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1.
Squalene synthase (SS) catalyzes the biosynthesis of squalene, the first specific intermediate in the cholesterol biosynthetic pathway. To test the feasibility of lowering plasma cholesterol by inhibiting hepatic SS, we generated mice in which SS is specifically knocked out in the liver (L-SSKO) using Cre-loxP technology. Hepatic SS activity of L-SSKO mice was reduced by >90%. In addition, cholesterol biosynthesis in the liver slices was almost eliminated. Although the hepatic squalene contents were markedly reduced in L-SSKO mice, the hepatic contents of cholesterol and its precursors distal to squalene were indistinguishable from those of control mice, indicating the presence of sufficient centripetal flow of cholesterol and/or its precursors from the extrahepatic tissues. L-SSKO mice showed a transient liver dysfunction with moderate hepatomegaly presumably secondary to increased farnesol production. In a fed state, the plasma total cholesterol and triglyceride were significantly reduced in L-SSKO mice, primarily owing to reduced hepatic VLDL secretion. In a fasted state, the hypolipidemic effect was lost. mRNA expression of liver X receptor α target genes was reduced, while that of sterol-regulatory element binding protein 2 target genes was increased. In conclusion, liver-specific ablation of SS inhibits hepatic cholesterol biosynthesis and induces hypolipidemia without increasing significant mortality.  相似文献   
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Recent studies have emphasized the important role of microRNA (miRNA) clusters and common target genes in disease progression. Despite the known involvement of the miR‐192/215 family in many human diseases, its biological role in Hirschsprung disease (HSCR) remains undefined. In this study, we explored the role of the miR‐192/215 family in the pathogenesis of HSCR. Quantitative real‐time PCR and western blotting measured relative expression levels of miRNAs, mRNAs, and proteins in 80 HSCR patients and 77 normal colon tissues. Targets were evaluated by dual‐luciferase reporter assays, and the functional effects of miR‐192/215 on human 293T and SH‐SY5Y cells were detected by the Transwell assay, CCK8 assay and flow cytometry. MiR‐192/215 was significantly down‐regulated in HSCR tissue samples, and their knockdown inhibited cell migration and proliferation in the human 293T and SH‐SY5Y cell lines. Nidogen 1 (NID1) was confirmed as a common target gene of miR‐192/215 by dual‐luciferase reporter gene assay and its expression was inversely correlated with that of miR‐192/215 in tissue samples and cell lines. Silencing of NID1 could rescue the extent of the suppressing effects by miR‐192/215 inhibitor. The down‐regulation of miR‐192/215 may contribute to HSCR development by targeting NID1.

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4.
There is a diverse range of microbiological challenges facing the food, healthcare and clinical sectors. The increasing and pervasive resistance to broad‐spectrum antibiotics and health‐related concerns with many biocidal agents drives research for novel and complementary antimicrobial approaches. Biofilms display increased mechanical and antimicrobial stability and are the subject of extensive research. Cold plasmas (CP) have rapidly evolved as a technology for microbial decontamination, wound healing and cancer treatment, owing to the chemical and bio‐active radicals generated known collectively as reactive oxygen and nitrogen species. This review outlines the basics of CP technology and discusses the interactions with a range of microbiological targets. Advances in mechanistic insights are presented and applications to food and clinical issues are discussed. The possibility of tailoring CP to control specific microbiological challenges is apparent. This review focuses on microbiological issues in relation to food‐ and healthcare‐associated human infections, the role of CP in their elimination and the current status of plasma mechanisms of action.  相似文献   
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Exposure of endothelial cells (ECs) to agents such as oxidized glycerophospholipids (oxGPs) and cytokines, known to accumulate in atherosclerotic lesions, perturbs the expression of hundreds of genes in ECs involved in inflammatory and other biological processes. We hypothesized that microRNAs (miRNAs) are involved in regulating the inflammatory response in human aortic endothelial cells (HAECs) in response to oxGPs and interleukin 1β (IL-1β). Using next-generation sequencing and RT-quantitative PCR, we characterized the profile of expressed miRNAs in HAECs pre- and postexposure to oxGPs. Using this data, we identified miR-21-3p and miR-27a-5p to be induced 3- to 4-fold in response to oxGP and IL-1β treatment compared with control treatment. Transient overexpression of miR-21-3p and miR-27a-5p resulted in the downregulation of 1,253 genes with 922 genes overlapping between the two miRNAs. Gene Ontology functional enrichment analysis predicted that the two miRNAs were involved in the regulation of nuclear factor κB (NF-κB) signaling. Overexpression of these two miRNAs leads to changes in p65 nuclear translocation. Using 3′ untranslated region luciferase assay, we identified 20 genes within the NF-κB signaling cascade as putative targets of miRs-21-3p and -27a-5p, implicating these two miRNAs as modulators of NF-κB signaling in ECs.  相似文献   
7.
