Regulation of PKM2 and Nrf2-ARE Pathway during Benzoquinone Induced Oxidative Stress in Yolk Sac Hematopoietic Stem Cells

Benzene is an occupational toxicant and an environmental pollutant that is able to induce the production of reactive oxygen species (ROS), causing oxidative stress and damages of the macromolecules in target cells, such as the hematopoietic stem cells. We had previously found that embryonic yolk sac hematopoietic stem cells (YS-HSCs) are more sensitive to benzene toxicity than the adult bone marrow hematopoietic stem cells, and that nuclear factor-erythroid-2-related factor 2 (Nrf2) is the major regulator of cytoprotective responses to oxidative stress. In the present report, we investigated the effect of PKM2 and Nrf2-ARE pathway on the cellular antioxidant response to oxidative stress induced by benzene metabolite benzoquinone (BQ) in YS-HSC isolated from embryonic yolk sac and enriched by magnetic-activated cell sorting (MACS). Treatment of the YS-HSC with various concentrations of BQ for 6 hours induces ROS generation in a dose-dependent manner. Additional tests showed that BQ is also capable of inducing expression of NADPH oxidase1 (NOX1), and several other antioxidant enzymes or drug-metabolizing enzymes, including heme oxygenase 1 (HMOX1), superoxide dismutase (SOD), catalase and NAD(P)H dehydrogenase quinone 1 (NQO1). Concomitantly, only the expression of PKM2 protein was decreased by the treatment of BQ but not the PKM2 mRNA, which suggested that BQ may induce PKM2 degradation. Pretreatment of the cells with antioxidant N-acetylcysteine (NAC) decreased ROS generation and prevented BQ-induced PKM2 degradation, suggesting involvement of ROS in the PKM2 protein degradation in cellular response to BQ. These findings suggest that BQ is a potent inducer of ROS generation and the subsequent antioxidant responses of the YS-HSC. The accumulated ROS may attenuate the expression of PKM2, a key regulator of the pyruvate metabolism and glycolysis.     

An efficient molecular docking method for adsorbent screening

In this study, with flavonol glycosides (FG) and terpene lactones (TL) in ginkgo biloba extract (GBE) as the targets for separation, we investigated the effectiveness of molecular docking in adsorbent screening. Several polyamine-modified methyl acylate-co-divinylbenzene (MA-co-DVB) adsorbent models were built, and their affinity to rutin, quercetin and ginkgolide B (GB) was evaluated via molecular docking. The model of ethylenediamine-modified adsorbent showed the largest difference in affinity between to GB and to quercetin as well as rutin, and thus this adsorbent could have the best separation performance. The results of the subsequently conducted static adsorption and dynamic adsorption experiments correlated well with docking results. Finally, using ethylenediamine-modified MA-co-DVB adsorbent, nearly complete separation of the FG and TL in GBE was simply achieved by one step of adsorption-desorption. Thus, the reported molecular docking method is expected to be helpful for rapid adsorbent screening.     

鸡β-防御素-1真核表达载体构建及其在Flp-In-293细胞中的表达

以防御素为研究对象探讨其药用开发价值已是国内外生物医学研究领域的热点之一,鸡β-防御素-1 (Gallinacin-1,Gal-1)对许多致病菌都具有杀菌作用,具有很好的研究开发价值.本研究以含有Gal-1成熟肽全长cDNA的重组质粒pMD19-T-Gal-1为模板,通过PCR在Gal-1成熟肽基因C末端引入6×His-tag,并将其克隆到pcDNA3.1 (+)载体中,构建真核表达质粒pcDNA3.1 (+)-Gal-1-His;经脂质体介导转染Flp-In-293细胞,48 h后间接免疫荧光分析,结果重组质粒pcDNA3.1 (+)-Gal-1-His转染的Flp-In-293细胞中有绿色荧光,而阴性对照中无荧光,表明Gal-1-His在Flp-In-293细胞中得到了表达.本研究结果将为进一步研究Gal-1的功能奠定基础.     

甘蓝型油菜雌性不育突变体FS-M1乳突细胞的细胞学观察

雌性不育突变体FS-M1是从甘蓝型油菜(Brassica napus)品种宁油10号中发现的.为了从细胞学角度研究FS-M1的雌性不育机理,利用荧光显微镜、扫描和透射电子显微镜观察分析了FS-M1柱头乳突细胞的授粉行为和超微结构.结果表明:花粉粒能在FS-M1乳突细胞上附着和萌发形成花粉管,但花粉管无法穿越柱头乳突细胞;开花后的FS-M1乳突细胞迅速衰退而呈干瘪萎蔫状,在衰退过程中,FS-M1柱头乳突细胞的细胞器数量减少,细胞液泡化明显,高尔基体、内质网和线粒体等一些细胞器结构被逐渐破坏.因此,推测FS-M1的雌性不育性是由于柱头乳突细胞发育异常造成的.     

