Determination of the expression of fish antifreeze protein (AFP) in 7th generation transgenic mice tissues and serum

In this study, the presence of antifreeze protein (AFP) gene expression through successive generations in transgenic mice carrying the chimeric gene construct of the coding sequence for the AFP protein from ocean pout was investigated. AFP transgenic hemizygote mice were used for AFP gene expression. AFP genome expressions in transgenic mice were analyzed by Western blotting, and tissue location of AFP protein was shown by immunohistochemical and immunofluorescence techniques. Seventh transgenic mice from the established founders demonstrated the expression of AFP in organs such as the skin, oviduct, lung, kidney and liver tissues and serum except for the heart. Our results demonstrate successful expression of AFP gene products in several tissues and serum of transgenic mice, the association of in vivo expressed AFP protein, for the first time. These results indicate that the coding sequence for the AFP protein gene (ocean pout type III AFP gene) could be integrated and stably transcribed and expressed in the 7th generation of transgenic mice. In conclusion transgenic mouse lines would be a good model for the cryostudy of AFP and for the determination of AFP roles in several organs and tissues.     

Elevation of body temperature is an essential factor for exercise-increased extracellular heat shock protein 72 level in rat plasma

This study examined whether the exercise-increased extracellular heat shock protein 72 (eHsp72) levels in rats was associated with body temperature elevation during exercise. In all, 26 female Sprague-Dawley rats (3 mo old) were assigned randomly to control (CON; n = 8), exercise under warm temperature (WEx; n = 9), or exercise under cold temperature (CEx; n = 9). The WEx and CEx were trained at 25 degrees C or 4 degrees C, respectively, for nine days using a treadmill. Before and immediately after the final exercise bout, the colonic temperatures were measured as an index of body temperature. The animals were subsequently anesthetized, and blood samples were collected and centrifuged. Plasma samples were obtained to assess their eHsp72 levels. Only the colonic temperature in WEx was increased significantly (P < 0.05) by exercise. The eHsp72 level in WEx was significantly higher (P < 0.05) than that of either the CON or CEx. However, no significant difference was found between CON and CEx. Regression analyses revealed that the eHsp72 level increased as a function of the body temperature. In another experiment, the eHsp72 level of animals with body temperature that was passively elevated through similar kinetics to those of the exercise was studied. Results of this experiment showed that mere body temperature elevation was insufficient to induce eHsp72 responses. Collectively, our results suggest that body temperature elevation during exercise is important for induction of exercise-increased eHsp72. In addition, the possible role of body temperature elevation is displayed when the exercise stressor is combined with it.     

The Fibrin Matrix Regulates Angiogenic Responses within the Hemostatic Microenvironment through Biochemical Control

Conceptually, premature initiation of post-wound angiogenesis could interfere with hemostasis, as it relies on fibrinolysis. The mechanisms facilitating orchestration of these events remain poorly understood, however, likely due to limitations in discerning the individual contribution of cells and extracellular matrix. Here, we designed an in vitro Hemostatic-Components-Model (HCM) to investigate the role of the fibrin matrix as protein factor-carrier, independent of its cell-scaffold function. After characterizing the proteomic profile of HCM-harvested matrix releasates, we demonstrate that the key pro-/anti-angiogenic factors, VEGF and PF4, are differentially bound by the matrix. Changing matrix fibrin mass consequently alters the balance of releasate factor concentrations, with differential effects on basic endothelial cell (EC) behaviors. While increasing mass, and releasate VEGF levels, promoted EC chemotactic migration, it progressively inhibited tube formation, a response that was dependent on PF4. These results indicate that the clot’s matrix component initially serves as biochemical anti-angiogenic barrier, suggesting that post-hemostatic angiogenesis follows fibrinolysis-mediated angiogenic disinhibition. Beyond their significance towards understanding the spatiotemporal regulation of wound healing, our findings could inform the study of other pathophysiological processes in which coagulation and angiogenesis are prominent features, such as cardiovascular and malignant disease.     

Different Patterns of White Matter Degeneration Using Multiple Diffusion Indices and Volumetric Data in Mild Cognitive Impairment and Alzheimer Patients

Alzheimeŕs disease (AD) represents the most prevalent neurodegenerative disorder that causes cognitive decline in old age. In its early stages, AD is associated with microstructural abnormalities in white matter (WM). In the current study, multiple indices of diffusion tensor imaging (DTI) and brain volumetric measurements were employed to comprehensively investigate the landscape of AD pathology. The sample comprised 58 individuals including cognitively normal subjects (controls), amnestic mild cognitive impairment (MCI) and AD patients. Relative to controls, both MCI and AD subjects showed widespread changes of anisotropic fraction (FA) in the corpus callosum, cingulate and uncinate fasciculus. Mean diffusivity and radial changes were also observed in AD patients in comparison with controls. After controlling for the gray matter atrophy the number of regions of significantly lower FA in AD patients relative to controls was decreased; nonetheless, unique areas of microstructural damage remained, e.g., the corpus callosum and uncinate fasciculus. Despite sample size limitations, the current results suggest that a combination of secondary and primary degeneration occurrs in MCI and AD, although the secondary degeneration appears to have a more critical role during the stages of disease involving dementia.     

The micronucleus assay in exfoliated buccal cells: application to occupational exposure to polycyclic aromatic hydrocarbons.

