磷酸钙法对哺乳动物非贴壁细胞的质粒共转染

对一种哺乳动物的非贴壁细胞Bal b/c近交系小鼠肥大细胞瘤细胞株P815进行了G418敏感浓度及甘油耐受性的测定;并通过磷酸钙法用含HBVS基因的质粒pRc/CMVS与含筛选标记基因neo的质粒pSV_2-neo进行了共转染。经G418筛选,获得了G418抗性细胞P851-S,在G418选择压力下,已连续筛选80余天,传代12次;斑点杂交和ELISA实验分别表明P815-S细胞内有HBVS基因的存在,培养上清中有HBsAg的表达。     

Global Establishment Risk of Economically Important Fruit Fly Species (Tephritidae)

The global invasion of Tephritidae (fruit flies) attracts a great deal of attention in the field of plant quarantine and invasion biology because of their economic importance. Predicting which one in hundreds of potential invasive fruit fly species is most likely to establish in a region presents a significant challenge, but can be facilitated using a self organising map (SOM), which is able to analyse species associations to rank large numbers of species simultaneously with an index of establishment. A global presence/absence dataset including 180 economically significant fruit fly species in 118 countries was analysed using a SOM. We compare and contrast ranked lists from six countries selected from each continent, and also show that those countries geographically close were clustered together by the SOM analysis because they have similar fruit fly assemblages. These closely clustered countries therefore represent greater threats to each other as sources of invasive fruit fly species. Finally, we indicate how this SOM method could be utilized as an initial screen to support prioritizing fruit fly species for further research into their potential to invade a region.     

Oral Administered Particulate Yeast-Derived Glucan Promotes Hepatitis B Virus Clearance in a Hydrodynamic Injection Mouse Model

Hepatitis B virus (HBV) persistent infection is associated with ineffective immune response for the clearance of virus. Immunomodulators represent an important class of therapeutics, which potentially could be beneficial for the treatment of HBV infection. The particulate yeast-derived glucan (PYDG) has been shown to enhance the innate and adaptive immune responses. We therefore, assessed the efficacy of PYDG in enhancing HBV specific immune responses by employing the hydrodynamic injection-based (HDI) HBV transfection mouse model. Mice were intragatric administered PYDG daily for 9 weeks post pAAV/HBV1.2 hydrodynamic injection. PYDG treatment significantly promoted HBV DNA clearance and production of HBsAb compared to control mice. PYDG treatment resulted in recruitment of macrophages, dendritic cells (DCs) and effector T cells to the liver microenvironment, accompanied by a significantly augmented DCs maturation and HBV-specific IFN-γ and TNF-α production by T cell. In addition, enhanced production of Th1 cytokines in liver tissue interstitial fluid (TIF) was associated with PYDG administration. Live imaging showed the accumulation of PYDG in the mouse liver. Our results demonstrate that PYDG treatment significantly enhances HBV-specific Th1 immune responses, accompanied by clearance of HBV DNA, and therefore holds promise for further development of therapeutics against chronic hepatitis B.     

Adaptive Tensor-Based Principal Component Analysis for Low-Dose CT Image Denoising

Computed tomography (CT) has a revolutionized diagnostic radiology but involves large radiation doses that directly impact image quality. In this paper, we propose adaptive tensor-based principal component analysis (AT-PCA) algorithm for low-dose CT image denoising. Pixels in the image are presented by their nearby neighbors, and are modeled as a patch. Adaptive searching windows are calculated to find similar patches as training groups for further processing. Tensor-based PCA is used to obtain transformation matrices, and coefficients are sequentially shrunk by the linear minimum mean square error. Reconstructed patches are obtained, and a denoised image is finally achieved by aggregating all of these patches. The experimental results of the standard test image show that the best results are obtained with two denoising rounds according to six quantitative measures. For the experiment on the clinical images, the proposed AT-PCA method can suppress the noise, enhance the edge, and improve the image quality more effectively than NLM and KSVD denoising methods.     

