不同环境温度对有机磷杀虫剂毒死蜱引起大鼠体温变化的影响

目的: 观察不同环境温度对有机磷杀虫剂毒死蜱(CHP)引起大鼠体温变化的影响.方法: 用数字体温仪测量大鼠的结肠和尾部皮肤温度,观察25℃、16℃和32℃环境温度对口饲CHP引起结肠温度和尾部皮肤温度变化的影响.结果: 给置于25℃和16℃环境中的大鼠口饲CHP (20 mg·kg-1) 后,均可引起结肠温度明显的降低,尾部皮肤温度明显升高;但32℃环境中大鼠口饲CHP后却引起了明显的升温作用,而尾部皮肤温度无明显影响.结论: CHP对大鼠体温的影响与环境温度有关.     

人肺鳞癌组织的血清蛋白质组学的比较分析

采用以肿瘤免疫学与蛋白质组学(proteomics)研究技术有机地结合为基础的血清蛋白质组学研究体系(serologicproteomeanalysis ,SERPA)筛选肺癌分子标志物.对10例人肺鳞癌组织,应用双向凝胶电泳(two dimensionalelectrophoresis ,2 DE)技术对同一肺鳞癌组织的细胞总蛋白同时进行电泳后获得3张相同的凝胶,其中一块2 DE凝胶经银染显色作为平行胶,其余两块2 DE凝胶经电转膜将凝胶中的蛋白质转至硝酸纤维素(NC)膜上,然后分别与肺癌患者的自身血清以及正常对照血清进行Western印迹分析,获取Western印迹反应图谱.经计算机图像分析识别差异反应的蛋白质,然后与平行胶比较找出相应的差异反应蛋白质点.获得了分辨率较高的人肺鳞癌组织与患者的自身血清以及正常对照血清的Western印迹反应图谱;图像分析共识别36±8个差异反应的蛋白质;在平行胶上找到了匹配的差异反应蛋白质点.对2 0个差异蛋白质点进行了肽质指纹图分析,鉴定出14个与细胞生长增殖、细胞代谢、细胞周期调控、信号转导等有关的肺鳞癌相关抗原.通过血清蛋白质组技术对肺鳞癌组织进行的研究,建立了分辨率较高的人肺鳞癌组织与患者的自身血清以及正常血清的Western印迹反应图谱,成功鉴定14个肺鳞癌相关抗原,为进一步筛选用于肺鳞癌诊断、治疗和预后评估     

Regulation of polymorphonuclear leukocyte apoptosis: role of lung endothelium-epithelium bilayer transmigration

Delayed polymorphonuclear leukocyte (PMN) apoptosis exacerbates acute lung injury. To reach the alveolar spaces, PMNs must migrate across both pulmonary endothelial and epithelial cell layers. We hypothesized that transmigration across the endothelium-epithelium bilayer suppresses PMN apoptosis and sought to elucidate the underlying mechanisms. PMNs freshly isolated from normal volunteers were allowed to migrate across polycarbonate membranes alone or membranes coated with a bilayer of human lung endothelial and epithelial cells. After migration toward different chemoattractants (IL-8, formyl-Met-Leu-Phe, or leukotriene B(4)), PMN apoptosis and caspase activities were assessed by annexin V, histology, and enzymatic assays, respectively. Messenger RNA and specific protein expression in three receptor ligand-mediated, apoptosis-inducing pathways (Fas, TNF-alpha, and TNF-related apoptosis-inducing ligand) were further examined by gene array, RT-PCR, flow cytometry, and Western blot analyses. The data demonstrated that transbilayer migration suppressed PMN apoptosis, and this effect was not chemoattractant type specific. Kinetic analyses further showed that the delay of apoptosis was sustained to at least 18 h. Transbilayer migration caused significant decreases in caspase (-3, -8, and -9) activities. The changes in apoptosis-related gene expression support the survival role of transbilayer migration. Furthermore, the reduced apoptosis was correlated with downregulation of Fas ligand and TNF receptor 1 expression. Our data reveal that migration across a lung endothelium-epithelium bilayer suppresses PMN apoptosis. The decreased activity and/or expression of proapoptotic proteins may provide possible targets for the regulation of inappropriate delay in PMN apoptosis during lung inflammation and injury.     

