Effects of Mycobacterium bovis on monocyte-derived macrophages from bovine tuberculosis infection and healthy cattle

A PCR-restriction fragment length polymorphism (PCR-RFLP) method for identifying vegetative insecticidal protein (vip) 1-type genes from Bacillus cereus was developed by designing specific primers based on the conserved regions of the genes to amplify vip1-type gene fragments. PCR products were digested with endonuclease AciI, and four known vip1-type genes were identified. Vip1Ac and vip1Aa-type genes appeared in 17 of 26 B. cereus strains. A novel vip1-type gene, vip1Ac1, was identified from B. cereus strain HL12. The vip1Ac1 and vip2Ae3 genes were co-expressed in Escherichia coli strain BL21 by vector pCOLADuet-1. The binary toxin showed activity only against Aphis gossypii (Homoptera), but not for Coleptera (Tenebrio molitor, Holotrichia oblita), Lepidoptera (Spodoptera exigua, Helicoverpa armigera, and Chilo suppressalis), Diptera (Culex quinquefasciatus). The LC(50) of this binary toxin for A. gossypii is 87.5 (34.2-145.3) ng mL(-1) . This is probably only the second report that Vip1 and Vip2 binary toxin shows toxicity against homopteran pests. The PCR-RFLP method developed could be very useful for identifying novel Vip1-Vip2-type binary toxins, and the novel binary toxins, Vip1Ac1 and Vip2Ae3, identified in this study may have applications in biological control of insects, thus avoiding potential problems of resistance.     

Tetrahydroxystilbene glucoside improves learning and (or) memory ability of aged rats and may be connected to the APP pathway

The aim of this study is to evaluate the protective effects of 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG) on learning and (or) memory deficit in aged rats, as well as to explore the possible connection between TSG and the β-amyloid precursor protein (APP) pathway. Sprague-Dawley rats were randomly divided into a young control group (age, 4?months), an aged control group (age, 22?months), and a TSG-treated group (age, 22?months). TSG at doses of 50?mg·kg(-1)·day(-1) was intragastrically administered to 22-month-old rats for 4?weeks. The learning and (or) memory ability was measured using the Morris water maze (MWM) test, and the mRNA and protein expression of APP pathway proteins was measured by real-time polymerase chain reaction (RT-PCR) and Western blot, respectively. The aged rats exhibited obvious learning and (or) memory deficit when compared with the young rats, but TSG treatment significantly improved the learning and (or) memory ability in the aged rats, as noted from the MWM test. RT-PCR and Western blot analysis showed an increase in the expression of beta-site APP cleaving enzyme 1 (BACE1) and A Disintegrin And Metalloproteinase 17 (ADAM17) in aged rats, and a decrease in ADAM10; however, TSG treatment significantly increased the mRNA and protein expression of ADAM10 (p?< 0.01, compared with aged control rats). These results provide solid evidence for the therapeutic effect of TSG on age-related cognitive impairment, especially spatial learning and memory deficit. TSG might exert this effect through the APP pathway, although further studies on the topic are required.     

Ghrelin inhibits insulin resistance induced by glucotoxicity and lipotoxicity in cardiomyocyte

Ghrelin has wide effects on cardiovascular and endocrine system. The aims of this study are to investigate the direct damage effect of high glucose and high palmitate on cardiomyocyte, and to study the effect of ghrelin on insulin resistance induced by glucotoxicity/lipotoxicity in cardiomyocyte and the possible mechanism underlying the cardioprotective activities of ghrelin. The changes of [³H]-2-deoxy-d-glucose (³H-G) intake rates were detected by isotope tracer method and the gene expressions in insulin signal transduction pathway were detected by real-time PCR and Western blot assay. The ³H-G intake rate significantly reduced in high glucose (25 mmol/l) or high palmitate (0.5 mmol/l) treated primary rat ventricular myocytes. After the treatment of ghrelin (10⁻⁷ mol/l), the ³H-G intake rate recovered to the normal level. In addition, the phosphorylation of AKT occurred in 10 min and was the highest in 30 min after the stimulation with ghrelin, which can be blocked by phosphoinositide 3-kinase (PI3K) inhibitor, LY2940002. Ghrelin also increased the mRNA levels of glucose transporter 4 (GLUT4), peroxisome proliferators (PPARr) and AMP activated protein kinase (AMPK) genes in insulin signal transduction pathway. These results indicate that the direct damage of high glucose and high palmitate on cardiomyocyte might be through insulin resistance (IR). Ghrelin can inhibit gluco/lipotoxicity induced insulin resistance by PI3K/AKT pathway. This may provide a clue for therapy for myocardial disease in diabetes mellitus.     

