Major linear antibody epitopes and capsid proteins differentially induce protective immunity against Theiler's virus-induced demyelinating disease.

Theiler's murine encephalomyelitis virus-induced immunologically mediated demyelinating disease (TMEV-IDD) in susceptible mice provides a relevant infectious model for multiple sclerosis. Previously, we have identified six major linear antibody epitopes on the viral capsid proteins. In this study, we utilized fusion proteins containing individual capsid proteins and synthetic peptides containing the linear antibody epitopes to determine the potential role of antibody response in the course of virus-induced demyelination. Preimmunization of susceptible mice with VPI and VP2 fusion proteins, but not VP3, resulted in the protection from subsequent development of TMEV-IDD. Mice free of clinical symptoms following preimmunizations with fusion proteins displayed high levels of antibodies to the capsid proteins corresponding to the immunogens. In contrast, the level of antibodies to a particular linear epitope, A1C (VP1(262-276)), capable of efficiently neutralizing virus in vitro increased with the progression of disease. Further immunization with synthetic peptides containing individual antibody epitopes indicated that antibodies to the epitopes are differentially effective in protecting from virus-induced demyelination. Taken together, these results suggest that antibodies to only certain linear epitopes are protective and such protection may be restricted during the early stages of viral infection.     

Nuclear magnetic resonance and molecular modeling studies on O-beta-D-galactopyranosyl-(1----4)-O-beta-D-xylopyranosyl-(1----0)-L-se rine, a carbohydrate-protein linkage region fragment from connective tissue proteoglycans

The solution conformation of O-beta-D-galactopyranosyl-(1----4)-O-beta-D-xylopyranosyl-(1----0)-L-ser ine (GXS), a carbohydrate-protein linkage region fragment from connective tissue proteoglycans, was investigated by two-dimensional NMR spectroscopy and molecular modeling calculations. Specifically, the 1H and 13C resonances were assigned by 2D-COSY and by 1H-13C heteronuclear correlation spectroscopy methods. 2D-NOESY was used to generate distance constraints between the galactose and xylose and between the xylose and serine residues. The 1H vicinal coupling constants for the sugars and the serine were also determined. A general molecular modeling methodology suitable for complex carbohydrates was developed. This methodology employed molecular dynamics and energy minimization procedures together with the application of inter-residue spatial constraints across the linkages derived from 2D-NOESY. The first step in this methodology is the generation of a wide variety of starting conformations that span the (phi, psi) space for each linkage. In the present study, nine such conformations were constructed for each linkage using the torsion angles phi and psi corresponding to the gauche+, gauche-, and trans configurations across each of the two bonds constituting the linkage. These conformations were subjected to a combined molecular dynamics/energy minimization refinement using the NOESY derived constraints as pseudoenergy functions. Families of conformations for the whole molecule were then constructed from the structures derived for each linkage. Characterization of GXS using this methodology identified a single family of conformations that are consistent with the solution phase NMR data on this molecule.     

MA 160 and EB 33 cell lines: HeLa cell contaminants,hybrids or prostatic epithelial cells?

Summary Studies of acid phosphates produced by cell lines MA 160 and EB 33 demonstrated immunochemically their prostatic origin. MA 160 and EB 33, rather than being HeLa contaminants, may be hybrids of prostatic epithelial and HeLa cells or true prostatic cell lines with chromosomal changes common to all long-term cultivated cell lines. This research was supported by NIH (Cancer) Research Grants Nos. 18748 and 16426; and Detroit General Hospital Research Corporation.     

The Metabolism of Oat Leaves during Senescence: III. The Senescence of Isolated Chloroplasts

The changes in chlorophyll and protein in senescing chloroplasts isolated from the first leaves of 7-day-old oat (Avena sativa) seedlings have been investigated. In darkness the chlorophyll in these plastids is highly stable, losing only 5 to 10% of its content after 7 days at 26 C. This result contrasts with the behavior of chlorophyll in intact leaves, in which about 80% of the pigment would have disappeared in that time. The protein is less stable than the chlorophyll, though more stable than in the leaf; probably a small amount of protease is present in the plastids. Some protein is also being synthesized in the chloroplasts along with its breakdown; gains of up to 38% in protein and 13% in chlorophyll were observed under different conditions. l-Serine, which actively promotes senescence in the leaf, has only a very slight effect on the chloroplasts, and kinetin antagonizes it. Kinetin also has a small but significant effect in preserving the protein from breakdown. Acid pH somewhat promotes the breakdown, both of chlorophyll and protein. A loss of chlorophyll and protein comparable to that occurring in the senescence of the leaf could not be induced in the chloroplasts by suspending them in malate, in cytoplasmic extract, or in any of a number of enzymes tested alone. Incubation with a mixture of four enzymes was the only treatment which approximated the senescent process in the leaf, causing 34% loss of chlorophyll at pH 5 and 40% loss of protein at pH 7.4, both in 72 hours.In white light, the chlorophyll and the carotenoids, but not the protein, disappear rapidly. This disappearance was shown to be prevented in an atmosphere of nitrogen or in air by a number of reducing agents, of which ascorbic acid was the most effective. It is, therefore, ascribed to photooxidation rather than to normal senescence.     

