A Genetic Screen Using the PiggyBac Transposon in Haploid Cells Identifies Parp1 as a Mediator of Olaparib Toxicity

Genetic perturbation screens have the potential to dissect a wide range of cellular phenotypes. Such screens have historically been difficult in diploid mammalian cells. The recent derivation of haploid embryonic stem cells provides an opportunity to cause loss of function mutants with a random mutagen in a mammalian cell with a normal genetic background. We describe an approach to genetic screens that exploits the highly active piggyBac transposon in haploid mammalian cells. As an example of haploid transposon (HTP) screening, we apply this approach to identifying determinants of cancer drug toxicity and resistance. In a screen for 6-thioguanine resistance we recovered components of the DNA mismatch repair pathway, a known requirement for toxicity. In a further screen for resistance to the clinical poly(ADP-ribose) polymerase (PARP) inhibitor olaparib we recovered multiple Parp1 mutants. Our results show that olaparib toxicity to normal cells is mediated predominantly via Parp1, and suggest that the clinical side effects of olaparib may be on target. The transposon mutant libraries are stable and can be readily reused to screen other drugs. The screening protocol described has several advantages over other methods such as RNA interference: it is rapid and low cost, and mutations can be easily reverted to establish causality.     

Helicobacter pylori cag Pathogenicity Island's Role in B7-H1 Induction and Immune Evasion

During Helicobacter pylori (H. pylori) infection CD4⁺ T cells in the gastric lamina propria are hyporesponsive and polarized by Th1/Th17 cell responses controlled by T_reg cells. We have previously shown that H. pylori upregulates B7-H1 expression on GEC, which, in turn, suppress T cell proliferation, effector function, and induce T_reg cells in vitro. In this study, we investigated the underlying mechanisms and the functional relevance of B7-H1 induction by H. pylori infection to chronic infection. Using H. pylori wild type (WT), cag pathogenicity island (cag PAI^-) and cagA ^- isogenic mutant strains we demonstrated that H. pylori requires its type 4 secretion system (T4SS) as well as its effector protein CagA and peptidoglycan (PG) fragments for B7-H1 upregulation on GEC. Our study also showed that H. pylori uses the p38 MAPK pathway to upregulate B7-H1 expression in GEC. In vivo confirmation was obtained when infection of C57BL/6 mice with H. pylori PMSS1 strain, which has a functional T4SS delivery system, but not with H. pylori SS1 strain lacking a functional T4SS, led to a strong upregulation of B7-H1 expression in the gastric mucosa, increased bacterial load, induction of T_reg cells in the stomach, increased IL-10 in the serum. Interestingly, B7-H1^-/- mice showed less T_reg cells and reduced bacterial loads after infection. These studies demonstrate how H. pylori T4SS components activate the p38 MAPK pathway, upregulate B7-H1 expression by GEC, and cause T_reg cell induction; thus, contribute to establishing a persistent infection characteristic of H. pylori.     

Fabrication and Characteristics of Reduced Graphene Oxide Produced with Different Green Reductants

There has been an upsurge of green reductants for the preparation of graphene materials taking consideration of human health and the environment in recent years. In this paper, reduced graphene oxides (RGOs) were prepared by chemical reduction of graphene oxide (GO) with three green reductants, L-ascorbic acid (L-AA), D-glucose (D-GLC) and tea polyphenol (TP), and comparatively characterized by X-ray photoelectron spectroscopy (XPS), Fourier transform infrared (FTIR) spectra, Raman spectra and electrical conductivity analysis. Results showed that all these three reductants were effective to remove oxygen-containing functional groups in GO and restore the electrical conductivity of the obtained RGO. The RGO sample with L-ascorbic acid as a reductant and reduced with the existence of ammonia had the highest electrical conductivity (9.8 S·cm^-1) among all the obtained RGO samples. The mechanisms regarding to the reduction of GO and the dispersion of RGO in water were also proposed. It is the good dispersibility of reduced graphene oxide in water that will facilitate its further use in composite materials and conductive ink.     

