Changes of the hepatic proteome in murine models for toxically induced fibrogenesis and sclerosing cholangitis

We investigated the changes in the hepatic proteome in murine models for toxic-induced fibrogenesis and sclerosing cholangitis. A comprehensive comparison of protein changes observed is made and the mechanistical basis of the expression changes is discussed. Hepatic fibrosis was induced by repetitive intraperitoneal CCl4 treatment of BALB/c mice or developed spontaneously in BALB/c-ATP-binding cassette, subfamily B, member 4 (Abcb4) knock out mice. Fibrosis was verified by a morphometric score and assessment of hydroxyproline content of liver tissue, respectively. The innovative difference in-gel electrophoresis (DIGE) technique was used to analyse protein expression levels of the mouse proteome. Results were confirmed by Western blotting and real-time RT-PCR. In CCl4-induced fibrosis 20 out of 40 and in BALB/c-Abcb4(-/-) mice 8 out of 28 differentially expressed proteins were identified utilizing DIGE. Only two proteins, selenium-binding protein (Sbp2) and carbonic anhydrase 3, have been unidirectionally expressed (i.e. down-regulated) in both models. Relevant differences in the pathogenesis of toxically induced liver fibrosis and sclerosing cholangitis exist. The only novel protein with regard to liver fibrosis depicting a unidirectional expression pattern in both animal models was Sbp2. An explicit protein function could not be clarified yet.     

Pctaire1 phosphorylates N-ethylmaleimide-sensitive fusion protein: implications in the regulation of its hexamerization and exocytosis

Pctaire1, a member of the cyclin-dependent kinase (Cdk)-related family, has recently been shown to be phosphorylated and regulated by Cdk5/p35. Although Pctaire1 is expressed in both neuronal and non-neuronal cells, its precise functions remain elusive. We performed a yeast two-hybrid screen to identify proteins that interact with Pctaire1. N-Ethylmaleimide-sensitive fusion protein (NSF), a crucial factor in vesicular transport and membrane fusion, was identified as one of the Pctaire1 interacting proteins. We demonstrate that the D2 domain of NSF, which is required for the oligomerization of NSF subunits, binds directly to and is phosphorylated by Pctaire1 on serine 569. Mutation of this phosphorylation site on NSF (S569A) augments its ability to oligomerize. Moreover, inhibition of Pctaire1 activity by transfecting its kinase-dead (KD) mutant into COS-7 cells enhances the self-association of NSF. Interestingly, Pctaire1 associates with NSF and synaptic vesicle-associated proteins in adult rat brain. To investigate whether Pctaire1 phosphorylation of NSF is involved in regulation of Ca(2+)-dependent exocytosis, we examined the effect of expressing Pctaire1 or NSF phosphorylation mutants on the regulated secretion of growth hormone from PC12 cells. Interestingly, expression of either Pctaire1-KD or NSF-S569A in PC12 cells significantly increases high K(+)-stimulated growth hormone release. Taken together, our findings provide the first demonstration that Pctaire1 phosphorylation of NSF regulates the ability of NSF to oligomerize, implicating an unexpected role of this kinase in modulating exocytosis. These findings open a new avenue of research in studying the functional roles of Pctaire1 in the nervous system.     

Prostasin regulates PD-L1 expression in human lung cancer cells

The serine protease prostasin is a negative regulator of lipopolysaccharide-induced inflammation and has a role in the regulation of cellular immunity. Prostasin expression in cancer cells inhibits migration and metastasis, and reduces epithelial–mesenchymal transition. Programmed death-ligand 1 (PD-L1) is a negative regulator of the immune response and its expression in cancer cells interferes with immune surveillance. The aim of the present study was to investigate if prostasin regulates PD-L1 expression. We established sublines overexpressing various forms of prostasin as well as a subline deficient for the prostasin gene from the Calu-3 human lung cancer cells. We report here that PD-L1 expression induced by interferon-γ (IFNγ) is further enhanced in cells overexpressing the wildtype membrane-anchored prostasin. The PD-L1 protein was localized on the cell surface and released into the culture medium in extracellular vesicles (EVs) with the protease-active prostasin. The epidermal growth factor-epidermal growth factor receptor (EGF-EGFR), protein kinase C (PKC), and mitogen-activated protein kinase (MAPK) participated in the prostasin-mediated up-regulation of PD-L1 expression. A Gene Set Enrichment Analysis (GSEA) of patient lung tumors in The Cancer Genome Atlas (TCGA) database revealed that prostasin and PD-L1 regulate common signaling pathways during tumorigenesis and tumor progression.     

