Restriction analysis of the prototype strain of enteric adenovirus type 41 using exonuclease III.

Enteric adenoviruses 40 and 41 (Ad40 and Ad41) are a prominent cause of gastroenteritis in young children. Diagnosis of these enteric types by conventional means is complicated by their fastidious growth characteristics. Enteric adenovirus growth was enhanced by cocultivation. Typing of enteric isolates currently entails analysis of the extracted viral DNA with restriction enzymes. Restriction endonuclease fragments of the Ad41 strain Tak genome were ordered by (i) double digestion, (ii) release of restriction fragments from plasmids containing 84% of the Ad41 genome in EcoRI fragments A, B and C, (iii) hybridization of Southern blotted Ad41 fragments with EcoRI fragment containing plasmids and various segments of the Ad2 genome, (iv) sequential reduction of the genome beginning with terminal restriction fragments with exonuclease III and S1 nuclease. The termini of adenovirus genomes are difficult to clone and the use of exonuclease III is a useful alternative to conventional restriction mapping. DNA restriction patterns, fragment sizes and restriction maps of the Ad4 1 strain Tak with enzymes BamHI, BglII, ClaI, EcoRI, HindIII, PstI, SalI, SmaI and XhoI are presented. Prototype strain restriction maps should enable better understanding of adenovirus type 41 and its epidemiology.     

Management of Brain Metastases in the New Era of Checkpoint Inhibition

Purpose of the Review

Brain metastasis is a common complication of advanced malignancies, especially, lung cancer, breast cancer, renal cell carcinoma, and melanoma. Traditionally surgery, when indicated, and radiation therapy, either as whole-brain radiation therapy or stereotactic radiosurgery, constituted the major treatment options for brain metastases. Until recently, most of the systemic chemotherapy agents had limited activity for brain metastases. However, with the advent of small molecule tyrosine kinase inhibitors and immunotherapy agents, there has been renewed interest in using these agents in the management of brain metastases.

Recent Findings

Immune checkpoint inhibitors have revolutionized the treatment of metastatic melanoma, lung cancer, kidney cancer, and bladder cancer among others. They modulate the immune system to recognize tumor antigens as “non-self” antigens and mount an immune response against them.

Summary

Initial studies of using immune checkpoint inhibitors in brain metastases have shown promising activity, and several clinical trials are currently underway. Studies are also assessing the combination of radiation therapy and immunotherapy in brain metastases. The results of these ongoing clinical trials have the potential to change the therapeutic paradigm in patients with brain metastases.

     

Expression and purification of functional recombinant epitopes for the platelet antigens, PlA1 and PlA2

The platelet antigens, PlA1 and PlA2, are responsible for most cases of posttransfusion purpura (PTP) and neonatal alloimmune thrombocytopenia (NAIT) in the caucasian population and are determined by two allelic forms of the platelet glycoprotein GPIIIa gene. To study the interaction between these antigens and their respective antibodies, we inserted the sequence that encodes the signal peptide and the N- terminal 66 amino acids of the PlA1 form of GPIIIa into the expression vector pGEX1. To express the PlA2 antigen, nucleotide 196 of the PlA1 coding sequence was mutated to the PlA2 allelic form. When transformed and induced in Escherichia coli, the two constructs produce glutathione S-transferase (GST)/N-terminal GPIIIa fusion proteins, one containing leucine at position 33 (PlA1), the other proline (PlA2). These proteins are easily purified in milligram quantities using glutathione-Sepharose and react specifically with their respective antibodies by immunoblot and enzyme-linked immunosorbent assay. Antigenicity of the PlA1 fusion protein in reduced glutathione increases with time; moreover, the addition of oxidized glutathione accelerates this process, presumably because of formation of the native disulfide bonds. Neutralization assays indicate that the PlA1 fusion protein competes for all of the anti-PlA1 antibody in the serum of patients with PTP and NAIT that is capable of interacting with the surface of intact platelets. This study shows that the GST/N-terminal GPIIIa fusion proteins contain conformational epitopes that mimic those involved in alloimmunization, and that regions other than the amino terminal 66 amino acids of GPIIIa are not likely to contain or be required for the development of functional PlA1 epitopes. Furthermore, these recombinant proteins can be used for the affinity-purification of clinical anti-PlA1 antibodies and specific antibody identification by western blotting, making them useful in the diagnosis of patients alloimmunized to PlA1 alloantigens.     