Arylsulfonamidopiperidone derivatives as a novel class of factor Xa inhibitors

The design, synthesis and SAR of a novel class of valerolactam-based arylsulfonamides as potent and selective FXa inhibitors is reported. The arylsulfonamide–valerolactam scaffold was derived based on the proposed bioisosterism to the arylcyanoguanidine-caprolactam core in known FXa inhibitors. The SAR study led to compound 46 as the most potent FXa inhibitor in this series, with an IC₅₀ of 7 nM and EC_2×PT of 1.7 μM. The X-ray structure of compound 40 bound to FXa shows that the sulfonamide–valerolactam scaffold anchors the aryl group in the S1 and the novel acylcytisine pharmacophore in the S4 pockets.     

Abnormal brain default-mode network functional connectivity in drug addicts

Background

The default mode network (DMN) is a set of brain regions that exhibit synchronized low frequency oscillations at resting-state, and is believed to be relevant to attention and self-monitoring. As the anterior cingulate cortex and hippocampus are impaired in drug addiction and meanwhile are parts of the DMN, the present study examined addiction-related alteration of functional connectivity of the DMN.

Methodology

Resting-state functional magnetic resonance imaging data of chronic heroin users (14 males, age: 30.1±5.3 years, range from 22 to 39 years) and non-addicted controls (13 males, age: 29.8±7.2 years, range from 20 to 39 years) were investigated with independent component analysis to address their functional connectivity of the DMN.

Principal Findings

Compared with controls, heroin users showed increased functional connectivity in right hippocampus and decreased functional connectivity in right dorsal anterior cingulate cortex and left caudate in the DMN.

Conclusions

These findings suggest drug addicts'' abnormal functional organization of the DMN, and are discussed as addiction-related abnormally increased memory processing but diminished cognitive control related to attention and self-monitoring, which may underlie the hypersensitivity toward drug related cues but weakened strength of cognitive control in the state of addiction.     

Subtractive hybridization analysis of gastric diseases-associated Helicobacter pylori identifies peptidyl-prolyl isomerase as a potential marker for gastric cancer

Helicobacter pylori, a microaerophilic Gram-negative bacterium, is known to cause chronic gastritis, peptic ulcer and gastric cancer. Genes that are present in certain isolates may determine strain-specific traits such as disease association and drug resistance. In order to understand the pathogenic mechanisms of gastric diseases, identify molecular markers of the diseases associated with H. pylori strains and provide clues for target treatment of H. pylori-related diseases, a subtracted DNA library was constructed from a gastric cancer-associated H. pylori strain and a superficial gastritis-associated H. pylori strain by suppression subtractive hybridization. The presence of gastric cancer-specific genes was identified by dot blot hybridization, DNA sequencing and PCR-based screening. Twelve gastric cancer-specific high-copy genes and nine low-copy genes were found in gastric cancer compared with the superficial gastritis strain. These genes were confirmed by PCR analysis of H. pylori isolates. Notably, peptidyl-prolyl cis-trans isomerase (PPIase) was detected positively in 11 out of 22 (50%) gastric cancer-associated H. pylori strains. In contrast, <24% of the H. pylori strains from superficial gastritis showed positive results. Given the potential role of PPIases in cell growth, apoptosis and oncogenic transformation, our results suggest that PPIase may represent a novel marker and potential therapeutic target for gastric cancer.     

