The Challenges of Vaccine Development against Betacoronaviruses: Antibody Dependent Enhancement and Sendai Virus as a Possible Vaccine Vector

Molecular Biology - To design an effective and safe vaccine against betacoronaviruses, it is necessary to use their evolutionarily conservative antigenic determinants that will elicit the...     

In vitro and in vivo study of pluripotency in intraspecific hybrid cells obtained by fusion of murine embryonic stem cells with splenocytes

Hypoxanthine phosphoribosyltransferase–deficient (HPRT^-) mouse embryonic stem (ES) cells, HM-1 cells (genotype XY), were fused with adult female DD/c mouse spleen cells. As a result, a set of HAT-resistant clones was isolated. Four hybrid clones most similar in morphology and growth characteristics to the HM-1 cells were studied in detail with respect to their pluripotency. Of these, three clones contained 41–43 chromosomes, and one clone was nearly tetraploid. All the clones had the XXY set of sex chromosomes and expressed the HPRT of the somatic partner only. The hybrid clones shared features with the HM-1 cells, indicating that they retained their pluripotent properties: (1) embryonic ECMA-7 antigen, not TROMA-1 antigen, was present in most cells; (2) the hybrid cells showed high activity of endogenous alkaline phosphatase (AP); (3) all the hybrid clones were able to form complex embryoid bodies containing derivatives of all the embryonic germinal layers; (4) the hybrid cells contained synchronously replicating X chromosomes, indicating that they were in an active state; and (5) a set of chimeric animals was generated by injecting hybrid cells into BALB/c and C57BL/6J mouse blastocysts. Evidence for chimerism was provided by the spotted coat derived from 129/Ola mice and identification of 129/Ola glucose phosphate isomerase (GPI) in many organs. Thus the results obtained demonstrated that the hybrid cells retain their high pluripotency level despite the close contact of the “pluripotent” HM-1 genome with the “somatic” spleen cell genome during hybrid cell formation and the presence of the “somatic” X chromosome during many cell generations. The presence of HPRT of the somatic partner in many organs and tissues, including the testes in chimeric animals, shows that the “somatic” X chromosome segregates weakly, if at all, during development of the chimeras. There were no individuals with the 129/Ola genotype among the more than 50 offspring from chimeric mice. The lack of the 129/Ola genotype is explained by the imbalance of the sex chromosomes in the hybrid cells rendering the passage of hybrid cell descendants through meiosis in chimeras impossible. As a result, chimeras become unable to produce gametes of the hybrid cell genotype. Mol. Reprod. Dev. 50:128–138, 1998. © 1998 Wiley-Liss, Inc.     

Evaluation of the hypomagnetic environment effects on capillary blood circulation,blood pressure,and heart rate

The impact of attenuated magnetic field (МF) on human health is a burning issue of present-day cosmonautics. A series of experiments with animals exposed to attenuated MF revealed violent disorders in the development of the cardiovascular system. The purpose of this study was to estimate the effects of hypomagnetic environment (HME) on capillary blood circulation, blood pressure (BP), and heart rate (HR) in healthy humans. Participants (n = 34) were 24 men and 10 women without cardiovascular symptoms. The mean age was 43.3 ± 15.4 years. Thirteen participants, eight men and five women, were randomly selected for a repeated investigation under natural conditions (sham exposure). The mean age in this group was 47.9 ± 18 years. Cardiac rhythm and heart rate were recorded using an Astrocard cardiac monitor (Russia). BP was measured by means of a Tonocard automatic blood pressure monitor (Russia). Capillary circulation was determined using a digital capillaroscope (Russia) with a high-speed CMOS camera (100 frames/s). The duration of HME exposure was 60 min. It has been demonstrated that HME increases capillary circulation rate by 22.4% in healthy humans without cardiovascular symptoms as compared to the records made under natural conditions. There was a significant HR reduction by the end of HME exposure as compared to the measurements taken at the beginning. At the end of the exposure, diastolic BP dropped considerably relative to mid-exposure values; on the contrary, systolic BP significantly increased by the end.     

45-KDa GTP-binding Protein from Rat Olfactory Epithelium: Purification, Characterization and Localization

The rat olfactory epithelium contains a specific water-soluble45-kDa protein. This protein is recognized by anti-peptide antibodieswhich react with -subunits of the known G-proteins. The 45-kDaprotein has been isolated using DEAE-chromatography and gel-exclusionchromatography. The content of 45-kDa protein is about 2% ofthe total soluble proteins of the olfactory mucosa and it islocated at the mucociliary surface. According to photo-affinitylabeling, the 45-kDa protein possesses a high affinity to GTPand exhibits low GTP hydrolytic activity. The functions of the45-kDa protein are discussed. Chem, Senses 21: 181–188,1996.     

