C-Series Coelenterazine-Driven Bioluminescence Signature Imaging

The present study introduces a unique BL signature imaging system with novel CTZ analogues named “C-series.” Nine kinds of C-series CTZ analogues were first synthesized, and BL intensity patterns and spectra were then examined according to the marine luciferases. The results show that the four CTZ analogues named C3, C4, C6, and C7, individually or collectively luminesce with completely distinctive BL spectral signatures and intensity patterns according to the luciferases: Renilla luciferase (RLuc), NanoLuc, and artificial luciferase (ALuc). The signatural reporters were multiplexed into a multi-reporter system comprising RLuc8.6-535SG and ALuc16. The usefulness of the signatural reporters was further determined with a multi-probe system that consists of two single-chain probes embedding RLuc8 and ALuc23. This study is a great addition to the study of conventional bioassays with a unique methodology, and for the specification of each signal in a single- or multi-reporter system using unique BL signatures and patterns of reporter luciferases.     

Highly Diverse Protein Library Based on the Ubiquitous (β/α)8 Enzyme Fold Yields Well‐Structured Proteins through in Vitro Folding Selection

Proper protein folding is a prerequisite for protein stability and enzymatic activity. Although directed evolution can be a powerful tool to investigate enzymatic function and to isolate novel activities, well‐designed libraries of folded proteins are essential. In vitro selection methods are particularly capable of searching for enzymatic activities in libraries of trillions of protein variants, yet high‐quality libraries of well‐folded enzymes with such high diversity are lacking. We describe the construction and detailed characterization of a folding‐enriched protein library based on the ubiquitous (β/α)₈ barrel fold, which is found in five of the six enzyme classes. We introduced seven randomized loops on the catalytic face of the monomeric, thermostable (β/α)₈ barrel of glycerophosphodiester phosphodiesterase (GDPD) from Thermotoga maritima. We employed in vitro folding selection based on protease digestion to enrich intermediate libraries containing three to four randomized loops for folded variants, and then combined them to assemble the final library (10¹⁴ DNA sequences). The resulting library was analyzed by using the in vitro protease assay and an in vivo GFP‐folding assay; it contains ～10¹² soluble monomeric protein variants. We isolated six library members and demonstrated that these proteins are soluble, monomeric and show (β/α)₈‐barrel fold‐like secondary and tertiary structure. The quality of the folding‐enriched library improved up to 50‐fold compared to a control library that was assembled without the folding selection. To the best of our knowledge, this work is the first example of combining the ultra‐high throughput mRNA display method with selection for folding. The resulting (β/α)₈ barrel libraries provide a valuable starting point to study the unique catalytic capabilities of the (β/α)₈ fold, and to isolate novel enzymes.     

Guangdong Metal Transaction Center has Become the Second Largest Precious Metal Transaction Platform Nationwide

正The reporter learned from Guangdong Province Precious Metal Transaction Center that in 2013the Center maintained healthy development momentum,and had become the second largest precious metal transaction platform in China in terms of market size;currently Guangdong is quickening steps of financial innovation for the Center,aiming to build a transaction center with international core competitiveness.     

