Short exposure to paclitaxel induces multipolar spindle formation and aneuploidy through promotion of acentrosomal pole assembly

Paclitaxel is a widely used microtubule drug and cancer medicine. Here we report that by short exposure to paclitaxel at a low dose, multipolar spindles were induced in mitotic cells without centrosome amplification. Both TPX2 depletion and Aurora-A overexpression antagonized the multipolarity. Live cell imaging showed that some paclitaxel-treated cells accomplished multipolar cell division and a portion of the daughter cells went on to the next round of mitosis. The surviving cells grew into clones with varied genome content. The results indicated that an aneuploidy population could be induced by short exposure to paclitaxel at a low dose, implicating potential side effects of paclitaxel.     

QCM detection of DNA targets with single-base mutation based on DNA ligase reaction and biocatalyzed deposition amplification

A novel biosensing technique for highly specific identification of gene with single-base mutation is proposed based on the implementation of the DNA ligase reaction and the biocatalyzed deposition of an insoluble product. The target gene mediated deposition of an insoluble precipitate is then transduced by quartz crystal microbalance (QCM) measurements. In this method, the DNA target hybridizes with a capture DNA probe tethered onto the gold electrode and then with a biotinylated allele-specific detection DNA. A ligase reaction is performed to generate the ligation between the capture and the detection probes, provided there is perfect match between the DNA target and the detection probe. Otherwise even when there is an allele mismatch between them, no ligation would take place. After thermal treatment at an elevated temperature, the formed duplex melts apart that merely allows the detection probe perfectly matched with the target to remain on the electrode surface. The presence of the biotinylated allele-matched probe is then detected by the QCM via the binding to streptavidin-peroxide horseradish (SA-HRP), which catalyzes the oxidative precipitation of 3,3-diaminobenzidine (DAB) by H2O2 on the electrode and provides an amplified frequency response. The proposed approach has been successfully implemented for the identification of single-base mutation in -28 site of the beta-thalassemia gene with a detection limit of 0.1 nM, demonstrating that this method provides a highly specific and cost-efficient approach for point mutation detection.     

Cardiac mitochondrial ATP-sensitive potassium channel is activated by nitric oxide in vitro

Previous observations on the activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) by nitric oxide (NO) in myocardial preconditioning were based on indirect evidence. In this study, we have investigated the direct effect of NO on the rat cardiac mitoK(ATP) after reconstitution of the inner mitochondrial membranes into lipid bilayers. We found that the mitoK(ATP) was activated by exogenous NO donor S-nitroso-N-acetyl penicillamine or PAPA NONOate. This activation was inhibited by mitoK(ATP) blockers 5-hydroxydecanoate or glibenclamide. Our observations confirm that NO can directly activate the cardiac mitoK(ATP), which may underlie its contribution to myocardial preconditioning.     

Regulation of alternative splicing of Bcl-x by IL-6, GM-CSF and TPA

The splicing of many alternative exons in the precursor messenger RNA (pre-mRNA) is regulated by extracellular factors but the underlying molecular bases remain unclear. Here we report the differential regulation of Bcl-x pre-mRNA splicing by extracellular factors and their distinct requirements for pre-mRNA elements. In K562 leukemia cells, treatment with interleukin-6 (IL-6) or granulocyte-macrophage colony stimulating factor (GM-CSF) reduced the proportion of the Bcl-xL variant mRNA while treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) had no effect. In U251 glioma cells, however, TPA efficiently increased the Bcl-xL level. These regulations were also seen for a transfected splicing reporter mini-gene. Further analyses of deletion mutants indicate that nucleotides 1-176 of the downstream intron are required for the IL-6 effect, whereas additional nucleotides 177-284 are essential for the GM-CSF effect. As for the TPA effect, only nucleotides 1-76 are required in the downstream intron. Thus, IL-6, GM-CSF and TPA differentially regulate Bcl-x splicing and require specific intronic pre-mRNA sequences for their respective effects.     

Role of lysosomal acid lipase in the intracellular metabolism of LDL-transported dehydroepiandrosterone-fatty acyl esters

Dehydroepiandrosterone-fatty acyl esters (DHEA-FAE) belong to a unique family of naturally occurring hydrophobic steroid hormone derivatives that are transported in circulating lipoproteins and may act as a source of dehydroepiendrosterone (DHEA) and other biologically active steroid hormones in cells. Here, we studied the metabolic fate of low-density lipoprotein-associated [(3)H]DHEA-FAE ([(3)H]DHEA-FAE-LDL) and the possible role of lysosomal acid lipase (LAL) in the hydrolysis of DHEA-FAE in cultured human cells. When HeLa cells were incubated with [(3)H]DHEA-FAE-LDL, the accumulation of label in the cellular fraction increased with incubation time and could be inhibited by excess unlabeled LDL, suggesting LDL receptor or LDL receptor-related receptor-dependent uptake. During 48 h of chase, decreasing amounts of [(3)H]DHEA-FAE were found in the cellular fraction, while in the medium increasing amounts of unesterified [(3)H]DHEA and its two metabolites, [(3)H]-5alpha-androstanedione (5alpha-adione) and [(3)H]androstenedione (4-adione), appeared. As LDL-cholesteryl ester hydrolysis is dependent on LAL activity, we depleted LAL from HeLa cells using small interfering RNAs and compared the hydrolysis of [(3)H]DHEA-FAE-LDL and [(3)H]cholesteryl-FAE-LDL. The results demonstrated a more modest but significant reducing effect on the hydrolysis of [(3)H]DHEA-FAE compared with [(3)H]cholesteryl-FAE. Moreover, experiments in LAL-deficient human fibroblasts (Wolman disease patient cells) showed that [(3)H]DHEA-FAE hydrolysis was not completely dependent on LAL activity. In summary, LDL-transported [(3)H]DHEA-FAE entered cells via LDL receptor or LDL receptor-related receptor-mediated uptake, followed by intracellular hydrolysis and further metabolism into 5alpha-adione and 4-adione that were excreted from cells. Although LAL contributed to the deesterification of DHEA-FAE, it was not solely responsible for the hydrolysis.     

