BJ-48, a novel thrombin-like enzyme from the Bothrops jararacussu venom with high selectivity for Arg over Lys in P1: Role of N-glycosylation in thermostability and active site accessibility.

BJ-48, a serine protease from the venom of Bothrops jararacussu, was purified to homogeneity using affinity chromatography on p-aminobenzamidine-agarose followed by HPLC gel filtration. BJ-48 presented 52kDa by SDS-PAGE analysis and 48,036Da by electron spray mass spectrometry. The enzyme was shown to be highly glycosylated with 42% of N-linked carbohydrates composed of Fuc(1):GalN(4):GlcN(5):Gal(1):Man(2) and a high content of sialic acid residues (8-12%). BJ-48 had optimal esterase activity at pH 7.5 and displayed maximum catalytic rate at 50 degrees C. Its hydrolytic activity was strongly inhibited by aprotinin and dithiothreitol while N-tosyl-l-phenylalanine chloromethyl ketone, 6-aminocaproic acid, E-64 and soybean trypsin inhibitor (SBTI) were ineffective. The kinetics of BJ-48 with chromogenic substrates revealed an unprecedented selectivity (10(4)-fold) for Arg over Lys in P1. BJ-48 proved to be a thrombin-like enzyme (TLE) with a specific fibrinogen-clotting activity of 73.4NIH units/mg. The TLE rapidly digested human fibrinogen Bbeta chain, but the Aalpha chain was cleaved specifically to release fibrinopeptide A with k(cat)/K(m)=2.1muM(-1)s(-1). The TLE showed no activity toward other thrombin substrates like protein C, protease-activated receptor-1 or inhibitors such as hirudin and antithrombin. A non-denaturing procedure using PNGase F and neuraminidase followed by hydrophobic interaction chromatography was employed to obtain active BJ-48 forms with variable carbohydrate content. Compared to the native enzyme, total or partially deglycosylated BJ-48 forms presented up to 2-fold reduction in their specific activities upon heating at 55/65 degrees C or treatment with SBTI. These results point out a role for BJ-48 glycosylation in thermostability and controlling the access of some canonical protein inhibitors to the active site.     

Bilateral arthrography of the wrist

In order to determine the true significance of a positive arthrogram in the investigation of wrist pain, a prospective trial was carried out in 60 patients, using the opposite asymptomatic wrist as a control. Of the 46 patients with positive findings in the symptomatic wrist, 34 (74%), had positive arthrograms in the opposite, asymptomatic, wrist. As a result of this, we conclude that a unilateral arthrogram is of little diagnostic value and we recommend the use of the opposite, asymptomatic, wrist as a control.     

Morphologic characterization of osteoblast-like cell cultures isolated from newborn rat calvaria

Summary Two methods for harvesting osteoblast-like cell populations from newborn (10 days) rat calvaria were compared. The first one consisted in culturing the periosteum-free bones and then trypsinizing the cells on the bone surface. The second one involved the migration of the osteoblasts on glass fragments before trypsinization. Since the plating efficiency, the proportion of alkaline phosphatase-positive cells, the population doubling time, and the calcium deposition were more adequate, the second method was used to further characterize the behavior of the cultures. During the first week of culture, the cells featured shapes similar to those observedin vivo on the surface of periosteum-free calvaria. They formed multilayers and, in the presence of ascorbic acid, synthetized an organic matrix containing exclusively type I collagen. Later, small amounts of type III collagen appeared. The cells were embedded in the matrix and progressively acquired the morphologic phenotype of osteocyte-like cells. The matrix mineralized in the presence of β-glycerophosphate. The technique of dropinoculation (high concentration of cells in a small volume of medium) promoted the multilayer formation and the achievement of large mineralized plates (about 1 cm²) in 3 weeks of culture.     

Clinicopathologic analysis of eight cases of pancreatic carcinoid tumors

Carcinoids of the pancreas are exceedingly rare tumors that orieinate from the enterochromaffin cells of the gastroenteropancreatic neuroendocrine system. According to a recent report,1 pancreatic carcinoids are found in only 0.58% (79/13 715 cases) of the entire carcinoid group. Todate, very limited information regarding the detection and diagnosis of this entity has been reported in the available literature. Although pancreatic carcinoid tumors grow.     