七姊妹山自然保护区黄杉林群落学特征

黄杉为国家Ⅱ级保护植物,是我国特有的第三纪孑遗的珍贵树种,主要分布在我国亚热带的中山地带。采用Braun-Blanquet的植物社会学调查方法并结合典型样方法对七姊妹山自然保护区的原生黄杉群落进行调查,并对其群落特征进行了分析。结果表明:在2300 m~2的调查样地,共有维管束植物52种,隶属24科38属,群落物种组成主要以杜鹃花科、豆科、五加科、壳斗科等种类为主;群落生活型组成分析显示,该黄杉群落高位芽植物占绝对优势,高位芽植物、地面芽植物占比分别为76.9%、13.5%,地上芽和地下芽植物较少,植物的生活型谱大致呈"L"型;群落重要值分析表明,黄杉重要值为53.6,占较大优势,是群落建群种,乌冈栎、石韦分别是灌木层及草本层的优势种,其重要值分别为24.0、54.8;该黄杉群落物种多样性指数整体较低,除均匀度指数外,优势度指数、多样性指数、丰富度指数总体表现出灌木层乔木层草本层,上坡山脊的规律;种群年龄结构分析表明,七姊妹山自然保护区黄杉种群年龄结构呈"L"型分布,属于增长型种群,在一定时间内,表明黄杉仍为该群落的优势种群,林窗的出现使该黄杉群落保持持续更新。     

Myostatin三维结构模建及分子进化分析

Myostatin(MST)为肌肉生长负调节因子,其功能受抑制可导致肌肉量增加.对MST核酸序列进行序列比对,构建进化树;采用同源模建方法首次模建MST成熟肽生物活性二聚体的四级结构,并预测MST与其受体ActRIIB的相互作用模式.进化树将肌肉生长抑制素基因(MSTN)分成4个亚家族:哺乳动物MSTN,鸟类MSTN以及鱼类MSTN 1和2.MST受纯化选择作用,在不同物种的直系同源基因具有较高的刚源性,其中哺乳动物、鸟类MST C端活性肽氨基酸序列高度保守.表明哺乳动物、鸟类MST的结构、功能类似,且信号传导路径可能一致;而鱼类MST的调控机制可能存在较大差异.MST结构及其表面静电势和疏水氨基酸分布表明静电力和疏水相互作用在MST与其受体结合过程中可能起到十分重要的作用.     

浙江马鞍列岛海域潮间带底栖海藻分布特征

2007年3—7月对浙江马鞍列岛海域潮间带底栖海藻进行了调查,初步查明该海域潮间带底栖海藻的组成、分布情况和温度属性,并利用相似性指数（S_c）和相对重要性指数（IRI_c）分析了调查海域潮间带底栖海藻的优势种组成.结果表明：调查海域采集到的31种海藻隶属于3门24属,其中绿藻门5属7种,褐藻门5属8种,红藻门14属16种;在波浪和潮汐作用下,潮间带出现局限分布种和选择性分布种,孔石莼、鼠尾藻等在调查岛礁区域广有分布;红藻门种类在调查海域的出现频率为61.1%,为优势门类,绿藻门种类在该海域的总体水平分布基本呈均匀状态;81%的调查种类分布在低潮带,其中包括一些中潮带延伸种类,中、低潮带的海藻组成相似性值为0.47,并且中、低潮带的生境趋同效果大于高、中潮带.马鞍列岛潮间带底栖海藻具有明显的垂直分带现象,温水性种类占绝对优势,优势种多为暖温性种.该海域底栖海藻属于暖温带-亚热带过渡型海洋植物区系.     

Preparation and Efficacy of Newcastle Disease Virus DNA Vaccine Encapsulated in PLGA Nanoparticles

Background

Although the Newcastle disease virus (NDV) inactivated vaccines and attenuated live vaccines have been used to prevent and control Newcastle disease (ND) for years, there are some disadvantages. Recently, newly developed DNA vaccines have the potential to overcome these disadvantages. The low delivery efficiency, however, hindered the application of DNA vaccines for ND in practice.

Methodology/Principal Findings

The eukaryotic expression plasmid pVAX1-F (o) DNA that expressed the F gene of NDV encapsulated in PLGA nanoparticles (pFNDV-PLGA-NPs) were prepared by a double emulsion-solvent evaporation method and optimal preparation conditions of the pFNDV-PLGA-NPs were determined. Under the optimal conditions, the pFNDV-PLGA-NPs were produced in good morphology and had high stability with a mean diameter of 433.5±7.5 nm, with encapsulation efficiency of 91.8±0.3% and a Zeta potential of +2.7 mV. Release assay in vitro showed that the fusion gene plasmid DNA could be sustainably released from the pFNDV-PLGA-NPs up to 93.14% of the total amount. Cell transfection test indicated that the vaccine expressed and maintained its bioactivity. Immunization results showed that better immune responses of SPF chickens immunized with the pFNDV-PLGA-NPs were induced compared to the chickens immunized with the DNA vaccine alone. In addition, the safety of mucosal immunity delivery system of the pFNDV-PLGA-NPs was also tested in an in vitro cytotoxicity assay.

Conclusions/Significance

The pFNDV-PLGA-NPs could induce stronger cellular, humoral, and mucosal immune responses and reached the sustained release effect. These results laid a foundation for further development of vaccines and drugs in PLGA nanoparticles.