Ultrasonic Elastography Research Based on a Multicenter Study: Adding Strain Ratio after 5-Point Scoring Evaluation or Not

Background

This study aimed to confirm whether strain ratio should be added after evaluation of lesions with 5-point elasticity scoring for differentiating benign and malignant breast lesions on ultrasonographic elastography(UE).

Materials and Methods

From June 2010 to March 2012, 1080 consecutive female patients with breast lesions were recruited into a multicenter retrospective study, which involved 8 centers across China. Each institutional ethic review board approved the study, and all the patients gave written informed consent. All the patients underwent the UE procedure and the strain ratios were calculated and the final diagnosis was made by histological findings. The sensitivity, specificity, accuracy, PPV and NPV were calculated for each of the two evaluation systems and the areas under the ROC curve were compared.

Results

The strain ratios of benign lesions (mean, 2.6±2.0) and malignant lesions (mean,7.9±5.8) were significantly different (p <0.01). When the cutoff point was 3.01, strain ratio method had 79.8% sensitivity, 82.8% specificity, and 81.3% accuracy, while the 5-point scoring method had 93.1% sensitivity, 73.0% specificity, and 76.8% accuracy. The areas under the ROC curve with the strain ratio method and 5-point scoring method were 0.863 and 0.865, respectively(p>0.05). The strain ratio method shows better a diagnosis performance of the lesions with elasticity score 3 and 4.

Conclusions

Although the two UE methods have similar diagnostic performance, separate calculation of the strain ratios seems compulsory, especially for the large solid breast lesions and the lesions with elasticity score 3 and 4.     

Dynamic 1H NMR-based extracellular metabonomic analysis of oligodendroglia cells infected with herpes simplex virus type 1

Herpes simplex virus type 1 (HSV-1) is a large, neurotropic, double-stranded DNA virus that establishes a lifelong latent infection in neurons and glial cells. Previous studies reveal that several metabolic perturbations are associated with HSV-1 infection. However, the extracellular metabolic alterations associated with HSV-1 infection have not been systematically profiled in human cells. Here, a proton nuclear magnetic resonance-based metabonomic approach was applied to differentiate the extracellular metabonomic profiles of HSV-1 infected human oligodendroglia cells (n = 18) and matched control cells (n = 18) at three time points (12, 24, and 36 h post-infection). Resulting spectra were analyzed by chemometric and statistical methods. Metabonomic profiling revealed perturbations in 21 extracellular metabolites. Partial least squares discriminant analysis demonstrated that the whole metabolic patterns enabled statistical discrimination between HSV-1 infected human oligodendroglia cells and control cells. Eight extracellular metabolites, seven of which were amino acids, were primarily responsible for score plot discrimination between HSV-1 infected human oligodendroglia cells and control cells at 36 h post-infection: alanine, glycine, isoleucine, leucine, glutamate, glutamine, histidine, and lactate. HSV-1 infection alters amino acid metabolism in human oligodendroglia cells cultured in vitro. HSV-1 infection may disturb these host cellular pathways to support viral replication. Through elucidating the extracellular metabolic changes incident to HSV-1 infection, this study also provides future directions for investigation into the pathogenic mechanism of HSV-1.     

Induction of multiple cytotoxic T lymphocyte responses in mice by a multiepitope DNA vaccine against dengue virus serotype 1

Dengue virus (DENV) is still a major threat to human health in most tropical and subtropical countries and regions. In the present study, a multi‐epitope DNA vaccine that encodes 15 immunogenic and conserved HLA‐A*0201‐, HLA‐A*1101‐, HLA‐A*2402‐restricted CTL epitopes from DENV serotype 1 (DENV‐1) was constructed based on the eukaryotic expressing plasmid pcDNA^TM3.1/myc‐His(?) A. Immunization of HLA‐A*0201, HLA‐A*1101 and HLA‐A*2402 transgenic mice with the recombinant plasmid pcDNA^TM3.1/myc‐His(?) A‐DENV‐1‐Meg resulted in significantly greater IFN‐γ‐secreting T‐cell responses against most (14/15) CTL epitopes than occurred in mice immunized with the empty plasmid pcDNA^TM3.1/myc‐His(?) A. Additionally, the epitope‐specific T cells directed to some epitopes secreted not only IFN‐γ but also IL‐6 and/or TNF‐α. Finally, the induced epitope‐specific T cells also efficiently lysed epitope‐pulsed splenocytes and DENV‐1‐infected splenic monocytes. The present study confirms the immunogenicity of multi‐epitope DENV vaccine, suggesting that it may contribute to the development of a universal DENV vaccine.