乳杆菌抑制金黄色葡萄球菌对HeLa细胞的粘附

对弯曲乳杆菌Lactobacillus crispatus T79-3和T90-1、詹氏乳杆菌Lactobacillus jensenii T118-3和T231-1四株乳杆菌对金黄色葡萄球菌生长的抑制效果以及抑菌成分进行了分析,比较乳杆菌排除、竞争、置换3种不同作用方式对金黄色葡萄球菌粘附HeLa细胞的抑制作用。结果表明4株乳杆菌皆能抑制金黄色葡萄球菌的生长及其粘附HeLa细胞的能力,分析发现4株乳杆菌发挥抑制作用的主要成分是有机酸,同时比较分析乳杆菌3种不同作用方式发现它们对金黄色葡萄球菌粘附HeLa细胞的抑制效果不同,其中,排除作用方式效果最好。另外,乳杆菌对金黄色葡萄球菌粘附HeLa细胞的抑制作用具有浓度依赖性,随着乳杆菌浓度增大,抑制作用增强并逐渐达到饱和。4株乳杆菌中,T79-3粘附能力最强,对金黄色葡萄球菌的抑制作用最强,排除作用方式抑制金黄色葡萄球菌粘附HeLa细胞作用效果较好,提示乳杆菌T79-3有可能作为益生菌防治妇女泌尿生殖道感染。     

Reverse regulation of soluble receptor for advanced glycation end products and proinflammatory factor resistin and S100A12 in Kawasaki disease

Introduction

Kawasaki disease (KD), an acute febrile disease, characterized by systemic vasculitis, predominantly affects infants and children under 5 years of age. Coronary artery lesions (CALs) are its most critical complication, and the etiology remains unknown yet. In order to explore the value of resistin, S100A12 and soluble receptor for advanced glycation end products (sRAGE) in the pathophysiology of KD, we studied the serum levels of resistin, S100A12 and sRAGE in different stages of KD.

Methods

Serum levels of resistin, S100A12 and sRAGE were measured by enzyme-linked immunosorbent assay (ELISA) method in 15 healthy children and 40 KD patients at acute, afebrile and subacute stage.

Results

The resistin and S100A12 levels, including the ratio of resistin to sRAGE and S100A12 to sRAGE increased significantly in the acute stage, and decreased progressively in the afebrile and subacute stage. However, the sRAGE levels decreased significantly in the acute stage, and increased progressively in the afebrile and subacute stage. In the acute, afebrile and subacute stage, the resistin levels were higher in intravenous immunoglobulin (IVIG) non-responders (0.64 ± 0.30, 0.48 ± 0.35, 0.28 ± 0.19, × 10² ng/ml) than in IVIG responders (0.35 ± 0.24, 0.21 ± 0.19, 0.12 ± 0.05, × 10² ng/ml). In the acute and subacute stage, the S100A12 levels were higher in IVIG non-responders (7.92 ± 2.61, 4.98 ± 4.75, × 10² ng/ml) than in IVIG responders (5.05 ± 3.22, 2.35 ± 2.26, × 10² ng/ml). In the afebrile and subacute stage, the sRAGE levels were lower in IVIG non-responders (3.51 ± 2.64, 3.65 ± 3.27, × 10² pg/ml) than in IVIG responders (6.00 ± 2.78, 7.19 ± 2.88, × 10² pg/ml). The resistin levels were positively correlated with S100A12 levels. The sRAGE levels were negatively related with S100A12 and resistin levels.

Conclusions

Resistin, S100A12 and sRAGE are involved in the pathophysiology of KD.     

Structural and functional analysis of the antiparallel strands in the lumenal loop of the major light-harvesting chlorophyll a/b complex of photosystem II (LHCIIb) by site-directed mutagenesis

The light-harvesting chlorophyll a/b-binding protein of photosystem II (LHCIIb) fulfills multiple functions, such as light harvesting and energy dissipation under different illuminations. The crystal structure of LHCIIb at the near atomic resolution reveals an antiparallel strands structure in the lumenal loop between the transmembrane helices B/C. To study the structural and functional significances of this structure, three amino acids (Val-119, His-120, and Ser-123) in this region have been exchanged to Phe, Leu, and Gly, respectively, and the influence of the mutagenesis on the structure and function of LHCIIb has been investigated. The results are as follows. 1) Circular dichroism spectra of the mutations reveals that the antiparallel strands in the lumenal region are very important for adjusting pigment conformation in the neoxanthin domain of LHCIIb. Although the mutagenesis causes only a slight loss of the Neo binding in the complexes (V119F, 0.09; S123G, 0.19; and H120L, 0.27), it imparts remarkable changes to the pigment conformation. 2) Substituting Ser-123 with Gly results in a higher susceptibility to photodamage, an increased tendency to aggregate, and enhanced fluorescence quenching induced by the medium acidification. These results demonstrate that this antiparallel strands domain plays an important role in regulating the pigment conformation and in adjusting the aggregation and the fluorescence yield of LHCIIb.     

Specific expression of a β-tubulin gene (GhTub1) in developing cotton fibers Specific expression of a b-tubulin gene (GhTub1) in developing cotton fibers

Eukaryotic microtubular cytoskeleton plays crucial roles in various cellular processes, in-cluding cell division, polar cell expansion and cell differentiation[1, 2]. The basic filament-forming unit of microtubules is a heterodimer protein composed of a-tubulin and b-tubulin polypeptides. In the higher plants studied, both a- and b-tubulin genes form multigene families. While some of these tubulin genes are expressed constitutively, many others exhibit tissue-, organ- or cell type-specific exp…