Targeted Treatment of Brain Metastases

Purpose of Review

Brain metastases are the most common intracranial tumors in adults. Historically, the median survival after the diagnosis of brain metastases has been dismal and medical therapies had a limited role in the management of these patients.

Recent Findings

The advent of targeted therapy has ushered in an era of increased hope for patients with brain metastases. The most common malignancies that result in brain metastases—melanoma, lung cancer, and breast cancer, often have actionable mutations, which make them good candidates for targeted systemic therapy. These brain metastases have been shown to have relevant and sometimes divergent genetic alterations, and there has been a resurgence of interest in targeted drug delivery to the brain by using standard or pulsatile dosing to achieve adequate concentration in the brain.

Summary

An increased understanding of oncogenic alterations, a surge in targeted drug development with good blood barrier penetration, and inclusion of patients with active brain metastases on clinical trials have led to improved outcomes for patients with brain metastases.

     

Tear film profile in Graves' ophthalmopathy.

Tear film profile was studied in 30 patients with Graves' ophthalmopathy. Tear film pH, fluorescein staining, marginal tear strip and Schimer test values in patients with Graves' ophthalmopathy were comparable with controls, indicating normal tear secretion. Tear film break-up-time (BUT) in late Graves' ophthalmopathy was significantly low suggesting unstable tear film. Rose bengal as well as lissamine green staining intensity scores were significantly high, indicating presence of drying epithelial cells in early as well as late Graves' ophthalmopathy patients.     

心肌坏死标记物在检测心肌坏死中的应用与地位

目的:评估各种心肌坏死标记物对判断心肌坏死的价值。资料来源:应用计算机检索LWW全文英文数据库1994-01/2006-08与心肌坏死标记物相关的文献,检索词“Myocardial damage,myocardiolysis,marker”,并限定文章语言种类为“English”。资料选择:对资料进行初审,选对于临床有意义的心肌坏死物进行较详细的总结。选择临床研究的文章,无论有无对照组、是否为随机对照文献均纳入;排除观察对象为动物的基础研究和综述类文献。资料提炼:对收集到的文献进一步查找全文,凡是对于心肌坏死检测快速且可靠的心肌坏死物优先选择,研究内容相似的,以近3年且发表在较权威杂志者优先,最后纳入30篇关于心肌坏死标记物的文章进行综述。资料综合:急性血栓形成及其随后的心肌损伤、坏死是急性心肌梗死的最主要特征,准确、快速、可靠地检测血栓形成和心肌坏死标志物对内科医生诊断急性心肌梗死极具价值。研究表明缺血修饰白蛋白、心肌型脂肪酸结合蛋白、髓过氧化物酶、CK-MB和心肌肌钙蛋白对急性心肌梗死的诊断各具特点,而联合检测更有助于早期、准确诊断急性心肌梗死,为尽早抢救急性心肌梗死创造时机。结论:缺血修饰白蛋白、心肌型脂肪酸结合蛋白、髓过氧化物酶、CK-MB和心肌肌钙蛋白是较敏感的心肌坏死标记物,联合应用诊断价值更大。     

Effect of B-vitamins and n-3 PUFA supplementation for 5 years on blood pressure in patients with CVD

Certain epidemiological and experimental studies suggest that n-3 fatty acids and folate can reduce blood pressure (BP). We investigated the effect of a daily supplementation with dietary doses of B-vitamins or n-3 fatty acids for 5 years on BP in patients with a history of CVD who participated in the Supplémentation en Folates et Omega-3 trial. The patients (n 2501; 1987 men and 514 women) were randomly assigned in a 2 × 2 factorial design to one of four groups: B-vitamins (5-methyl-THF (560 μg); vitamin B? (3 mg) and vitamin B?? (20 μg)) and a placebo capsule for n-3 fatty acids; n-3 fatty acids (600 mg of EPA and DHA at a ratio of 2:1) and a placebo capsule for B-vitamins; both B-vitamins and n-3 fatty acids; or placebo capsules for both treatments. The patients took two capsules daily in a double-blind manner for a median duration of 4·7 years. At baseline and annual examination for 5 years, the patients underwent a clinical examination where BP and clinical and biological parameters were assessed. No effect of supplementation with either n-3 PUFA or B-vitamins on BP was observed in crude and adjusted multivariate models. Change in BP was not associated with change in homocysteine. In conclusion, the present results do not support the routine use of dietary supplements containing B-vitamins, or of n-3 fatty acids, to reduce BP in people with prior CVD.