Cellular signals activating muscle proteolysis in chronic kidney disease: a two-stage process

Muscle atrophy is a prominent feature of catabolic conditions and in animal models of these conditions there is accelerated muscle proteolysis that is dependent on the ubiquitin-proteasome system. However, ubiquitin system cannot degrade actomyosin or myofibrils even though it rapidly degrades actin or myosin. We identified caspase-3 as the initial and potentially rate-limiting proteolytic step that cleaves actomyosin/myofibrils. In rodent models of catabolic conditions, we find that caspase-3 is activated to cleave muscle proteins and actomyosin to fragments that are rapidly degraded by the ubiquitin system. This initial proteolytic step in muscle can be recognized because it leaves a footprint of a characteristic 14-kDa actin band. Stimulation of caspase-3 activity depends on activation of phosphatidylinositol 3-kinase. When we suppressed this enzyme in muscle cells, protein breakdown increased as did the expression of caspase-3. In addition, there was increased expression of E3-ubiquitin-conjugating enzymes that are involved in muscle proteolysis, atrogin-1/MAFbx and MuRF1. Thus, when phosphatidylinositol 3-kinase activity is low in muscle cells or rat muscle, both caspase-3 and the ubiquitin-proteasome system are stimulated to degrade protein. Additional investigations will be needed to define the cell signaling processes that activate muscle proteolysis in uremia and catabolic conditions.     

Langmuir monolayer approaches to protein recognition through molecular imprinting

Surface plasmon resonance (SPR) spectroscopy and atomic force microscopy (AFM) have been employed to investigate ferritin adsorption to binary surfactant monolayers of cationic dioctadecyldimethylammonium bromide (DOMA) and non-ionic methyl stearate (SME). Surfactant molar ratios, miscibility, and lateral mobility were controlled to define the number, size, and distribution of "binding sites" for ferritin, which under the low ionic strength conditions investigated, adsorbed to the monolayers predominantly through electrostatic interactions. Successive adsorption/desorption cycles revealed that fluid monolayers, capable of laterally restructuring during the initial protein adsorption event, bound up to 60% more ferritin (dependent on SME:DOMA ratios) as compared to monolayers that were immobilized on a hydrophobic support during this first adsorption step. The enhanced binding of ferritin to fluid monolayers was accentuated in films having non-ionic SME as the principal component. These findings support the premise that the surfactants reorganize to form favorable interactions with an adsorbing protein, leading to protein specific charge patterns, or templates, in the films. Template assessment, however, was complicated by the presence of an irreversibly bound protein fraction, which AFM revealed to be locally ordered protein clusters.     

Application of response surface methodology in medium optimization for spore production of <Emphasis Type="Italic">Coniothyrium minitans</Emphasis> in solid-state fermentation

Summary Spore production of Coniothyrium minitans was optimized by using response surface methodology (RSM), which is a powerful mathematical approach widely applied in the optimization of fermentation process. In the first step of optimization, with Plackett–Burman design, soluble starch, urea and KH²PO⁴ were found to be the important factors affecting C. minitans spore production significantly. In the second step, a 2³ full factorial central composite design and RSM were applied to determine the optimal concentration of each significant variable. A second-order polynomial was determined by the multiple regression analysis of the experimental data. The optimum values for the critical components for the maximum were obtained as follows: soluble starch 0.643 (36.43 g. l⁻¹), urea −0.544 (3.91 g l⁻¹) and KH²PO⁴ 0.049 (1.02 g l⁻¹) with a predicted value of maximum spore production of 9.94 × 10⁹ spores/g IDM. Under the optimal conditions, the practical spore production was 1.04 × 10¹⁰ spores/g IDM. The determination coefficient (R²) was 0.923, which ensure an adequate credibility of the model.     