延胡索的化学成分研究(英文)

采用现代分离技术和方法,从延胡索根茎中分离得到17个化合物,通过波谱分析鉴定其结构。包括11个生物碱类化合物,二氢白屈菜红碱(1)、去氢紫堇碱(2)、四氢非洲防己胺(3)、异紫堇球碱(4)、紫堇碱(5)、四氢黄连碱(6)、药根碱(7)、黄连碱(8)、小檗碱(12)、巴马汀(13)和延胡索乙素(14);2个蒽醌类化合物,大黄素(9)和大黄素甲醚(10);1个三萜类化合物,即3β-羟基-齐墩果烷-111,3(18)-二烯-28-酸(11)和3个甾醇类化合物,豆甾醇(15)、β-谷甾醇(16)和胡萝卜苷(17)。其中,化合物9～11首次从该属植物中分离得到,1首次从该种中分离得到。     

Enhanced functional properties of corneal epithelial cells by coculture with embryonic stem cells via the integrin β1-FAK-PI3K/Akt pathway

Adult stem cells are important cell sources in regenerative medicine, but isolating them is technically challenging. This study employed a novel strategy to generate stem-like corneal epithelial cells and promote the functional properties of these cells by coculture with embryonic stem cells. The primary corneal epithelial cells were labelled with GFP and cocultured with embryonic stem cells in a transwell or by direct cell-cell contact. The embryonic stem cells were pre-transfected with HSV-tk-puro plasmids and became sensitive to ganciclovir. After 10 days of coculture, the corneal epithelial cells were isolated by treating the cultures with ganciclovir to kill the embryonic stem cells. The expression of stem cell-associated markers (ABCG2, p63) increased whereas the differentiation mark (Keratin 3) decreased in corneal epithelial cells isolated from the cocultures as evaluated by RT-PCR and flow cytometry. Their functional properties of corneal epithelial cells, including cell adhesion, migration and proliferation, were also enhanced. These cells could regenerate a functional stratified corneal epithelial equivalent but did not form tumors. Integrin β1, phosphorylated focal adhesion kinase and Akt were significantly upregulated in corneal epithelial cells. FAK Inhibitor 14 that suppressed the expression of phosphorylated focal adhesion kinase and Akt inhibited cell adhesion, migration and proliferation. LY294002 that suppressed phosphorylated Akt but not phosphorylated focal adhesion kinase inhibited cell proliferation but had no effect on cell adhesion or migration. These findings demonstrated that the functional properties of stem-like corneal epithelial cells were enhanced by cocultured embryonic stem cells via activation of the integrin β1-FAK-PI3K/Akt signalling pathway.     

Ht31, a protein kinase A anchoring inhibitor, induces robust cholesterol efflux and reverses macrophage foam cell formation through ATP-binding cassette transporter A1

Macrophage foam cell is the predominant cell type in atherosclerotic lesions. Removal of excess cholesterol from macrophages thus offers effective protection against atherosclerosis. Here we report that a protein kinase A (PKA)-anchoring inhibitor, st-Ht31, induces robust cholesterol/phospholipid efflux, and ATP-binding cassette transporter A1 (ABCA1) greatly facilitates this process. Remarkably, we found that st-Ht31 completely reverses foam cell formation, and this process is ABCA1-dependent. The reversal is also accompanied by the restoration of well modulated inflammatory response to LPS. There is no detectable toxicity associated with st-Ht31, even when cells export up to 20% cellular cholesterol per hour. Using FRET-based PKA biosensors in live cells, we provide evidence that st-Ht31 drives cholesterol efflux by elevating PKA activity specifically in the cytoplasm. Furthermore, ABCA1 facilitates st-Ht31 uptake. This allows st-Ht31 to effectively remove cholesterol from ABCA1-expressing cells. We speculate that de-anchoring of PKA offers a novel therapeutic strategy to remove excess cholesterol from lipid-laden lesion macrophages.