     

UV-B辐射方向对白三叶克隆整合的影响

增强UV-B辐射会对植物生长和生理生化过程产生有害效应。克隆植物中,相连的克隆分株对经常共享资源和激素,然而鲜有关于异质性UV-B辐射下UV-B辐射方向对克隆整合的影响及克隆植物形态结构变化的报道。模拟同质(克隆分株片段均处于自然背景辐射)和异质(克隆分株一端处于自然背景辐射,另一端处于补加的UV-B辐射)UV-B辐射,以克隆植物白三叶为材料,进行连接和隔断处理,研究UV-B辐射方向对克隆整合强度变化、叶片形态结构特化及生理可塑性的影响。结果表明:异质性UV-B辐射下,15N同位素标记端保留的15N百分比高于同质UV-B辐射处理,转移到无标记相连端的15N含量则降低,紫外辐射处理和同位素标记是否处于同一分株端对结果无显著性影响,说明克隆植物白三叶生理整合存在但整合强度降低,辐射方向与克隆整合强度无关;隔断处理组气孔长度增加,栅栏组织增厚,但连接处理组却无此变化,表明生理整合在白三叶叶片形态结构特化中发挥作用。UV-B辐射下,最小荧光、电子传递速率及光化学淬灭系数降低但非光化学淬灭系数升高,而生理整合却使结果相反;叶绿素和紫外吸收物可在异质性UV-B辐射相连的两端运输分享。以上均表明异质UV-B辐射环境中,UV-B辐射胁迫端克隆分株通过生理整合从非胁迫端获益,并以此提高胁迫环境中克隆植物对资源的利用效率。     

Somatic Embryogenesis and in vitro Flowering in <Emphasis Type="Italic">Saposhnikovia divaricata</Emphasis>

Efficient somatic embryogenesis (SE) and in vitro flowering and fruiting were achieved in Saposhnikovia divaricata (Turcz.) Schischk. Friable embryogenic callus developed from the root, internode, and leaf explants on Murashige and Skoog medium (MS) with 2.26 μM 2,4-dichlorophenoxyacetic acid (2,4-D), and subsequently developed into somatic embryos on MS medium containing 4–5% sucrose, 1.74 μM naphthaleneacetic acid (NAA), 4.44 μM 6-benzylaminopurine (BA), and 1.90 μM abscisic acid (ABA). Then the mature embryos were separated and transferred onto MS with 3% sucrose and 0.6% agar for further development and conversion to plantlets. In vitro flowering and fruiting were obtained when the subcultures were carried out for over 15 months. Paclobutrazol (PP333) or ethephon (ETH) at low levels promoted flowering significantly. Also, abnormal rootless somatic embryos of S. divaricata could form flowers and fruits in vitro.     

Coexistence of nitrifiers, denitrifiers and Anammox bacteria in a sequencing batch biofilm reactor as revealed by PCR-DGGE

Aims:  The bacterial diversity in a sequencing batch biofilm reactor (SBBR) treating landfill leachate was studied to explain the mechanism of nitrogen removal.
Methods and Results:  The total microbial DNA was extracted from samples collected from landfill leachate and biofilm of the reactor with the removal efficiencies of NH₄⁺-N higher than 97% and that of chemical oxygen demand (determined by K₂Cr₂O₇, COD_Cr) higher than 86%. Denaturing gradient gel electrophoresis (DGGE) fingerprints based on total community 16S rRNA genes were analyzed with statistical methods, and excised DNA bands were sequenced. The results of phylogenetic analyses revealed high diversity within the SBBR biofilm community, and DGGE banding patterns showed that the community structure in the biofilm remained stable during the running period.
Conclusions:  A coexistence of nitrifiers, including ammonia-oxidizing bacteria and nitrite-oxidizing bacteria, denitrifiers, including aerobic or anaerobic denitrifying bacteria and Anammox bacteria were detected, which might be the real matter of high removal efficiencies of NH₄⁺-N and COD_Cr in the reactor.
Significance and Impact of the Study:  The findings in this study indicated that PCR-DGGE analysis could be used for microbial community detection as prior method, and the SBBR technique could provide preferable growing environment for bacteria with N removal function.     