A Raman Study of the Time Variability for the A-to-B Transition in Wet-spun Films of Calf-thymus DNA

Synopsis

The time evolution of the A-to-B transition has been monitored by Raman spectroscopy and found to vary significantly for different samples. Though all samples were prepared in the identical fashion, some samples completed the transition on the time scale of several hours while other samples took days. The shortest time required for the A conformation to disappear was about 2 hours, as determined by the disappearance of the A-form Raman band at 807 cm^?1. For these fastest transforming samples, the B-form Raman band at 835 cm^?1 was clearly evident after about 5 hours. These data are consistent with the hypothesis that the A conformation of DNA is stabilized by intermolecular interactions.     

卵巢上皮癌中ING4 基因启动子的甲基化状态及其临床意义

目的：探讨卵巢上皮癌中ING4 基因启动子的甲基化状态及其临床意义。方法：收集2005 年7 月至2012 年6 月哈尔滨医科 大学附属第一医院行全面分期手术并经病理检查确诊的150 例卵巢上皮癌组织标本,并以同期因子宫肌瘤或子宫腺肌症行子宫 全切除术或次全切除术并经病理检查确诊为正常卵巢组织的150 例标本作为对照组。采用甲基化特异性PCR（MSP）技术检测卵 巢上皮癌组织与正常卵巢组织中ING4 基因启动子的甲基化状态,蛋白印迹法检测ING4 蛋白的表达,并分析ING4 基因启动子 的甲基化状态与卵巢上皮癌临床病例特征的关系。结果：卵巢上皮癌组织中ING4 基因启动子的甲基化阳性率为42.7%（64/150）, 明显高于正常卵巢组织（4%,6/150）,差异有统计学意义（P<0.05）。ING4 基因启动子甲基化阳性的卵巢上皮癌组织中ING4蛋白 表达阴性或弱阳性;ING4 基因启动子甲基化阴性的卵巢上皮癌和正常卵巢组织中ING4 蛋白表达阳性;在64 例ING4 基因启动 子甲基化的卵巢上皮癌组织中,ING4 蛋白表达强度与ING4 基因启动子的甲基化程度呈负相关（r=-0.435,P<0.05）。卵巢上皮癌 组织中,ING4 基因甲基化的阳性率随着手术病理分期和组织学分级的增加而增加（P<0.05）;卵巢透明细胞癌（55.6%,10/18）和卵 巢子宫内膜样癌（59.3%,16/27）中ING4 基因甲基化的阳性率显著高于浆液性囊腺癌（33.9%,20/59）和粘液性囊腺癌（39.1%, 18/46）（P<0.05）;ING4基因启动子的甲基化状态与患者的年龄、有无腹水及淋巴结转移均无显著相关性（P>0.05）。结论：ING4 基 因启动子的甲基化可能促进了其在卵巢上皮癌组织中的表达失活,进而促进了卵巢上皮癌的生长和分化。     

RAGE mediates amyloid-beta peptide transport across the blood-brain barrier and accumulation in brain

Amyloid-beta peptide (Abeta) interacts with the vasculature to influence Abeta levels in the brain and cerebral blood flow, providing a means of amplifying the Abeta-induced cellular stress underlying neuronal dysfunction and dementia. Systemic Abeta infusion and studies in genetically manipulated mice show that Abeta interaction with receptor for advanced glycation end products (RAGE)-bearing cells in the vessel wall results in transport of Abeta across the blood-brain barrier (BBB) and expression of proinflammatory cytokines and endothelin-1 (ET-1), the latter mediating Abeta-induced vasoconstriction. Inhibition of RAGE-ligand interaction suppresses accumulation of Abeta in brain parenchyma in a mouse transgenic model. These findings suggest that vascular RAGE is a target for inhibiting pathogenic consequences of Abeta-vascular interactions, including development of cerebral amyloidosis.     

西南岩溶地区黄荆和檵木叶片结构对其生态环境的响应

应用常规石蜡切片法对生长于桂林毛村岩溶区和非岩溶区的黄荆(Vitex negundo)和檵木(Loropetalumchinense)的解剖特征进行了比较研究,并对两区的黄荆叶片表皮形态进行了扫描电镜观察.结果显示:(1)两地的黄荆叶片背面均有浓密的绒毛,但致密程度有差异,岩溶区黄荆叶片的气孔深藏于绒毛间隙,这种结构可减少水分蒸发,降低因岩溶干旱带来的水分缺失.(2)岩溶区黄荆和檵木的叶片厚度、上下表皮厚度、栅栏组织的厚度以及栅栏组织的致密程度均大于非岩溶区,这些特征有利于减少水分蒸腾.(3)岩溶区黄荆和檵木叶片的维管组织发达程度高于非岩溶区,有利于在蒸腾减小的情况下促进水分运输和营养元素的迁移,说明2种植物叶片结构特征在不同生境区的改变是其长期在岩溶区干旱环境条件下形成的适应性变化.