Development and Applications of VSV Vectors Based on Cell Tropism

Viral vectors have been available in various fields such as medical and biological research or gene therapy applications. Targeting vectors pseudotyped with distinct viral envelope proteins that influence cell tropism and transfection efficiency are useful tools not only for examining entry mechanisms or cell tropisms but also for vaccine vector development. Vesicular stomatitis virus (VSV) is an excellent candidate for development as a pseudotype vector. A recombinant VSV lacking its own envelope (G) gene has been used to produce a pseudotype or recombinant VSV possessing the envelope proteins of heterologous viruses. These viruses possess a reporter gene instead of a VSV G gene in their genome, and therefore it is easy to evaluate their infectivity in the study of viral entry, including identification of viral receptors. Furthermore, advantage can be taken of a property of the pseudotype VSV, which is competence for single-round infection, in handling many different viruses that are either difficult to amplify in cultured cells or animals or that require specialized containment facilities. Here we describe procedures for producing pseudotype or recombinant VSVs and a few of the more prominent examples from envelope viruses, such as hepatitis C virus, Japanese encephalitis virus, baculovirus, and hemorrhagic fever viruses

Studies on children’s lifestyle circumstances and health (First report)

In this first report we aimed to clarify the factors of life environments which affect children’s health. Our subjects were the guardians of 93 elder (6 years-old) male and female preschoolers of K nursery school in Tokyo met. (abbreviated as KNS) and Y nursery school in Yamaguchi pref. (abbreviated as YNS). Questionnaires regarding the life environments of the preschoolers covered 29 items concerning fundamental life habits including their wake-up time, bedtime, and sleeping period, dietary habits, activity habits, and circumstances of playing. In both preschools more than 80％ of preschoolers live with parents, and the number of preschoolers living with only their mothers was less than 15％, therefore no significant difference of family constitution was observed between YNS and KNS. The average wake-up time of preschoolers attending YNS was significantly earlier (p＜0.01) than that of KNS. The average bedtime of KNS was significantly later (p＜0.001) than that of YNS. The average sleeping period of YNS was significantly longer (p＜0.001) than that of KNS. The means of going to school showed much difference between the two schools. The percentage taken by car was greater in YNS than KNS, while the percentages of being taken by bicycle or walking were greater in KNS than YNS. All means of transportation to school had significant difference of rates (p＜0.05) between YNS and KNS. Results of this study clarified that the preschool located in the city area is characterized by life habits such as short sleeping period, late wake-up time and bedtime, and the preschool located in the mountain area is characterized by longer sleeping period, early wake-up time and bedtime. However, the location of playing spaces did not show significant difference, probably because of the social backgrounds including TV games rather than environmental factors. Considering the health of preschoolers, emphasis on physical activities by playing outside and ensuring sufficient sleep are important factorsdepartmental bulletin pape

Off-pump waffle procedure using an ultrasonic scalpel for constrictive pericarditis †

Abstract Radical pericardiectomy alone is not effective in patients with constrictive pericarditis accompanied by epicardial constriction. However, a waffle procedure is very effective in these patients, as we demonstrate in this paper. Firstly, we performed a radical pericardiectomy via a median sternotomy. We used an ultrasonic scalpel for this procedure, and an apical suction device was useful for lifting the heart. This technique is similar to that used in off-pump coronary artery bypass grafting. After the whole heart was dissected from the markedly thickened parietal pericardium, a waffle procedure was performed on the left ventricle first. Multiple longitudinal and transverse incisions were made on the thickened epicardium using an ultrasonic scalpel. After the left ventricle was fully relieved of constriction, the same procedure was performed on the right ventricle. Thereafter, a remarkable improvement of our patient's haemodynamic status was obtained. The whole procedure can be done without cardiopulmonary bypass. We describe tips for performing the waffle procedure without cardiopulmonary bypass using an ultrasonic scalpel and apical suction device, as well as pitfalls to avoid

Experimental Evaluation of Bretschneider's Solution for Myocardial Preservation in Cardiac Transplantation

Myocardial preservation by Bretschneider's solution (BR solution, Group I) with an intracellular like electrolyte and histidine buffer action was compared with EC-Bi solution (Group II) which has an extracellular like electrolyte and bicarbonate buffer action in the mongrel dogs. The PH and PCO2 of the effluent from the coronary sinus were maintained during myocardial ischemia for 3 hr but the lactate rose gradually in group I. The oxygen and lactate up-take ratio of the myocardium after re-perfusion was satisfactory and LV max dp/dt was also maintained at a high level in group I. Morphologically, myelin figure and mitochondrial deformation were more found in group II on electron microscopy. A donor heart preserved for 3 hr in cold BR solution was transplanted in the left thoracic cavity in four mongrel dogs by the technique of the heterotopic cardiac transplantation. Resuscitation of cardiac pulsation was smooth and maintenance of the systemic circulation after transplantation was possible in every case. From these findings, it might be concluded that myocardial preservation using cold BR solution was useful for cardiac transplantation

Critical Point Mutations for Hepatitis C Virus NS3 Proteinase

AbstractThe hepatitis C virus NS3 proteinase plays an essential role in processing of HCV nonstructural precursor polyprotein. To detect its processing activity, we developed a simpletrans-cleavage assay. Two recombinant plasmids expressing the NS3 proteinase region and a chimeric substrate polyprotein containing the NS5A/5B cleavage site between maltose binding protein and protein A were co-introduced intoEscherichia colicells. The proteinase processed the substrate at the single site during their polyprotein expression. Deletion analysis indicated that the functionally minimal domain of the NS3 proteinase was composed of 146 amino acids, 1059 to 1204. We isolated several cDNA clones encoding the functional domain of the NS3 proteinase from the sera of patients chronically infected with HCV and determined their proteinase activity by thistrans-cleavage assay. Both active and inactive clones existed in the same patients. Comparative sequence analyses of these clones suggested that certain point mutations seemed to be related to the loss of proteolytic activity. This was confirmed by back mutation experiments. Among the critical mutations, Pro-1168 to Thr and Arg-1135 to Gly were intriguing. These amino acids, which are situated near the oxyanion hole, seem to be essential for maintaining the conformation of the active center of the NS3 proteinase

GDP/GTP exchange reaction-stimulating activity of Rabphilin-3A for Rab3A small GTP-binding protein

AbstractRabphilin-3A is a putative target protein for Rab3A, a small GTP-binding protein particularly implicated in neurotransmitter release. Rabphilin-3A interacts more preferentially with GTP-Rab3A than with GDP-Rab3A. Moreover, Rabphilin-3A shows a weak activity to stimulate the GTPase activity of Rab3A and a strong activity to inhibit the Rab3A GTPase-activating protein (GAP)-stimulated GTPase activity of Rab3A. Here, we show that Rabphilin-3A has another activity to stimulate the GDP/GTP exchange reaction of Rab3A. Rabphifin-3A may keep Rab3A continuously in the GTP-bound form by converting again GDP-Rab3A, which may be converted from GTP-Rab3A by Rab3A GAP, to GTP-Rab3A, until the function of Rab3A is accomplished