Lucigenin-enhanced chemiluminescence of the animal tissues

Lucigenin-enhanced chemiluminescence (LcCL) allows one to investigate the reactions of superoxide anion radical (*O2-) generated by mitochondria and is applied to study the superoxide production in enzymatic and membrane systems by isolated mitochondria and cells, and in whole organs. The application of lucigenin-enhanced chemiluminescence to estimate the respiration of human tissues involves the use of small tissue pieces, which can be obtained, for instance, by biopsia; however, no systematic investigations have been performed on these objects. In the present paper, a comparative study of lucigenin-enhanced chemiluminescence of tissues isolated from different organs of the rat was carried out to elucidate its dependence on the extent of tissue defragmentation, storage time, and access for oxygen. It was shown that the addition of lucigenin to a piece of tissue, a suspension of fine tissue fragments, and homogenates greatly enhanced chemiluminescence, and a whole piece of tissue possessed a much lesser (by 1-1.5 order of magnitude) intensity of chemiluminescence than homogenate or gruel. In the absence of stirring of the surrounding solution, the lucigenin-enhanced chemiluminescence of tissue quickly decreased, apparently due to a decrease in the level of oxygen in the tissue, as the result of its consumption. The chemiluminescence consisted of two components: a lucigenin-dependent and lucigenin-independent one (intrinsic chemiluminescence). Thus, the tissue was a source of lucigenin-enhanced chemiluminescence, and this luminescence was observed only at a sufficient access for oxygen. The lucigenin-independent component did not practically depend on oxygen and was determined by the components coming out of the tissue into the surrounding solution. Nitric oxide (NO) inhibited chemiluminescence as its concentration increased and did not affect considerably the rate of oxygen consumption by the tissue. The results obtained allow one to conclude that lucigenin can be used as a rather effective chemiluminescent probe for the production of superoxide radicals by tissue pieces.     

Widespread occurrence of natural genetic transformation of plants by Agrobacterium

Plant Molecular Biology - Naturally transgenic plant species occur on an unexpectedly large scale. Agrobacterium-mediated gene transfer leads to the formation of crown galls or hairy roots, due to...     

Effect of space flight factors simulated in ground-based experiments on the behavior,discriminant learning,and exchange of monoamines in different brain structures of rats

Experimental treatment (long-term fractionated γ-irradiation, antiorthostatic hypodynamia, and the combination of these factors) simulating the effect of space flight in ground-based experiments rapidly restored the motor and orienting-investigative activity of animals (rats) in “open-field” tests. The study of the dynamics of discriminant learning of rats of experimental groups did not show significant differences from the control animals. It was found that the minor effect of these factors on the cognitive performance of animals correlated with slight changes in the concentration of monoamines in the brain structures responsible for the cognitive, emotional, and motivational functions.     

Degradation of proteinaceous substrates by xylotrophic basidiomycetes

The ability of various xylotrophs to produce extracellular proteolytic enzymes has been studied, with emphasis on medium-related factors regulating their secretion. Direct measurement of proteolytic activity in the culture liquid and postelectrophoresis determination of protease activity in polyacrylamide gel copolymerized with gelatin demonstrated that the secreted enzymes are quantitatively and qualitatively diverse. Activity levels of extracellular proteolytic enzymes strongly depend on pH and contents of protein and carbohydrate in the medium. All secreted proteases notably differed in molecular weight (of 51 kDa or higher and in excess of 95 kDa) and belonged mostly to two classes of proteolytic enzymes (serine proteases and metalloproteinases).     

Changes of trace element content in MARC-145 cells before and after infection with virulent and attenuated strains of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV)

The influence of virulent and attenuated strains of Porcine Reproductive and Respiratory Syndrome Virus with defined genomic differences on trace element profile of MARC-145 cells was investigated to elucidate the process of infection in vitro. The concentrations of seven trace elements (Al, Cr, Mn, Ni, Fe, Cu, Zn) in infected and non-infected cells were determined. Although viral replication was similar in all cases, there were distinct features of difference in the trace element profiles (changes of concentrations of Ni, Mn, Fe) in infected cells. In particular, cells infected with the attenuated strain of PRRSV (NADC8-251) contained very low level of Ni, compared with its pathogenic counterpart NADC8-252K. The infection with the Lelystad strain also led to a moderately high level of this trace element.     

Oncolytic Paramyxoviruses: Mechanism of Action,Preclinical and Clinical Studies

Preclinical studies demonstrate that a broad spectrum of human and animal malignant cells can be killed by oncolytic paramyxoviruses, which includes cells of ecto-, endo- and mesodermal origin. In clinical trials, significant reduction or even complete elimination of primary tumors and established metastases has been reported. Different routes of virus administration (intratumoral, intravenous, intradermal, intraperitoneal, or intrapleural) and single- vs. multiple-dose administration schemes have been explored. The reported side effects were grades 1 and 2, with the most common among them being mild fever. There are certain advantages in using paramyxoviruses as oncolytic agents compared to members of other virus families exist. Thanks to cytoplasmic replication, paramyxoviruses do not integrate the host genome or engage in recombination, which makes them safer and more attractive candidates for widely used therapeutic oncolysis than retroviruses or some DNA viruses. The list of oncolytic Paramyxoviridae members includes the attenuated measles virus, mumps virus, low pathogenic Newcastle disease, and Sendai viruses. Metastatic cancer cells frequently overexpress certain surface molecules that can serve as receptors for oncolytic paramyxoviruses. This promotes specific viral attachment to these malignant cells. Paramyxoviruses are capable of inducing efficient syncytium-mediated lysis of cancer cells and elicit strong immune stimulation, which dramatically enforces anticancer immune surveillance. In general, preclinical studies and phases I–III of clinical trials yield very encouraging results and warrant continued research of oncolytic paramyxoviruses as a particularly valuable addition to the existing panel of cancer-fighting approaches.