多维度碳纳米相增强铝基复合材料研究进展

目的: 研究微小RNA（miR-136）通过靶向CD163抑制CD68+M0巨噬细胞向CD68+CD163+M2巨噬细胞极化的作用机制。方法: 采用流式细胞术检测20例肝细胞肝癌患者肿瘤组织及癌旁组织中CD68+CD163+M2巨噬细胞的比例,实时荧光定量PCR（RT-qPCR）检测肿瘤及癌旁组织中miR-136的水平。分离人脾脏中CD68+M0巨噬细胞,将细胞分为对照组（Control）,miRNA模拟物组（miRNA mimic）和miRNA抑制物组（miRNA inhibitor）。IL-4和IL-13 10 ng/mL诱导巨噬细胞M2型极化。采用流式细胞术检测CD68+CD163+M2巨噬细胞比例,Western blot检测细胞中CD163、TNF-α、诱导型一氧化氮合酶（iNOS）的表达以及M2巨噬细胞标志物精氨酸酶（Arg1）的表达,机制研究中检测JAK/STAT信号中蛋白酪氨酸激酶1（JAK1）、信号转导子与转录激活子1（STAT1）和STAT6的水平,PI3K/AKT信号中磷脂酰肌醇-3激酶（PI3K）、丝氨酸-苏氨酸蛋白激酶1（AKT1）的表达。荧光素酶报告基因法确定miR-136与CD163的靶向关系。结果: 肝癌组织中M2型巨噬细胞比例显著高于癌旁组织,而miR-136的表达显著低于癌旁组织,两者表达具有负相关。荧光素酶报告基因结果显示CD163是miR-136的靶基因。miRNA mimic组中CD68+CD163+M2巨噬细胞比例显著低于miRNA inhibitor组,与Control组比较无统计学差异。机制检测中发现miRNA mimic中JAK1、STAT6、PI3K、AKT1低表达,说明miR-136可以间接抑制JAK1-STAT6以及PI3K-AKT1的表达,抑制M2巨噬细胞极化。结论: miR-136可以靶向CD163抑制M2型巨噬细胞极化,其作用机制与JAK1-STAT6以及PI3K-AKT1抑制有关,这是M2巨噬细胞在肝癌免疫中的调节机制之一。     

Molecular Imaging of Human Skeletal Myoblasts (huSKM) in Mouse Post-Infarction Myocardium

Current treatment protocols for myocardial infarction improve the outcome of disease to some extent but do not provide the clue for full regeneration of the heart tissues. An increasing body of evidence has shown that transplantation of cells may lead to some organ recovery. However, the optimal stem cell population has not been yet identified. We would like to propose a novel pro-regenerative treatment for post-infarction heart based on the combination of human skeletal myoblasts (huSkM) and mesenchymal stem cells (MSCs). huSkM native or overexpressing gene coding for Cx43 (huSKMCx43) alone or combined with MSCs were delivered in four cellular therapeutic variants into the healthy and post-infarction heart of mice while using molecular reporter probes. Single-Photon Emission Computed Tomography/Computed Tomography (SPECT/CT) performed right after cell delivery and 24 h later revealed a trend towards an increase in the isotopic uptake in the post-infarction group of animals treated by a combination of huSkMCx43 with MSC. Bioluminescent imaging (BLI) showed the highest increase in firefly luciferase (fluc) signal intensity in post-infarction heart treated with combination of huSkM and MSCs vs. huSkM alone (p < 0.0001). In healthy myocardium, however, nanoluciferase signal (nanoluc) intensity varied markedly between animals treated with stem cell populations either alone or in combinations with the tendency to be simply decreased. Therefore, our observations seem to show that MSCs supported viability, engraftment, and even proliferation of huSkM in the post-infarction heart.     

Central Geological Prospecting Fund Newly Added 16.47 Million Tonnes of Bauxite in 2013

正In 2013,the Central Geological Prospecting Fund(CGPF)Project newly discovered 15 ore fields,including 3 large above ore fields,and 5medium sized ore fields.The reporter learned from the recently published"Annual Report of Central Geological Prospecting Fund Project(2013)"that,from 2006 to 2013 the CGPF     

半滑舌鳎肝脏细胞系的建立与鉴定

以半滑舌鳎肝脏组织为材料,探索了组织细胞分离培养条件,建立了半滑舌鳎组织细胞培养技术及半滑舌鳎肝脏细胞系（HTLC）。该细胞系的培养基是添加了抗生素、胎牛血清（FBS）、花鲈血清（SPS）、成纤维生长因子（bFGF）的MEM。HTLC形态呈纤维状,在培养基中生长迅速,经200多天的培养,成功传代30多代。检测了温度、FBS浓度、bFGF对其生长的影响,结果表明,在24～30℃生长良好,但当温度低于12℃时细胞生长速度明显减慢;在一定浓度范围内,其生长速度随血清浓度的升高而增快,血清浓度过高或过低对其生长不利;在培养基中添加bFGF可以使细胞生长速度显著提高;其二倍体核型为2n=21t。将绿色荧光蛋白（GFP）报告基因通过脂质体介导的方法转入HTLC中并成功地获得了表达,转化率为20％左右。该细胞系的建立为研究鱼类病毒学、免疫学、遗传学、功能基因组学提供了很好的实验材料。