In vitro culture development and polyphenolics production of Artemisia alba Turra

Open Access Journal Second National Youth Conference “Biological sciences for a better future”, Plovdiv, October 30-31, 2015.Artemisia alba Turra is an aromatic plant, characterized by a high variability of the terpenoid profile of its essential oil. In previous research, in vitro shoots of the plant were developed, aiming at elucidation of the effects of plant growth regulators on essential oil production. Though less information is available in literature regarding the non-volatile components of the plant, a number of works report on the presence of compounds with coumarin, flavonoid and sesquiterpene structure which might attribute to the pharmacological activity of the plant. In the present work, different lines of differentiated and non-differentiated in vitro cultures of the plant have been developed in solid and liquid media. The potential of these lines to produce compounds with phenolic and flavonoid structure has been studied. In differentiated shoot cultures, low benzyl adenine (BA) concentration alone or in combination with different indole-3-butyric acid (IBA) concentrations increased the polyphenolic levels as compared with plant growth regulators free control, as well with media with high BA alone or combined with IBA. The content of these compounds was also low when IBA was applied alone. In non-differentiated cell aggregate cultures, 1-naphthaleneacetic acid (NAA) in combination with BA significantly increased polyphenolics as compared with IBA. Observations on the morphology of the aggregates formed in the two media suggested that the more compact structure and larger size of aggregates as a result of NAA supplementation might be decisive for the higher polyphenolics productivity, as compared with IBA

Calorimetric Study of Helix aspersa Maxima Hemocyanin Isoforms

The thermal unfolding of hemocyanin isoforms, β-HaH and αD+N-HaH, isolated from the hemolymph of garden snails Helix aspersa maxima, was studied by means of differential scanning calorimetry (DSC). One transition, with an apparent transition temperature (Tm) at 79.88°C, was detected in the thermogram of β-HaH in 20 mM HEPES buffer, containing 0.1 M NaCl, 5 mM CaCl2, and 5 mM MgCl2, pH 7.0, at scan rate of 1.0°C min−1. By means of successive annealing procedure, two individual transitions were identified in the thermogram of αD+N-HaH. Denaturation of both hemocyanins was found to be an irreversible process. The scan-rate dependence of the calorimetric profiles indicated that the thermal unfolding of investigated hemocyanins was kinetically controlled. The thermal denaturation of the isoforms β-HaH and αD+N-HaH was described by the two-state irreversible model, and parameters of the Arrhenius equation were calculated

Antitumor Properties of Epitope-Specific Engineered Vaccine in Murine Model of Melanoma

Finding new effective compounds of natural origin for composing anti-tumor vaccines is one of the main goals of antitumor research. Promising anti-cancer agents are the gastropodan hemocyanins–multimeric copper-containing glycoproteins used so far for therapy of different tumors. The properties of hemocyanins isolated from the marine snail Rapana thomasiana (RtH) and the terrestrial snail Helix aspersa (HaH) upon their use as carrier-proteins in conjugated vaccines, containing ganglioside mimotope GD3P4 peptide, were studied in the developed murine melanoma model. Murine melanoma cell line B16F10 was used for solid tumor establishment in C57BL/6 mice using various schemes of therapy. Protein engineering, flow cytometry, and cytotoxicity assays were also performed. The administration of the protein-engineered vaccines RtH-GD3P4 or HaH-GD3P4 under the three different regimens of therapy in the B16F10 murine melanoma model suppressed tumor growth, decreased tumor incidence, and prolonged the survival of treated animals. The immunization of experimental mice induced an infiltration of immunocompetent cells into the tumors and generated cytotoxic tumor-specific T cells in the spleen. The treatment also generates significantly higher levels of tumor-infiltrated M1 macrophages, compared to untreated tumor-bearing control mice. This study demonstrated a promising approach for cancer therapy having potential applications for cancer vaccine research