多沙唑嗪与特拉唑嗪治疗原发性高血压的疗效比较

目的:比较甲磺酸多沙唑嗪和特拉唑嗪治疗原发性高血压的疗效和安全性.方法:采用随机单盲平行对照试验,将符合条件的轻、中度高血压患者63例随机分为试验组(多沙唑嗪)和对照组(特拉唑嗪).试验剂量从2 mg开始逐渐递增至2～8 mg/d,疗程4周.治疗前后分别行心电图、常规体检、血尿常规和生化检查.结果:①治疗4周后,多沙唑嗪组降压总有效率80.7%,特拉唑嗪组降压总有效率70.0%.两组间无显著性差异.②多沙唑嗪组治疗前后收缩压、舒张压的下降幅度分别为10.81%、10.70%,特拉唑嗪组为10.01%、11.47%.两组间无显著性差异.③多沙唑嗪组有效剂量2～6 mg/d,平均3.04 mg/d,特拉唑嗪组有效剂量2～8 mg/d,平均4.19 mg/d.④每组不良事件发生率相似,多沙唑嗪组25.8%,特拉唑嗪组33.3%(P>0.05).最常见的不良反应为头晕、头痛、心悸、体位性低血压等,多为轻度,可迅速缓解,多沙唑嗪组未发生体位性低血压.结论:甲磺酸多沙唑嗪2～6 mg/d,一天一次口服治疗轻、中度高血压安全有效.     

自身免疫性白内障的晶状体代谢与生理改变研究

目的 :探讨自身免疫性白内障 (AIC)的病理生理学机制。　方法 :采用酶组织化学方法以及晶状体生理功能检查 ,观察AIC模型动物晶状体生理功能与组织内主要酶活性的变化。　结果 :晶状体组织内Na K ATP酶活性降低与晶状体调节力下降、裂隙灯下显微镜混浊改变之间存在着相关性。　结论 :自身免疫性晶状体损伤 ,进而造成晶状体组织内相关酶代谢异常是AIC的病理基础     

海洛因依赖者戒毒中应用护理程序的研究

目的:探讨护理程序对戒毒病人的护理模式,以促进病人积极参与戒毒,减轻戒断症状及改善吸毒导致生理、心理、社会的危害。方法:对应用护理程序工作方法的护理效果进行SPSS11.5统计分析。结果:开展后住院病人意外事件及暴力行为的发生率比开展前显著性降低,实施心理卫生教育后病人对毒品危害性及相关知识认识水平显著提高。结论:通过应用护理程序的工作方法能帮助病人进步且朝向预定目标,为病人走上康复之道打下基础。     

耐药性癫(癎)患者颞叶中细胞周期依赖性蛋白激酶5的表达

目的 研究细胞周期依赖性蛋白激酶5(cyclin dependent kinases 5,CDK5)在耐药性癫(癎)患者颞叶中的表达,探索其在耐药性癫(癎)发病机制中的作用.方法 收集耐药性癫(癎)患者术后脑组织,用荧光定量PCR、免疫组化和Western blot 3种检测方法从基因和蛋白水平分别测定CDK5在耐药性癫(癎)患者颞叶中的表达,并与对照组进行比较.结果 荧光定量PCR发现CDK5 mRAN比对照组明显增加,免疫组化检测显示这种基因的蛋白表达产物主要分布在神经元轴突和胶质细胞中,Western blot检测在相对分子质量35 000处有一蛋白条带,并且可见实验组(颞叶和海马中分别为1.4293±0.1839和2.0733±0.4738)高于对照组(颞叶和海马中分别为0.9680±0.4147和1.403±0.6163,P＜0.05).结论 CDK5在耐药性癫(癎)患者颞叶中表达增强,提示他们可能参与了耐药性癫(癎)的形成.     

Safe anesthesia for endoscopic laryngeal laser surgery

Anesthesia safety for endoscopic laryngeal laser surgery has been a major limiting factor for laser applications in the larynx and the hypopharynx. Several anesthesia techniques have been proposed and each technique appears to have its own limitations. This paper will deal with the distinct advantages offered by the malleable copper tube which is used for delivery of the open Venturi system anesthesia for endoscopic laser surgery. A retrospective study of 100 patients who have undergone this modality of anesthesia at our institution will be presented. Our conclusion from this study shows clearly the superiority of the copper tube over the conventional aluminium-foil-wrapped endotracheal tube in safety and the exposure of the larynx during surgery.