Many polycyclic aromatic hydrocarbons (PAHs) have been identified as cancer-inducing chemicals for animals and/or humans. Also, there is sufficient evidence that exposures in the occupational settings are carcinogenic or probably carcinogenic to human. Engine exhaust and used engine oils are major PAH sources in engine repair workshops and traffic. Analysis of micronucleus (MN) in exfoliated buccal cells is a sensitive method for monitoring genetic damage in human populations. In our study, we used three different occupational groups (Group 1; engine repair workers, Group 2; taxi drivers, Group 3; traffic police) and two controls (Control I for Group 1 and Control II for Group 2 and Group 3) for the exposed groups. We analysed MN frequencies in exfoliated buccal cells and compared the exposed groups (Group 1; n=34, Group 2; n=17, Group 3; n=15) and subjects not occupationally exposed to PAH (Control I; n=28, Control II; n=20). The mean (+/-S.D.) MN (%) frequencies in exfoliated buccal cells from Group 1 and Control I were 0.07+/-0.05 and 0. 05+/-0.04, respectively (p>0.05; Table 2). The mean (+/-S.D.) MN (%) frequencies in exfoliated buccal cells from Group 2, 3 and Control II were 0.12+/-0.05, 0.10+/-0.05 and 0.03+/-0.03, respectively (p<0. 0001, p<0.05; Table 2) Smokers and nonsmokers do not differ with respect to the incidence of MN in all groups.     

Seroprevalence of Borrelia burgdorferi infection among forestry workers and farmers in Duzce, north-western Turkey

Borrelia burgdorferi infection is the most frequent tick-transmitted disease worldwide. Our aim was to assess the seroprevalence of B. burgdorferi infection among forestry workers and farmers in Duzce, in the north-west region of Turkey. Blood samples from 349 forestry workers and farmers and 193 healthy blood donors were obtained to determine the presence of antibodies to B. burgdorferi. A two-step testing strategy was used; the sera were initially tested by ELISA and then by Western blot (WB) IgG. Demographic data regarding residence, age, gender, profession, tick bite history, contact with animals, and symptoms involving the skin, nervous system, and osteoarticular system were collected by questionnaire. All results were evaluated statistically using the chi2 test. The seroprevalence of B. burgdorferi was 10.9% (n=38) in forestry workers and farmers and 2.6% (n=5) in blood donors by ELISA, with a statistically significant difference (p<0.001). Seropositivity rates were related to age, gender, and common risk factors for the disease. IgG seropositivity was confirmed in four (1.1%) sera by WB. In this first seroepidemiological report from the northwest region of Turkey, tick bite exposure was found to be high, whereas B. burgdorferi infection was not common. Preventive measures against tick exposure and further studies to determine the distribution of Lyme disease in Turkey are proposed.     

SAR and biological evaluation of SEN12333/WAY-317538: Novel alpha 7 nicotinic acetylcholine receptor agonist

Alpha 7 nicotinic acetylcholine receptor (α₇ nAChR) agonists are promising therapeutic candidates for the treatment of cognitive impairment associated with a variety of disorders including Alzheimer’s disease and schizophrenia. Alpha 7 nAChRs are expressed in brain regions associated with cognitive function, regulate cholinergic neurotransmission and have been shown to be down regulated in both schizophrenia and Alzheimer’s disease. Herein we report a novel, potent small molecule agonist of the alpha 7 nAChR, SEN12333/WAY-317538. This compound is a selective agonist of the α₇ nAChR with excellent in vitro and in vivo profiles, excellent brain penetration and oral bioavailability, and demonstrates in vivo efficacy in multiple behavioural cognition models. The SAR and biological evaluation of this series of compounds are discussed.     

Role of the clathrin terminal domain in regulating coated pit dynamics revealed by small molecule inhibition

Clathrin-mediated endocytosis (CME) regulates many cell physiological processes such as the internalization of growth factors and receptors, entry of pathogens, and synaptic transmission. Within the endocytic network, clathrin functions as a central organizing platform for coated pit assembly and dissociation via its terminal domain (TD). We report the design and synthesis of two compounds named pitstops that selectively block endocytic ligand association with the clathrin TD as confirmed by X-ray crystallography. Pitstop-induced inhibition of clathrin TD function acutely interferes with receptor-mediated endocytosis, entry of HIV, and synaptic vesicle recycling. Endocytosis inhibition is caused by a dramatic increase in the lifetimes of clathrin coat components, including FCHo, clathrin, and dynamin, suggesting that the clathrin TD regulates coated pit dynamics. Pitstops provide new tools to address clathrin function in cell physiology with potential applications as inhibitors of virus and pathogen entry and as modulators of cell signaling.     

Automated Tracing of Horizontal Neuron Processes During Retinal Development

In the developing mammalian retina, horizontal neurons undergo a dramatic reorganization of their processes shortly after they migrate to their appropriate laminar position. This is an important process because it is now understood that the apical processes are important for establishing the regular mosaic of horizontal cells in the retina and proper reorganization during lamination is required for synaptogenesis with photoreceptors and bipolar neurons. However, this process is difficult to study because the analysis of horizontal neuron anatomy is labor intensive and time-consuming. In this paper, we present a computational method for automatically tracing the three-dimensional (3-D) dendritic structure of horizontal retinal neurons in two-photon laser scanning microscope (TPLSM) imagery. Our method is based on 3-D skeletonization and is thus able to preserve the complex structure of the dendritic arbor of these cells. We demonstrate the effectiveness of our approach by comparing our tracing results against two sets of semi-automated traces over a set of 10 horizontal neurons ranging in age from P1 to P5. We observe an average agreement level of 81% between our automated trace and the manual traces. This automated method will serve as an important starting point for further refinement and optimization.     

Correction to: Imaging features of immune-mediated genitourinary disease

Neurochemical Research - The authors regret that they neglected to cite their conference report on the technical part of a ‘preliminary study’ presented at, and published in, the...