Platelet adhesion to vascular endothelial cells is a pathophysiologically relevant cell-to-cell interaction. However, the mechanisms underlying this cellular interaction are incompletely understood. In search of the ligand for CD226 adhesion molecule expressed on platelets, we found that human umbilical vein endothelial cells (HUVEC) express significant amount of putative CD226 ligand. We demonstrated that thrombin-activated, but not resting, platelets bind to intact HUVEC. Anti-CD226 monoclonal antibody specifically inhibited the binding, indicating that CD226 mediates the intercellular binding between thrombin-activated platelets and HUVEC. We also demonstrated that platelet activation with thrombin induces tyrosine phosphorylation of CD226 as well as CD226-mediated platelet adhesion. Moreover, experiments using mutant transfectants suggested that the tyrosine at residue 322 of CD226 plays an important role for its adhesive function. CD226 was also expressed on primary megakaryocytes and megakaryocytic cell lines. Anti-CD226 monoclonal antibody inhibited binding of megakaryocytic cell lines to HUVEC. Taken together, these results reveal a novel mechanism for adhesion of platelets and megakaryocytic cells to vascular endothelial cells.

An established cell line originating from a Sarcophaga peregrina (fleshfly) embryo, NIH-Sape-4, was found to synthesize mRNAs for Sarcophaga lectin and sarcotoxin IA, but not those for storage protein or 25 kDa protein. These four proteins are known to be synthesized in the fat-body of third-instar larvae, and the two former in particular are known to participate in the defence mechanism of this insect and to be induced in response to injury of the body wall. Thus the embryonic cell line NIH-Sape-4 synthesizes certain defence proteins constitutively. This cell line will be useful for large-scale purification of Sarcophaga lectin, since 50 micrograms of purified Sarcophaga lectin could be obtained from about 400 ml of culture medium.

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<b><i>Introduction:</i></b> Pneumonia is one of the most frequently occurring complications after esophagectomy and is associated with increased operative mortality. Chronic obstructive pulmonary disease (COPD) is known to be a risk factor for pulmonary complications and operative mortality. However, in COPD patients preparing for esophagectomy, preventive measures against postoperative pneumonia have not yet been discovered. In this study, we evaluate the effect of perioperative inhaled tiotropium, a long-acting, antimuscarinic bronchodilator used in the management of COPD, on patients with COPD who undergo esophageal cancer surgery. <b><i>Methods/Design:</i></b> This study investigates the effect of perioperative inhaled tiotropium on patients with COPD who undergo esophagectomy. It is an open-label, randomized controlled trial conducted in a single center (EPITOPE study). A total of 32 enrolled patients are randomly assigned in a 1:1 ratio to either conventional management or inhalation of tiotropium in addition to the conventional management. Patients included in the intervention group receive tiotropium Respimat 5 μg (two inhalations of 2.5 μg) for at least 2 weeks before the esophagectomy. Following the esophagectomy, tiotropium is re-delivered, starting as early as possible and continuing until the postoperative evaluation (between 30 and 44 days after the operation). The primary outcome is the incidence of pneumonia within 30 days after esophagectomy. Secondary outcomes are the incidence of cardiovascular complications within 30 days after esophagectomy, the incidence of any postoperative complications within 30 days after esophagectomy, pulmonary function (preintervention, preoperative, and postoperative), walking distance in the incremental shuttle walking test (preintervention, preoperative, and postoperative), the incidence of adverse events, and mortality within 30 days after esophagectomy. <b><i>Discussion:</i></b> The EPITOPE study is the first pilot study on the effects of perioperative inhaled tiotropium on patients with COPD undergoing esophagectomy. After completing this study, we will plan a multicenter RCT with the appropriate outcomes in the future.

Traumatic events frequently produce false fear memories. We investigated the effect of hypothalamic corticotropin-releasing factor (CRF) knockdown (Hy-Crf-KD) or overexpression (Hy-CRF-OE) on contextual fear memory, as fear stress-released CRF and hypothalamic–pituitary–adrenal axis activation affects the memory system. Mice were placed in a chamber with an electric footshock as a conditioning stimulus (CS) in Context A, then exposed to a novel chamber without CS, as Context B, at 3 h (B-3h) or 24 h (B-24h). The freezing response in B-3h was intensified in the experimental mice, compared to control mice not exposed to CS, indicating that a false fear memory was formed at 3 h. The within-group freezing level at B-24h was higher than that at B-3h, indicating that false context fear memory was enhanced at B-24h. The difference in freezing levels between B-3h and B-24h in Hy-Crf-KD mice was larger than that of controls. In Hy-CRF-OE mice, the freezing level at B-3h was higher than that of control and Hy-Crf-KD mice, while the freezing level in B-24h was similar to that in B-3h. Locomotor activity before CS and freezing level during CS were similar among the groups. Therefore, we hypothesized that Hy-Crf-KD potentiates the induction of false context fear memory, while Hy-CRF-OE enhances the onset of false fear memory formation.

Previously we reported the discovery of the ASE/AIE-type age related regulatory mechanism of gene expression, the first molecular mechanism for age dimension homeostasis (Kurachi et al, Science 1999; Zhang et al, J Biol Chem 2002; Kurachi & Kurachi J Thromb Haemost 2005). Here we report a comprehensive analysis of the age associated changes of mouse liver nuclear proteins, an experimental model for studying the dynamic regulatory mechanisms of the age-dimension homeostasis. Liver nuclei obtained from mice (C57BL/6xSJL, male) at 1, 3, 6, 12, 18 and 21 months of age (n=10–20/age) were used for preparing protein extracts. The protein extracts were then subjected to 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE) (pH range of 4–11) and protein identification by MALDI-TOF/MS. As analyzed by the PDQuest program combined with Mascot Protein Identifier, out of over 5000 protein spots identified by 2D-PAGE, approximately 2800 protein spots were found unique including isomers. Intensity changes of these protein spots along the age axis were determined and analyzed in details with the PDQuest software. Twelve proteins were found to show no significant changes in their intensity throughout the life-span (smaller than 20% fluctuation). Besides some complex patterns, at least 6 unique age-related patterns were found, including those of puberty-onset increase, puberty-onset decrease, puberty-onset increase up to 6 months of age followed by decrease, unique increase only at around 12 months of age, ageing associated increase or decrease. Age related pattern changes were observed for some isomers among many, which were likely generated by differential posttranslational modifications or conformational changes. These findings support that in addition to the ASE/AIE type mechanism, there exist multiple novel age-dimension regulatory mechanisms, and their various combinations are responsible for producing complex and subtle age related regulatory patterns for a large number of genes. This is the first comprehensive analysis of the liver nuclear proteins in relation to age, and would provide a valuable basis for studying many age related phenomena, diseases as well as epigenetic effects.