Signal peptides are short peptides located at the N-terminus of secreted proteins. They characteristically have three domains; a basic region at the N-terminus (n-region), a central hydrophobic core (h-region) and a carboxy-terminal cleavage region (c-region). Although hundreds of different signal peptides have been identified, it has not been completely understood how their features enable signal peptides to influence protein expression. Antibody-derived signal peptides are often used to prepare recombinant antibodies expressed by eukaryotic cells, especially Chinese hamster ovary (CHO) cells. However, when prokaryotic Escherichia coli (E. coli) are utilized in drug discovery processes, such as for phage display selection or antibody humanization, signal peptides have been selected separately due to the differences in the expression systems between the species. In this study, we successfully established a signal peptide that enables a functional antibody to be expressed in both prokaryotic and eukaryotic cells by focusing on the importance of having an Ala residue in the c-region of the signal sequence. We found that changing Ser to Ala at only two positions significantly augmented the anti-HER2 antigen binding fragment (Fab) expression in E. coli. In addition, this altered signal peptide also retained the ability to express functional anti-HER2 antibody in CHO cells. Taken together, the present findings indicate that the signal peptide can promote functional antibody expression in both prokaryotic E. coli and eukaryotic CHO cells. This finding will contribute to the understanding of signal peptides and accelerate therapeutic antibody research.  相似文献   
8.
目的:构建s TACI-Fc-Myc重组质粒,并进行原核表达和纯化具有生物活性的融合蛋白。方法:通过PCR法获得s TACI-Fc-Myc重组片段,然后把融合基因片段与原核载体p ET28a连接在一起,并构建p ET28a-s TACI-Fc-Myc重组子,并转入BL21(DE3)中进行表达,用蛋白A凝胶亲和层析柱进行纯化及酶联免疫吸附剂(ELISA)法测定其生物学活性。结果:获得了s TACI-Fc-Myc重组质粒,且该质粒可以在BL21(DE3)中表达,亲和层析柱纯化后纯度可达到95%以上,与BAFF的结合活性具有剂量依赖性,浓度达到5 ng/μL时,两者的吸附达到饱和。结论:成功构建了s TACI-Fc-Myc原核表达载体,并使有生物学活性的融合蛋白在BL21(DE3)上获得了稳定表达,为进一步研究并筛选高活性BAFF拮抗肽奠定了基础。  相似文献   
9.
目的:静脉血栓是一种高复发风险和高致死率的疾病,其形成和复发的分子机制尚不明确。基于人类信号网络和基因表达谱数据可针对静脉血栓经华法令抗凝治疗后的复发机制进行研究。方法:结合表达谱数据和人类信号网络,设计差异模块筛选策略,通过功能分析、差异表达分析和已知血栓相关基因及药物靶基因的互作关联研究,获得与静脉血栓复发相关的显著差异模块。结果:最终获得8个与静脉血栓复发密切相关的显著差异模块,评估了华法令治疗静脉血栓的效能,提出了联合用药的3种可能途径。结论:应用本文提出的整合筛选策略,能识别与静脉血栓复发相关的模块,探究静脉血栓复发的分子机制和评估华法令的治疗效能。还提供了潜在的联合用药途径,这对治愈血栓、防治血栓复发及复发机制的研究具有重要意义。  相似文献   
10.
目的:探讨脓毒血症患者血清外周白细胞中的微小RNA(mi RNA)表达水平的变化及其在脓毒症患者中的表达意义以及免疫调控的关系。方法:采用流式细胞仪检测外周血CD4+CD25+Treg细胞表达,采用实时定量PCR(RT-PCR)方法检测110例脓毒症患者以及100例正常对照外周血白细胞中mi RNA以及Foxp3 m RNA表达量,酶联免疫吸附法测定TNF-α和IL-10浓度,序贯器官衰竭估计(SOFA)评分系统评价脓毒症患者的严重程度。对mi RNA与白细胞总数、TNF-α、IL-10和SOFA评分之间的相关性进行分析。结果:实验组mi RNA表达水平较对照组显著降低(P0.01),WBC、IL-10水平显著升高(P0.01)。实验组mi RNA表达水平以及SOFA评分、血清TNF-α和IL-10之间呈负相关关系(r值分别为-0.512、-0.623、-0.432,P0.05);与WBC无显著相关性(r=0.215,P0.05)。脓毒症患者外周血Treg表达率和Foxp3m RNA均显著高于对照组(P0.01);随病情严重而升高,轻、中、重度实验组间两两比较差异均有统计学意义(P0.01)。死亡均在重度脓毒症患者中,死亡组Treg、Foxp3 m RNA及IL-10均显著高于存活组(P0.01);mi RNA低于存活组(P0.01)。结论:脓毒症患者外周血的mi RNA表达量显著降低,表达水平在一定程度上可以反应机体的炎症反应情况,同时还可以判断疾病的严重度,且mi RNA参与对Treg细胞增殖的调节,在脓毒症免疫失衡机制中发挥一定的作用。  相似文献   
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