化学发光法筛查18-三体高危假阳性率(0.63豫)低于三联(0.82豫),差异有统计学意义(字2=4.776,P约0.05)。三联时间分辨荧光免疫法筛查DS、18-三体假阳性率(4.01豫、0.34豫)较二联(8.93豫、0.61豫)均明显降低(字2=6.992、4.776,P约0.05)。二联化学发光法检测DS、18-三体假阳性率分别为9.10豫、0.63豫,时间分辨荧光免疫法则分别为8.93豫、0.61豫,两种检测方法比较,差异无统计学意义(字2=1.787、0.000,P跃0.05);而三联时间荧光分辨法检测DS、18-三体假阳性率(4.01豫、0.34豫)均低于三联化学发光法(7.36豫、0.82豫),差异有统计学意义(字2=5.382、4.783,P约0.05)。结论化学发光法的检测系统发现筛查效率二联、三联及四联方案呈递增趋势。时间分辨荧光免疫法检测系统发现筛查效率三联高于二联方案,时间荧光分辨法三联筛查优于化学发光法。     

Factors affecting the transduction of pluripotent hematopoietic stem cells: long-term expression of a human adenosine deaminase gene in mice

An amphotropic retroviral vector, LgAL(delta Mo + PyF101) containing a human adenosine deaminase (ADA) cDNA was used to optimize procedures for the lasting genetic modification of the hematopoietic system of mice. The highest number of retrovirally infected cells in the hematopoietic tissues of long-term reconstituted mice was observed after transplantation of bone marrow (BM) cells that had been cocultured in the presence of both interleukin-1 alpha (IL-1 alpha) and IL-3. A significantly lower number was detected when IL-1 alpha was omitted from such cocultures. The yield of cells that generate spleen colony-forming cells (CFU-S) in the BM of lethally irradiated recipients (MRA-CFU-S) significantly improved on inclusion of the adherent cell fraction of cocultures in the transplant. Retroviral integration patterns in MRA-CFU-S-derived spleen colonies showed that an MRA-CFU-S can produce many CFU-S during BM regeneration. Expression of hADA was detected in the circulating white blood cells of long-term reconstituted animals, demonstrating that the LgAL(delta Mo + PyF101) vector is capable of directing the sustained expression of hADA, and in approximately 35% of the transduced MRA-CFU-S-derived spleen colonies. These results should facilitate the development of gene therapy protocols for the treatment of severe combined immunodeficiency caused by a lack of functional ADA.     

Detection and phasing of single base de novo mutations in biopsies from human in vitro fertilized embryos by advanced whole-genome sequencing

