Afroditi Sivropoulou

Antiviral properties of isoborneol, a potent inhibitor of herpes simplex virus type 1

Isoborneol, a monoterpene and a component of several plant essential oils, showed dual viricidal activity against herpes simplex virus 1 (HSV-1). First, it inactivated HSV-1 by almost 4 log10 values within 30 min of exposure, and second, isoborneol at a concentration of 0.06% completely inhibited viral replication, without affecting viral adsorption. Isoborneol did not exhibit significant cytotoxicity at concentrations ranging between 0.016% and 0.08% when tested against human and monkey cell lines. Isoborneol specifically inhibited glycosylation of viral polypeptides based on the following data: (1) the mature fully glycosylated forms of two viral glycoproteins gB and gD were not detected when the virus was replicated in the presence of isoborneol, (2) no major changes were observed in the glycosylation pattern of cellular polypeptides between untreated and isoborneol treated Vero cells, (3) isoborneol did not affect the glycosylation of gB produced from a copy of the gB gene resident in the cellular genome, and (4) other monoterpenes such as 1,8-cineole and borneol, a stereoisomer of isoborneol, did not inhibit HSV-1 glycosylation.

A single amino acid substitution in the cytoplasmic tail of the glycoprotein B of herpes simplex virus 1 affects both syncytium formation and binding to intracellular heparan sulfate

Herpes simplex virus 1 (HSV-1) (S) is a spontaneous syncytial mutant derived from the prototype HSV-1(F) after extensive plaque purification, and produces large syncytial plaques on Vero cells. Marker transfer experiments and DNA sequence analysis mapped the syncytial phenotype to a T-C base substitution at codon 787 of the cytoplasmic domain of mature gB, that results in Leu to Pro substitution and consequently belongs to the syn 3 locus. Both the cytoplasmic and the extracellular domains of gB are active in the fusion event since the addition of anti-gB monoclonal antibodies that recognize the extracellular domain of gB prevent HSV-1(S) induced cell fusion. Similarly, gD also participates in cell fusion since addition of anti-gD monoclonal antibodies also prevent HSV-1(S) induced cell fusion. Furthermore the glycoproteins B and D formed complexes in cells infected with mutant or wild type viruses. The amount of gB bound to total heparan sulfate is lower in the mutant than in the wild type strain. This difference becomes particularly profound when gB is associated with a portion of heparan sulfate intercalated to the membranes. The discrepancy in the binding of the mutant and wild type gB to heparan sulfate may be related to the mechanism of cell fusion induced by HSV-1(S).

Comparison of cell proliferation on modified dental ceramics

The aim of this study was to investigate the influence of substrate characteristics such as chemical composition and surface morphology of dental ceramics to support cell attachment and proliferation. Thus, body (B) and shoulder (S) porcelain differing on their surface morphology and composition were treated with oxides CaO or CaO and P(2)O(5) and four modified ceramics BCa, BCaP, SCa, SCaP were constructed, respectively. The modified ceramics differ from their controls concerning their surface morphology as evaluated by Scanning Electron Microscope (SEM), and their surface chemical composition (Na, KP and Ca) as evaluated by Energy Dispersing Spectroscopy (EDS). All modified ceramics support better than the control ceramics the cell proliferation over 72 h incubation period. Furthermore, higher rates of cell proliferation was detected in shoulder modified ceramics (SCa and SCaP) than in all other cases.

Correlation of the Insecticidal Activity of the Bacillus thuringiensis A4 Strain Against Bactrocera oleae (Diptera) with the 140-kDa Crystal Polypeptide

The crystals of the soil-isolated Bacillus thuringiensis (Bt) strain A4 consist of two polypeptides with molecular mass of 140 kDa and 32 kDa that exhibit insecticidal activity against adult flies of Bactrocera oleae (Diptera). Plasmid curing applied to this strain resulted in the isolation of several subclones exhibiting alterations in their crystal polypeptides as well as two acrystalliferous subclones. The crystals of subclone 1.1 lacked the 32-kDa polypeptide and consisted uniquely of a 140-kDa polypeptide antigenically related to the parental 140-kDa crystal polypeptide. Additionally, the crystals of this subclone exhibited insecticidal activity against B. oleae equivalent to that of the parental strain. Therefore, the 32-kDa crystal polypeptide is dispensable for insecticidal activity, which appears to be dependent on the presence of the 140-kDa crystal polypeptide.