Genetic diversity and its relationship to hybrid performance and heterosis in rice as revealed by PCR-based markers

Ten elite inbred lines (four japonica, six indica), chosen from those widely used in the hybrid rice breeding program at Human Hybrid Rice Research Center in China, were crossed to produce all possible hybrids excluding reciprocals. The 45 F₁ hybrids along with the ten parents were evaluated for eight traits of agronomic importance, including yield potential, in a replicated field trial. The ten parents were analyzed with 100 arbitrary decamer oligonucleotide primers and 22 microsatellite (simple sequence repeats, SSRs) primer sets via polymerase chain reaction (PCR). Out of the 100 random primers used, 74 were informative and amplified 202 non-redundant bands (variants) with a mean of 2.73 bands per polymorphic primer. All 22 microsatellite primer sets representing 23 loci in the rice genome showed polymorphisms among the ten parents and revealed 90 alleles with an average of 3.91 per SSR locus. Cluster analysis based on Nei's genetic distance calculated from the 291 (202 RAPDs, 89 SSRs) non-redundant variants separated the ten parental lines into two major groups that corresponds to indica and japonica subspecies, which is consistent with the pedigree information. Strong heterosis was observed in hybrids for most of the traits examined. For the 43 diallel crosses (excluding 2 crosses not heading), yield potential, its components (including panicles per plant, spikelets per panicle and 1000-grain weight) and their heterosis in F₁ hybrids showed a significant positive correlation with genetic distance. When separate analyses were performed for the three subsets, yield potential and its heterosis showed significant positive correlations with genetic distance for the 15 indica x indica crosses and the 6 japonica x japonica crosses; however, yield potential and its heterosis were not correlated with genetic distance for the 22 indica x japonica crosses. Results indicated that genetic distance measures based on RAPDs and SSRs may be useful for predicting yield potential and heterosis of intra-subspecific hybrids, but not inter-subspecies hybrids.     

Optimized production of high-titer recombinant adeno-associated virus in roller bottles

Adeno-associated viral (AAV) vectors are used for in vivo gene transfer in a number of preclinical models of genetic diseases (including large-animal models) and are currently being tested in clinical trials for treatment of hemophilia B and cystic fibrosis. Protocols for production of AAV vectors in a helper virus-free system are available and are based on transient transfection of HEK-293 cells with multiple plasmids. Scale-up of vector production has been labor intensive and inefficient because of a lack of larger culture vessels suitable for growth of adherent cells, large-scale transfection, and vector production. Here we report efficient production of AAV vector in roller bottles, which represents a 10-fold scale-up from the conventional flask or plate method. Optimized production yielded greater than 10(13) vector genomes per bottle and was as cost effective as published protocols using plates. Successful vector production by this method was dependent on optimization of transfection by calcium phosphate precipitation, of monitoring of cell growth (by measurement of glucose consumption), of cell culture conditions, and CO2/air exchange with the culture vessel.     

Kinetic FP-TDI assay for SNP allele frequency determination

Strategies for identifying genetic risk factors in complex diseases by association studies require the comparison of allele frequencies of numerous SNPs between affected and control populations. Theoretically, hundreds of thousands of SNP markers across the genome will have to be genotyped in these studies. Genotyping SNPs one sample at a time is extremely costly and time consuming. To streamline whole genome association studies, some have proposed to screen SNPs by pooling the DNA samples initially for allele frequency determination and perform individual genotyping only when there is a significant discrepancy in allele frequencies between the affected and control populations. Here we describe a new method for determining the allele frequency of SNPs in pooled DNA samples using a two-color primer extension assay with real-time monitoring of fluorescence polarization (named kinetic FP-TDI assay). By comparing the ratio of the rate of incorporation of the two allele-specific dye-terminators, one can calculate the relative amounts of each allele in the pooled sample. The accuracy of allele frequency determination with pooled samples is within 3.3 +/- 0.8% of that determined by genotyping individual samples that make up the pool.