Currently, the methods available for preimplantation genetic diagnosis (PGD) of in vitro fertilized (IVF) embryos do not detect de novo single-nucleotide and short indel mutations, which have been shown to cause a large fraction of genetic diseases. Detection of all these types of mutations requires whole-genome sequencing (WGS). In this study, advanced massively parallel WGS was performed on three 5- to 10-cell biopsies from two blastocyst-stage embryos. Both parents and paternal grandparents were also analyzed to allow for accurate measurements of false-positive and false-negative error rates. Overall, >95% of each genome was called. In the embryos, experimentally derived haplotypes and barcoded read data were used to detect and phase up to 82% of de novo single base mutations with a false-positive rate of about one error per Gb, resulting in fewer than 10 such errors per embryo. This represents a ∼100-fold lower error rate than previously published from 10 cells, and it is the first demonstration that advanced WGS can be used to accurately identify these de novo mutations in spite of the thousands of false-positive errors introduced by the extensive DNA amplification required for deep sequencing. Using haplotype information, we also demonstrate how small de novo deletions could be detected. These results suggest that phased WGS using barcoded DNA could be used in the future as part of the PGD process to maximize comprehensiveness in detecting disease-causing mutations and to reduce the incidence of genetic diseases.Worldwide, more than 5 million babies (Ferraretti et al. 2013) have been born through in vitro fertilization (IVF) since the birth of the first in 1978 (Steptoe and Edwards 1978). Exact numbers are difficult to determine, but it has been estimated that currently 350,000 babies are born yearly through IVF (de Mouzon et al. 2009, 2012; Centers for Disease Control and Prevention 2011; Ferraretti et al. 2013). That number is expected to rise, as advanced maternal age is associated with decreased fertility rates and women in developed countries continue to delay childbirth to later ages. In 95% of IVF procedures, no diagnostic testing of the embryos is performed (https://www.sartcorsonline.com/rptCSR_PublicMultYear.aspx?ClinicPKID=0). Couples with prior difficulties conceiving or those wishing to avoid the transmission of highly penetrant heritable diseases often choose to perform preimplantation genetic diagnosis (PGD). PGD involves the biopsy of one cell from a 3-d embryo or the recently more preferred method, due to improved implantation success rates (Scott et al. 2013b), of up to 10 cells from a 5- to 6-d blastocyst-stage embryo. Following biopsy, genetic analysis is performed on the isolated cell(s). Currently this is an assay for translocations and the correct chromosome copy number (Hodes-Wertz et al. 2012; Munne 2012; Yang et al. 2012; Scott et al. 2013a; Yin et al. 2013), a unique test designed and validated for each specific heritable disease (Gutierrez-Mateo et al. 2009), or a combination of both (Treff et al. 2013). Importantly, none of these approaches can detect de novo mutations.Advanced maternal age has long been associated with an increased risk of producing aneuploid embryos (Munne et al. 1995; Crow 2000; Hassold and Hunt 2009) and giving birth to a child afflicted with Down syndrome or other diseases resulting from chromosomal copy number alterations. Conversely, children of older fathers have been shown to have an increase in single base and short multibase insertion/deletion (indels) de novo mutations (Kong et al. 2012). Many recent large-scale sequencing studies have found that de novo variations spread across many different genes are likely to be the cause of a large fraction of autism cases (Michaelson et al. 2012; O’Roak et al. 2012; Sanders et al. 2012; De Rubeis et al. 2014; Iossifov et al. 2014), severe intellectual disability (Gilissen et al. 2014), epileptic encephalopathies (Epi4K Consortium and Epilepsy Phenome/Genome Project 2013), and many other congenital disorders (de Ligt et al. 2012; Veltman and Brunner 2012; Yang et al. 2013; Al Turki et al. 2014). Additionally rare and de novo variations have been suggested to be prevalent in patients with schizophrenia (Fromer et al. 2014; Purcell et al. 2014), and Michaelson et al. (2012) found that single base de novo mutations affect conserved regions of the genome and essential genes more often than regions of unknown function. Current targeted approaches to PGD would miss many of these important functional changes within the embryonic DNA sequence, and even a whole-genome sequencing (WGS)–based carrier screen of both parents would not enable comprehensive preimplantation or prenatal diagnoses due to de novo mutations. As more parents delay childbirth into their mid-30s and later, these studies suggest we should try to provide better diagnostic tests for improving the health of newborns. In this study, we demonstrate the use of an advanced WGS process that provides an accurate and phased genome sequence from about 10 cells, allowing highly sensitive and specific detection of single base de novo mutations from IVF blastocyst biopsies.     

Lean Body Mass Associated with Upper Body Strength in Healthy Older Adults While Higher Body Fat Limits Lower Extremity Performance and Endurance

Impaired strength adversely influences an older person’s ability to perform activities of daily living. A cross-sectional study of 117 independently living men and women (age = 73.4 ± 9.4 year; body mass index (BMI) = 27.6 ± 4.8 kg/m²) aimed to assess the association between body composition and: (1) upper body strength (handgrip strength, HGS); (2) lower extremity performance (timed up and go (TUG) and sit to stand test (STS)); and (3) endurance (6-minute walk (SMWT). Body composition (% fat; lean body mass (LBM)) was assessed using bioelectrical impedance. Habitual physical activity was measured using the Minnesota Leisure Time Physical Activity Questionnaire (MLTPA) and dietary macronutrient intake, assessed using 24 h recalls and 3-day food records. Regression analyses included the covariates, protein intake (g/kg), MLTPA, age and sex. For natural logarithm (Ln) of right HGS, LBM (p < 0.001) and % body fat (p < 0.005) were significant (r² = 46.5%; p < 0.000). For left LnHGS, LBM (p < 0.000), age (p = 0.036), protein intake (p = 0.015) and LnMLTPA (p = 0.015) were significant (r² = 0.535; p < 0.000). For SMW, % body fat, age and LnMLTPA were significant (r² = 0.346; p < 0.000). For STS, % body fat and age were significant (r² = 0.251; p < 0.000). LBM is a strong predictor of upper body strength while higher % body fat and lower physical activity are associated with poorer outcomes on tests of lower extremity performance.     