HMGB1 enhances embryonic neural stem cell proliferation by activating the MAPK signaling pathway

Neural stem cells (NSCs) are involved in neural tube formation. As the high-mobility group box 1 (HMGB1) protein is involved in neurulation and is present at elevated levels in neural tube defects (NTDs) induced by hyperthermia, we have now investigated the effects of HMGB1 on proliferation, differentiation, and MAPK signaling pathways of NSCs in vitro. We constructed a lentivirus vector with HMGB1 siRNA and used it to infect NSCs. Down-regulation of HMGB1 expression was confirmed. Proliferation of NSCs was determined by MTS and nestin/BrdU double-labeling. Differentiation of NSCs was assessed using β-tubulinIII and GFAP. Knockdown of HMGB1 significantly suppressed NSC proliferation but hardly affected differentiation, which was regulated by decreased expression of MAPK signaling pathways. Thus, HMGB1 has beneficial effects on neurulation and may serve as a new target for the prevention of NTDs.     

The Variation of Root Exudates from the Hyperaccumulator Sedum alfredii under Cadmium Stress: Metabonomics Analysis

Hydroponic experiments were conducted to investigate the variation of root exudates from the hyperaccumulator Sedum alfredii under the stress of cadmium (Cd). S. alfredii was cultured for 4 days in the nutrient solution spiked with CdCl₂ at concentrations of 0, 5, 10, 40, and 400 µM Cd after the pre-culture. The root exudates were collected and analyzed by GC-MS, and 62 compounds were identified. Of these compounds, the orthogonal partial least-squares discrimination analysis (OPLS-DA) showed that there were a distinct difference among the root exudates with different Cd treatments and 20 compounds resulting in this difference were found out. Changing tendencies in the relative content of these 20 compounds under the different Cd treatments were analyzed. These results indicated that trehalose, erythritol, naphthalene, d-pinitol and n-octacosane might be closely related to the Cd stabilization, phosphoric acid, tetradecanoic acid, oxalic acid, threonic acid and glycine could be attributed to the Cd mobilization, and mannitol, oleic acid, 3-hydroxybutanoic acid, fructose, octacosanol and ribitol could copy well with the Cd stress.     

Optimal Design of River Monitoring Network in Taizihe River by Matter Element Analysis

The objective of this study is to optimize the river monitoring network in Taizihe River, Northeast China. The situation of the network and water characteristics were studied in this work. During this study, water samples were collected once a month during January 2009 - December 2010 from seventeen sites. Futhermore, the 16 monitoring indexes were analyzed in the field and laboratory. The pH value of surface water sample was found to be in the range of 6.83 to 9.31, and the average concentrations of NH₄⁺-N, chemical oxygen demand (COD), volatile phenol and total phosphorus (TP) were found decreasing significantly. The water quality of the river has been improved from 2009 to 2010. Through the calculation of the data availability and the correlation between adjacent sections, it was found that the present monitoring network was inefficient as well as the optimization was indispensable. In order to improve the situation, the matter element analysis and gravity distance were applied in the optimization of river monitoring network, which were proved to be a useful method to optimize river quality monitoring network. The amount of monitoring sections were cut from 17 to 13 for the monitoring network was more cost-effective after being optimized. The results of this study could be used in developing effective management strategies to improve the environmental quality of Taizihe River. Also, the results show that the proposed model can be effectively used for the optimal design of monitoring networks in river systems.     

长江口滨岸带河蚬的时空分布特征及其指示作用

以长江口滨岸湿地生态系统习见的大型底栖动物河蚬为对象,研究了河蚬种群密度、生物量的时空分布特征,分析了河蚬及环境中悬浮颗粒物和沉积物中的重金属含量.结果表明,不同季节、不同采样点以及相同采样点的不同断面河蚬的分布都有差异.河蚬种群密度、生物量的季节分布趋势是春、秋两季＞夏季;其中,浒浦河蚬的年度平均种群密度与平均生物量最高;河蚬在崇明中潮滩的种群密度、生物量高于低潮滩和高潮滩.研究表明,河蚬对沉积物中的重金属元素Cu、Zn富集能力强,对Pb、Cr的富集能力弱,其中,河蚬体内Pb含量与沉积物中Pb含量呈显著负相关(R=-0.924,P<0.01).