Attachment and Proliferation of Human Periodontal Ligament Fibroblasts on Fibronectin-Coated Bioactive Glass Modified Ceramics

The effect of fibronectin (FN) on human periodontal ligament fibroblasts (PDLF) attachment and proliferation on Bioglass® (PerioGlas® Synthetic Bone Graft Particulate, US Biomaterials) modified dental ceramics, was investigated in vitro. FN introduced limited alterations in cell attachment on Bioglass®-modified dental ceramics in comparison with the corresponding non-FN-coated specimens but had a profound positive effect on Bioglass®-coated specimens that weakly supported both cell attachment and proliferation. The amount of protein adsorbed on the specimens was not proportional to its biological activity, i.e. cell attachment, spread and proliferation, probably due to surface energy variations and FN conformational changes induced by differences in surface composition and morphology of the different dental ceramics modifications.

Transformed cells producing the glycoprotein D of HSV-1 are resistant to infection with clinical strains of HSV

SummaryThe generality of the resistance exhibited by gD producing cells to HSV-1 infection was tested. We tested three different cell lines producing various amounts of gD for resistance against three HSV-1 strains. The strains used were the prototype laboratory F strain and two recently isolated low passage local clinical strains, VG and VD. The results indicate that: (i) the resistance of the cell lines is directly related to the amount of gD they produce, (ii) the cell lines showed greater resistance against the two local clinical HSV-1 strains than against the laboratory strain, and (iii) the resistance is not mediated at the level of virus adsorption to the cell membranes.

In vitro and in vivo study on the effect of antifungal agents on hematopoietic cells in mice

Liposomal amphotericin B, voriconazole, and caspofungin are currently used for systemic and severe fungal infections. Patients with malignant diseases are treated with granulocyte-colony stimulating factor (G-CSF) for the recovery of granulocytes after chemotherapy or hematopoietic cell (HC) transplantation. Since they have a high incidence of fungal infections, they inevitably receive antifungal drugs for treatment and prophylaxis. Despite their proven less toxicity for various cell types comparatively with amphotericin B and the decrease in the number of leukocytes that has been reported as a possible complication in clinical studies, the effect of liposomal amphotericin B, voriconazole, and caspofungin on HCs has not been clarified. The present study aimed to examine the in vitro and in vivo effect of these three modern antifungals on HCs. Colony-forming unit (CFU) assays of murine bone marrow cells were performed in methylcellulose medium with or without cytokines and in the presence or absence of various concentrations of liposomal amphotericin B, voriconazole, and caspofungin. In the in vivo experiments, the absolute number of granulocytes was determined during leukocyte recovery in sublethally irradiated mice receiving each antifungal agent separately, with or without G-CSF. In vitro, all three antifungal drugs were nontoxic and, interestingly, they significantly increased the number of CFU-granulocyte-macrophage colonies in the presence of cytokines, at all concentrations tested. This was contrary to the concentration-dependent toxicity and the significant decrease caused by conventional amphotericin B. In vivo, the number of granulocytes was significantly higher with caspofungin plus G-CSF treatment, higher and to a lesser extent higher, but not statistically significantly, with voriconazole plus G-CSF and liposomal amphotericin B plus G-CSF treatments, respectively, as compared with G-CSF alone. These data indicate a potential synergistic effect of these antifungals with the cytokines, in vitro and in vivo, with subsequent positive effect on hematopoiesis.

Application of a transformed cell line constitutively expressing HSV-1 polypeptides for the detection of HSV antibodies in human sera by an enzyme immunoassay

SummaryWe have previously reported the construction of a cell line BA4, constitutively producing the glycoproteins gD, gG, and α4, the major regulatory protein of HSV-1. These cells have been selected in stepwise increasing concentrations of methotrexate and shown to produce much higher amounts of gD than non-selected cells. Extracts of the selected cells were used in an enzyme linked immunosorbent assay to detect HSV antibodies in human sera obtained from Greek blood donors. We report here that (i) the assay developed is able to distinguish HSV antibody positive from negative human sera and (ii) that its application in an epidemiological survey showed that the incidence of HSV infection in the general population in Greece is 90.4%.