五种布鲁氏菌核酸检测试剂盒检测性能的评价北大核心CSCD

目的比较5种布鲁氏菌核酸实时荧光PCR检测试剂盒的一致性和检出能力,为临床实验室选择检测方法和布鲁氏菌的诊断提供参考依据。方法选用经病原学检测确定为布鲁氏菌阳性的血液样本38份,健康人的血液样本24份,潘氏变形杆菌、溶藻弧菌、河弧菌、铜绿假单胞菌、肺炎克雷伯菌DNA各1份,使用5种试剂盒(编号A-E)分别进行核酸检测,比较5种试剂盒临床样本检测的一致性;选择1份阳性样本核酸用无RNA酶水梯度稀释得到5个浓度(浓度1:4453.13 fg/μL,浓度2:1113.28 fg/μL,浓度3:278.32 fg/μL,浓度4:69.58 fg/μL,浓度5:17.40 fg/μL),每个浓度使用5种试剂盒(编号A-E)分别进行3次检测,比较5种试剂盒的阳性检出率及批内重复性。结果5种试剂盒检测67份DNA样品的符合率稍有不同,试剂盒ABDE的符合率均为100%,试剂盒C的符合率为98.51%。批内重复性显示5种试剂盒在浓度1、浓度2、浓度3水平重复检测DNA的Ct值变异系数均<5%;在浓度1与浓度4梯度区间,试剂盒的阳性检出能力比较显示试剂盒A、B、D较高,为11/12,试剂盒C和E较低,为8/12。结论5种试剂盒的真实性和可靠性较好,灵敏度和符合率稍有差别,特异度均为100%;重复性较好,检测性能良好。部分试剂盒对弱阳性样本的检出能力不强,该类样本可使用多种试剂盒复核,以保障结果的准确性。     

Leptin and total cholesterol are predictors of weight gain in pre-pubertal children

OBJECTIVE: The aim of this study was to identify specifically which biochemical indices predict excessive weight gain over time in a cohort of pre-pubertal children. SUBJECTS: Fifty nine healthy pre-pubertal children (age: 6.3-9.8y). MEASUREMENTS: Children were defined anthropometrically and biochemically at baseline. Height and weight measurements were then repeated after six (n=52) and 12 months (n=37). RESULTS: Weight change after six months (defined by a change in body mass index (BMI) z-score from baseline) demonstrated no correlation with fasting plasma levels of leptin, insulin, insulin:glucose (IG) ratio, cholesterol, triglyceride or high density lipoprotein (HDL) cholesterol. However, after 12 months there was a significant negative correlation between BMI z-score change and initial plasma leptin (r=-0.35, P=0.048) and this relationship strengthened when adjusted for body fat (from bio-electrical impedance; r=-0.46, P=0.009). In addition, there was a significant positive relationship between plasma total cholesterol and BMI z score change (r=0.38, P=0.03) and this relationship remained unchanged when adjusted for body fat. No relationship was observed between weight change after 12 months and plasma levels of insulin, IG ratio, HDL cholesterol or triglyceride. CONCLUSION: Plasma leptin and total cholesterol were found to be predictive of weight gain over 12 months in a cohort of pre-pubertal children. These two potential predictors can be readily measured in clinical practice and these findings may represent a method of defining the 'at risk of obesity' state in childhood.     

Unconventional biological threats and the molecular biological response to biological threats

This article concludes this symposium on potential agents of warfare and terrorism with discussion of 3 topics. First, influenza A virus is discussed as a potential biological weapon. Although it does not receive much attention in this role, the potential for mass casualties and public panic certainly exist if an epidemic of a virulent influenza A virus were initiated. Second, agroterrorism, terrorism directed at livestock or poultry or crops, is briefly discussed. Finally, the potential role of techniques of modern molecular biology to create new agents for bioterrorism or enhance the terrorist potential of available agents, and the known roles of these techniques in defense against biological warfare or terrorism are discussed.