猪PEG1基因多态性及其遗传印记

PEG1基因影响动物胚胎生长及母性行为,多数动物PEG1为父方表达的遗传印记特征,但出生后猪的PEG1印记表达尚不清楚。因此,文章选取长白、大白和蓝塘3个品种共166头纯种猪,在猪的PEG1基因外显子12区域内寻找SNP,采用PCR-SSCP方法对其多态性进行检测和基因频率分析;取带有PEG1基因该位点SNP为杂合的仔猪3头,对其胃、胸腺、胰、脾、肺、肌肉、肝、舌、肾、脑、膀胱、心脏等组织器官和胎盘的mRNA产物分别进行RT-PCR-SSCP分析,结果表明:PEG1基因外显子12存在一个由G突变为A的单核苷酸多态性位点;PEG1的外显子12在3头仔猪的主要组织器官仅表达父亲来源的等位基因,表明猪的PEG1基因呈母方印记、父方表达的遗传特征。     

应用排序分析藓类植物分类群分布与气候因素的关系

以我国21个山地藓类植物区系和气象资料为基础,应用典范对应分析(CCA)和除趋势典范对应分析(DCCA),比较了21个山地中藓类植物61个科,曲尾藓科(Dicranaceae)23个属、曲柄藓属(Campylopus)17种及曲尾藓属(Dicranum)35种分布与年均温度、>10℃积温、1月均温、7月均温、年均相对湿度、年均降雨量、年有雾天数、年有霜天数和年日照时数的关系,通过排序进行了直观的展示;同时还应用典范对应分析直观地反映了包括长白山在内的我国9个山地苔藓植物地理成分组成上的相似性及它们与气候因素间的关系.本文研究表明将DCCA和CCA应用到植物区系地理学研究上是可行的.     

Regulation of TRP channel TRPM2 by the tyrosine phosphatase PTPL1

TRPM2, a member of the transient receptor potential (TRP) superfamily, is a Ca²⁺-permeable channel, which mediates susceptibility to cell death following activation by oxidative stress, TNF, or -amyloid peptide. We determined that TRPM2 is rapidly tyrosine phosphorylated after stimulation with H₂O₂ or TNF. Inhibition of tyrosine phosphorylation with the tyrosine kinase inhibitors genistein or PP2 significantly reduced the increase in [Ca²⁺]_i observed after H₂O₂ or TNF treatment in TRPM2-expressing cells, suggesting that phosphorylation is important in TRPM2 activation. Utilizing a TransSignal PDZ domain array blot to identify proteins which interact with TRPM2, we identified PTPL1 as a potential binding protein. PTPL1 is a widely expressed tyrosine phosphatase, which has a role in cell survival and tumorigenesis. Immunoprecipitation and glutathione-S-transferase pull-down assays confirmed that TRPM2 and PTPL1 interact. To examine the ability of PTPL1 to modulate phosphorylation or activation of TRPM2, PTPL1 was coexpressed with TRPM2 in human embryonic kidney-293T cells. This resulted in significantly reduced TRPM2 tyrosine phosphorylation, and inhibited the rise in [Ca²⁺]_i and the loss of cell viability, which follow H₂O₂ or TNF treatment. Consistent with these findings, reduction in endogenous PTPL1 expression with small interfering RNA resulted in increased TRPM2 tyrosine phosphorylation, a significantly greater rise in [Ca²⁺]_i following H₂O₂ treatment, and enhanced susceptibility to H₂O₂-induced cell death. Endogenous TRPM2 and PTPL1 was associated in U937-ecoR cells, confirming the physiological relevance of this interaction. These data demonstrate that tyrosine phosphorylation of TRPM2 is important in its activation and function and that inhibition of TRPM2 tyrosine phosphorylation reduces Ca²⁺ influx and protects cell viability. They also suggest that modulation of TRPM2 tyrosine phosphorylation is a mechanism through which PTPL1 may mediate resistance to cell death. transient receptor potential channels; oxidative stress     

妊娠相关脑卒中39 例临床分析

目的:探讨妊娠相关脑卒中的发病原因、临床表现、母婴结局、治疗及预防措施。方法:回顾性分析2001年1月-2013年12月我院共收治的妊娠相关脑卒中39例患者的临床资料。结果:39例患者中,发生在妊娠期21例,产褥期18例。经电子计算机断层扫描(CT)、磁共振成像(MRI)、磁共振动脉血管造影(MRA)、磁共振脑静脉血管成像(MRV)、数字减影血管造影术(DSA)和腰椎穿刺等检查明确诊断,诊断出血性脑卒中7例,脑梗死3例,脑静脉窦血栓(CVST)29例,均给予相应的抢救及治疗。16例早期、中期妊娠患者行人工流产术或利凡诺羊膜腔内注射穿刺引产终止妊娠,5例患者行剖宫产终止妊娠。患者治愈出院15例,6例死亡,14例遗留不同程度的肢体活动障碍或语言障碍,家属放弃治疗出院4例。结论:妊娠相关脑卒中危险因素主要包括子痫前期、心源性栓塞、脑血管畸形、脑动脉瘤、水电解质紊乱等代谢障碍性疾病及产褥感染等。其发病急、病死率高,故需提高对本病的认识,定期产前检查,及时发现高危因素,早诊断及时治疗,选择适当的时机及方式终止妊娠是改善妊娠相关脑卒中患者预后的关键。     

Plasmid metagenome reveals high levels of antibiotic resistance genes and mobile genetic elements in activated sludge