Fgfbp1 is essential for the cellular survival during zebrafish embryogenesis

Fibroblast growth factor binding protein 1 (FGFBP1) is expressed in various tumors and may serve as a diagnostic marker and/or a therapeutic target. Previous studies suggested FGFBP1 functions as an angiogenic switch molecule by regulating the activity of FGF2, and it was later found to associate with a broad spectrum of FGFs. To study FGFBP1, we used zebrafish, in which the function of extracellular matrix protein can be easily studied in intact tissues or organisms. When Fgfbp1 expression was knocked down, morphants manifested massive cell death and structural abnormalities. Cell death was most prominent in the brain and the neural tube, but not limited to those regions. These findings suggest that the primary function of Fgfbp1 may be to sustain cellular survival throughout embryogenesis. For comparison, the expression of fgf2 was limited to the early stage of embryogenesis and fgf2 morphants showed more severe phenotype, with high morbidity before reaching 14-somites. Taken together, our work reveals the physiologic function of Fgfbp1, and that its function could be exerted in a Fgf2-independent manner.     

Association of myostatin deficiency with collagen related disease-umbilical hernia and tippy toe standing in pigs

Transgenic Research - Herein, we investigate the high incidence of umbilical hernia and tippy-toe standing and their underlying changes in gene expression and proliferation in myostatin knockout...     

Hydroxychloroquine-mediated inhibition of SARS-CoV-2 entry is attenuated by TMPRSS2

Hydroxychloroquine, used to treat malaria and some autoimmune disorders, potently inhibits viral infection of SARS coronavirus (SARS-CoV-1) and SARS-CoV-2 in cell-culture studies. However, human clinical trials of hydroxychloroquine failed to establish its usefulness as treatment for COVID-19. This compound is known to interfere with endosomal acidification necessary to the proteolytic activity of cathepsins. Following receptor binding and endocytosis, cathepsin L can cleave the SARS-CoV-1 and SARS-CoV-2 spike (S) proteins, thereby activating membrane fusion for cell entry. The plasma membrane-associated protease TMPRSS2 can similarly cleave these S proteins and activate viral entry at the cell surface. Here we show that the SARS-CoV-2 entry process is more dependent than that of SARS-CoV-1 on TMPRSS2 expression. This difference can be reversed when the furin-cleavage site of the SARS-CoV-2 S protein is ablated or when it is introduced into the SARS-CoV-1 S protein. We also show that hydroxychloroquine efficiently blocks viral entry mediated by cathepsin L, but not by TMPRSS2, and that a combination of hydroxychloroquine and a clinically-tested TMPRSS2 inhibitor prevents SARS-CoV-2 infection more potently than either drug alone. These studies identify functional differences between SARS-CoV-1 and -2 entry processes, and provide a mechanistic explanation for the limited in vivo utility of hydroxychloroquine as a treatment for COVID-19.     

Pairwise and Bipartite Entanglements of Three Non-Equally Separated Quantum Dots in Plasmonic Nanowaveguide System

Plasmonics - We theoretically investigate properties of the pairwise and bipartite entanglements of three non-equally separated quantum dots (QDs) coupled to one-dimensional plasmonic nanowaveguide...     

Regulation of ICAM‐1 expression in gingival fibroblasts infected with high‐glucose‐treated P. gingivalis

Porphyromonas gingivalis is a major pathogen in the initiation and progression of periodontal disease, which is recognized as a common complication of diabetes. ICAM‐1 expression by human gingival fibroblasts (HGFs) is crucial for regulating local inflammatory responses in inflamed periodontal tissues. However, the effect of P. gingivalis in a high‐glucose situation in regulating HGF function is not understood. The P. gingivalis strain CCUG25226 was used to study the mechanisms underlying the modulation of HGF ICAM‐1 expression by invasion of high‐glucose‐treated P. gingivalis (HGPg). A high‐glucose condition upregulated fimA mRNA expression in P. gingivalis and increased its invasion ability in HGFs. HGF invasion with HGPg induced increases in the expression of ICAM‐1. By using specific inhibitors and short hairpin RNA (shRNA), we have demonstrated that the activation of p38 MAPK and Akt pathways is critical for HGPg‐induced ICAM‐1 expression. Luciferase reporters and chromatin immunoprecipitation assays suggest that HGPg invasion increases NF‐κB‐ and Sp1‐DNA‐binding activities in HGFs. Inhibition of NF‐κB and Sp1 activations blocked the HGPg‐induced ICAM‐1 promoter activity and expression. The effect of HGPg on HGF signalling and ICAM‐1 expression is mediated by CXC chemokine receptor 4 (CXCR4). Our findings identify the molecular pathways underlying HGPg‐dependent ICAM‐1 expression in HGFs, providing insight into the effect of P. gingivalis invasion in HGFs.     

Impacts of Intergroup Interactions on Intragroup Behavioral Changes in Javan Gibbons (Hylobates moloch)

International Journal of Primatology - Agonistic intergroup interactions can cause individual costs such as physical injuries, increased physiological stress, and disrupted intragroup social...