Determination of Zygosity in Adult Chinese Twins Using the 450K Methylation Array versus Questionnaire Data

Previous studies have shown that both single nucleotide polymorphisms (SNPs) and questionnaires-based method can be used for twin zygosity determination, but few validation studies have been conducted using Chinese populations. In the current study, we recruited 192 same sex Chinese adult twin pairs to evaluate the validity of using genetic markers-based method and questionnaire-based method in zygosity determination. We considered the relatedness analysis based on more than 0.6 million SNPs genotyping as the golden standards for zygosity determination. After quality control, qualified twins were left for relatedness analysis based on identical by descent calculation. Then those same sex twin pairs were included in the zygosity questionnaire validation analysis. Logistic regression model was applied to assess the discriminant ability of age, sex and the three questions in zygosity determination. Leave one out cross-validation was used as a measurement of internal validation. The results of zygosity determination based on 65 SNPs in 450k methylation array were all consistent with genotyping. Age, gender, questions of appearance confused by strangers and previously perceived zygosity consisted of the most predictable model with a consistency rate of 0.8698, cross validation predictive error of 0.1347. For twin studies with genotyping and\or 450k methylation array, there would be no need to conduct other zygosity testing for the sake of costs consideration.     

TopBP1 Governs Hematopoietic Stem/Progenitor Cells Survival in Zebrafish Definitive Hematopoiesis

In vertebrate definitive hematopoiesis, nascent hematopoietic stem/progenitor cells (HSPCs) migrate to and reside in proliferative hematopoietic microenvironment for transitory expansion. In this process, well-established DNA damage response pathways are vital to resolve the replication stress, which is deleterious for genome stability and cell survival. However, the detailed mechanism on the response and repair of the replication stress-induced DNA damage during hematopoietic progenitor expansion remains elusive. Here we report that a novel zebrafish mutant^cas003 with nonsense mutation in topbp1 gene encoding topoisomerase II β binding protein 1 (TopBP1) exhibits severe definitive hematopoiesis failure. Homozygous topbp1^cas003 mutants manifest reduced number of HSPCs during definitive hematopoietic cell expansion, without affecting the formation and migration of HSPCs. Moreover, HSPCs in the caudal hematopoietic tissue (an equivalent of the fetal liver in mammals) in topbp1^cas003 mutant embryos are more sensitive to hydroxyurea (HU) treatment. Mechanistically, subcellular mislocalization of TopBP1^cas003 protein results in ATR/Chk1 activation failure and DNA damage accumulation in HSPCs, and eventually induces the p53-dependent apoptosis of HSPCs. Collectively, this study demonstrates a novel and vital role of TopBP1 in the maintenance of HSPCs genome integrity and survival during hematopoietic progenitor expansion.     

Protease‐activated receptor (PAR)‐2 is required for PAR‐1 signalling in pulmonary fibrosis

Idiopathic pulmonary fibrosis is the most devastating diffuse fibrosing lung disease of unknown aetiology. Compelling evidence suggests that both protease‐activated receptor (PAR)‐1 and PAR‐2 participate in the development of pulmonary fibrosis. Previous studies have shown that bleomycin‐induced lung fibrosis is diminished in both PAR‐1 and PAR‐2 deficient mice. We thus have been suggested that combined inactivation of PAR‐1 and PAR‐2 would be more effective in blocking pulmonary fibrosis. Human and murine fibroblasts were stimulated with PAR‐1 and PAR‐2 agonists in the absence or presence of specific PAR‐1 or PAR‐2 antagonists after which fibrotic markers like collagen and smooth muscle actin were analysed by Western blot. Pulmonary fibrosis was induced by intranasal instillation of bleomycin into wild‐type and PAR‐2 deficient mice with or without a specific PAR‐1 antagonist (P1pal‐12). Fibrosis was assessed by hydroxyproline quantification and (immuno)histochemical analysis. We show that specific PAR‐1 and/or PAR‐2 activating proteases induce fibroblast migration, differentiation and extracellular matrix production. Interestingly, however, combined activation of PAR‐1 and PAR‐2 did not show any additive effects on these pro‐fibrotic responses. Strikingly, PAR‐2 deficiency as well as pharmacological PAR‐1 inhibition reduced bleomycin‐induced pulmonary fibrosis to a similar extent. PAR‐1 inhibition in PAR‐2 deficient mice did not further diminish bleomycin‐induced pulmonary fibrosis. Finally, we show that the PAR‐1‐dependent pro‐fibrotic responses are inhibited by the PAR‐2 specific antagonist. Targeting PAR‐1 and PAR‐2 simultaneously is not superior to targeting either receptor alone in bleomycin‐induced pulmonary fibrosis. We postulate that the pro‐fibrotic effects of PAR‐1 require the presence of PAR‐2.