The overuse or misuse of antibiotics has accelerated antibiotic resistance, creating a major challenge for the public health in the world. Sewage treatment plants (STPs) are considered as important reservoirs for antibiotic resistance genes (ARGs) and activated sludge characterized with high microbial density and diversity facilitates ARG horizontal gene transfer (HGT) via mobile genetic elements (MGEs). However, little is known regarding the pool of ARGs and MGEs in sludge microbiome. In this study, the transposon aided capture (TRACA) system was employed to isolate novel plasmids from activated sludge of one STP in Hong Kong, China. We also used Illumina Hiseq 2000 high-throughput sequencing and metagenomics analysis to investigate the plasmid metagenome. Two novel plasmids were acquired from the sludge microbiome by using TRACA system and one novel plasmid was identified through metagenomics analysis. Our results revealed high levels of various ARGs as well as MGEs for HGT, including integrons, transposons and plasmids. The application of the TRACA system to isolate novel plasmids from the environmental metagenome, coupled with subsequent high-throughput sequencing and metagenomic analysis, highlighted the prevalence of ARGs and MGEs in microbial community of STPs.     

The ubiquinone-binding site in NADH:ubiquinone oxidoreductase from Escherichia coli

An azido-ubiquinone derivative, 3-azido-2-methyl-5-methoxy[3H]-6-decyl-1,4-benzoquinone ([3H]azido-Q), was used to study the ubiquinone/protein interaction and to identify the ubiquinone-binding site in Escherichia coli NADH:ubiquinone oxidoreductase (complex I). The purified complex I showed no loss of activity after incubation with a 20-fold molar excess of [3H]azido-Q in the dark. Illumination of the incubated sample with long wavelength UV light for 10 min at 0 degrees C caused a 40% decrease of NADH:ubiquinone oxidoreductase activity. SDS-PAGE of the complex labeled with [3H]azido-Q followed by analysis of the radioactivity distribution among the subunits revealed that subunit NuoM was heavily labeled, suggesting that this protein houses the Q-binding site. When the [3H]azido-Q-labeled NuoM was purified from the labeled reductase by means of preparative SDS-PAGE, a 3-azido-2-methyl-5-methoxy-6-decyl-1,4-benzoquinone-linked peptide, with a retention time of 41.4 min, was obtained by high performance liquid chromatography of the protease K digest of the labeled subunit. This peptide had a partial NH2-terminal amino acid sequence of NH2-VMLIAILALV-, which corresponds to amino acid residues 184-193 of NuoM. The secondary structure prediction of NuoM using the Toppred hydropathy analysis showed that the Q-binding peptide overlaps with a proposed Q-binding motif located in the middle of the transmembrane helix 5 toward the cytoplasmic side of the membrane. Using the PHDhtm hydropathy plot, the labeled peptide is located in the transmembrane helix 4 toward the periplasmic side of the membrane.     

TRAIL-R2 (DR5) mediates apoptosis of synovial fibroblasts in rheumatoid arthritis

TRAIL has been proposed as an anti-inflammatory cytokine in animal models of rheumatoid arthritis (RA). Using two agonistic mAbs specific for TRAIL-R1 (DR4) and TRAIL-R2 (DR5), we examined the expression and function of these death receptors in RA synovial fibroblast cells. The synovial tissues and primary synovial fibroblast cells isolated from patients with RA, but not those isolated from patients with osteoarthritis, selectively expressed high levels of cell surface DR5 and were highly susceptible to anti-DR5 Ab (TRA-8)-mediated apoptosis. In contrast, RA synoviocytes did not show increased expression of TRAIL-R1 (DR4), nor was there any difference in expression of Fas between RA and osteoarthritis synovial cells. In vitro TRA-8 induced apoptosis of RA synovial cells and inhibited production of matrix metalloproteinases induced by pro-inflammatory cytokines. In vivo TRA-8 effectively inhibited hypercellularity of a SV40-transformed RA synovial cell line and completely prevented bone erosion and cartilage destruction induced by these cells. These results indicate that increased DR5 expression and susceptibility to DR5-mediated apoptosis are characteristic of the proliferating synovial cells in RA. As highly proliferative transformed-appearing RA synovial cells play a crucial role in bone erosion and cartilage destruction in RA, the specific targeting of DR5 on RA synovial cells with an agonistic anti-DR5 Ab may be a potential therapy for RA.     

P—选择素凝集素—EGF功能域的克隆、表达及其单克隆抗体制备

制备特异性抗人P 选择素的凝集素 表皮生长因子 (L EGF)功能域的单克隆抗体。利用特异引物 ,通过RT PCR从外周血血小板中扩增出人P 选择素的L EGF功能域基因 ,将其克隆至pET42b( )载体中 ,测序验证后转染大肠杆菌BL2 1,经诱导表达了C端融合 6×His的蛋白质。融合蛋白质经分离纯化后 ,免疫Balb/c小鼠 ,应用杂交瘤技术 ,通过间接ELISA筛选阳性克隆。获得 3株可稳定分泌抗L EGF功能域单抗的杂交瘤细胞株 (B10、F3和H5 )。其亚型分别为IgG2 、IgG1和IgG3;轻链均为κ型。所获的单抗对LPS刺激活化的人脐静脉内皮细胞均有特异性结合反应 ,并可在体外阻断经凝血酶激活的血小板与中性粒细胞间的粘附。表明所获的单抗可特异性识别结合天然P 选择素 ,具有体外抗活化血小板与中性粒细胞粘附的功能 ,为进一步应用此单抗进行P 选择素结构和功能及抗粘附治疗研究提供了实验基础。