New dual inhibitors of EGFR and HER2 protein tyrosine kinases

A novel series of dual EGFR and HER2 inhibitors based on the pyrrolo[2,1-f][1,2,4]triazine nucleus is described. A general route toward their synthesis, which enables functionalization at multiple sites, has been developed. Biological evaluation in enzymatic and cell-based assays has identified a series of C-6 carbamates with potent biochemical and cellular activities.     

Synthesis and SAR of diazepine and thiazepine TACE and MMP inhibitors

Potent and selective TACE and MMP inhibitors utilizing the diazepine and thiazepine ring systems were synthesized and evaluated for biological activity in in vitro and in vivo models of TNF-alpha release. Oral activity in the mouse LPS model of TNF-alpha release was seen. Efficacy in the mouse collagen induced arthritis model was achieved with diazepine 20.     

The ligand-dependent interaction of mineralocorticoid receptor with coactivator and corepressor peptides suggests multiple activation mechanisms

We investigated the coregulator (coactivator and corepressor) interactions with the mineralocorticoid receptor (MR) that lead to activation and inhibition of the receptor in the presence of agonist and/or antagonist. Our results indicate that MR ligand binding domain (LBD) interacts strongly with only a few specific coactivator peptides in the presence of the agonist aldosterone and that these interactions are blocked by the antagonist eplerenone. We also discovered that cortisol, the preferred physiological ligand for the glucocorticoid receptor in humans, is a partial MR agonist/antagonist, providing a possible molecular explanation of the tissue-selective effects of glucocorticoids on MR. However, when we examined the coactivator and corepressor peptide interactions in the presence of cortisol, we found that MR bound with cortisol or aldosterone interacted with the same set of peptides. Thus, the partial agonism shown by cortisol is unlikely to be the result of differential interaction with known coactivators and corepressors. On the other hand, we have identified coactivator binding groove mutations that are critical for cortisol activation but not for aldosterone activation, suggesting that the two steroids induce different MR LBD conformations. In addition, we also show that cortisol becomes full agonist when S810L mutation is introduced in the LBD of MR. Interestingly, MR antagonists, such as eplerenone and progesterone, become partial agonist/antagonist of S810L but are still able to recruit LXXLL peptides to the mutant receptor. Together, these findings suggest a model to explain the MR activation by various ligands.     

Synthesis and anti-viral activity of a series of d- and l-2'-deoxy-2'-fluororibonucleosides in the subgenomic HCV replicon system

Based on the discovery of (2'R)-d-2'-deoxy-2'-fluorocytidine as a potent anti-hepatitis C virus (HCV) agent, a series of d- and l-2'-deoxy-2'-fluororibonucleosides with modifications at 5- and/or 4-positions were synthesized and evaluated for their in vitro activity against HCV and bovine viral diarrhea virus (BVDV). The key step in the synthesis, the introduction of 2'-fluoro group, was achieved by either fluorination of 2,2'-anhydronucleosides with hydrogen fluoride-pyridine or potassium fluoride, or a fluorination of arabinonucleosides with DAST. Among the 27 analogues synthesized, only the 5-fluoro compound, namely (2'R)-d-2'-deoxy-2',5-difluorocytidine (13), demonstrated potent anti-HCV activity and toxicity to ribosomal RNA. The replacement of the 4-amino group with a thiol group resulted in the loss of activity, while the 4-methylthio substituted analogue (25) exhibited inhibition of ribosomal RNA. As N(4)-hydroxycytidine (NHC) had previously shown potent anti-HCV activity, we combined the two functionalities of the N(4)-hydroxyl and the 2'-fluoro into one molecule, resulting (2'R)-d-2'-deoxy-2'-fluoro-N(4)-hydroxycytidine (23). However, this nucleoside showed neither anti-HCV activity nor toxicity. All the l-forms of the analogues were devoid of anti-HCV activity. None of the compounds showed anti-BVDV activity, suggesting that the BVDV system cannot always predict anti-HCV activity.