Anti-tumor effect investigation of obtustatin and crude Macrovipera lebetina obtusa venom in S-180 sarcoma bearing mice.

Over the last few decades, research on snake venom toxins has provided not only new tools to decipher molecular details of various physiological processes, but also inspiration to design and develop a number of therapeutic agents. Isolated from the venom of Macrovipera lebetina obutusa (MLO), obtustatin represents the shortest known snake venom monomeric disintegrin specific inhibitor of α1ß1 integrin. This low molecular weight peptide revealed a potent therapeutic effect on melanoma progression. Its oncostatic effect was related to the inhibition of angiogenesis. The aim of the proposed investigation was to study the influence of obtustatin and crude MLO venom on the S-180 sarcoma growth in vitro and in vivo. A S-180 sarcoma bearing mouse model, histological examination, DNA retardation assay were utilized to investigate the anti-tumor effects of MLO and obtustatin. In addition, some biochemical tests (chemiluminescence-ChL, TBA-test) were applied to elucidate the influence of obtustatin and crude MLO venom on the S-180 sarcoma. The size of tumor was significantly inhibited by MLO venom and obtustatin with the inhibitory rate of 50% and 33% at the doses of 10 µg/mouse and 1mg/kg/day respectively. Both ChL and MDA decrease in the two treated groups. Both obtustatin and MLO venom have an anticancer activity and might be candidates for the treatment of malignant sarcoma. All our results have shown that both obtustatin and MLO venom have an anticancer activity and might be candidates for the treatment of malignant sarcoma.

Phospholipases a2 from Viperidae snakes: Differences in membranotropic activity between enzymatically active toxin and its inactive isoforms.

We describe the interaction of various phospholipases A2 (PLA2) from snake venoms of the family Viperidae (Macrovipera lebetina obtusa, Vipera ursinii renardi, Bothrops asper) with giant unilamellar vesicles (GUVs) composed of natural brain phospholipids mixture, visualized through fluorescence microscopy. The membrane fluorescent probes 8-anilino-1-naphthalenesulfonicacid (ANS), LAUDRAN and PRODAN were used to assess the state of the membrane and specifically mark the lipid packing and membrane fluidity. Our results have shown that the three PLA2s which contain either of aspartic acid, serine, or lysine residues at position 49 in the catalytic center, have different effects on the vesicles. The PLA2 with aspartic acid at this position causes the oval deformation of the vesicles, while serine and lysine-containing enzymes lead to an appreciable increase of fluorescence intensity in the vesicles membrane, wherein the shape and dimensions of GUVs have not changed, but in this case GUV aggregation occurs. LAURDAN and PRODAN detect the extent of water penetration into the bilayer surface. We calculated generalized polarization function (GP), showing that for all cases (D49 PLA2, S49 PLA2 and K49 PLA2) both LAUDRAN and PRODAN GP values decrease. A higher LAURDAN GP is indicative of low water penetration in the lipid bilayer in case of K49 PLA2 compared with D49 PLA2, whereas the PRODAN mainly gives information when lipid is in liquid crystalline phase.

Lipid bilayer condition abnormalities following Macrovipera lebetina obtusa snake envenomation.

Viper bites is an endemic public health problem in Armenia, even in the cities. Human envenomation is often characterized by clotting disorders, hypofibrinogenemia, and local tissue necrosis. In this original study, we assess some changes of cell membranes plastic properties (namely, its microviscosity, thickness, permeability) in a rat envenomation model using the biophysical approaches. We describe the interaction of Macrovipera lebetina obtusa (MLO) venom with giant unilamellar vesicles (GUVs) composed of the native phospholipid mixtures visualized through fluorescent microscopy. GUVs with a mean diameter of 30 µm have a minimum curvature and mimic cell membranes in this respect. The membrane fluorescence probe, ANS and pyrene, were used to assess the state of membrane and specifically mark the phospholipid domains. Independent of their lipid composition, GUVs were enlarged in size as venom-dependent lipid hydrolysis proceeded. Except of the visible morphological changes, ANS and pyrene also allows us to quantify the fluidity changes in the membrane by measuring of the fluorescence intensity. The presence of viper venom in GUVs media reveals a noticeable decreasing of membrane fluidity compare the control, while the binding of fluorophores with GUVs modified by venom lead to appearance of channel activity. These studies also emphasize the importance of a membrane surface curvature for its interaction with enzymatic components of venom.

Molecular events associated with Macrovipera lebetina obtusa and Montivipera raddei venom intoxication and condition of biomembranes.

Studies on the interaction of snake venom and organized lipid interfaces have been conducted using a variety of systems, including BLMs, SUVs and GUVs. The present study was undertaken to elucidate how the plastic properties (namely, its microviscosity, thickness, permeability) of model membranes from native lipids of different tissues of rats change in the course of Macrovipera lebetina obtusa (MLO), Montivipera raddei (MR) and Naja kaouthia (NK) venoms processing. The presence of viper venom in organism leads to increasing of the electrical resistance of BLMs from liver and muscle lipids approximately on a sequence, while the BLMs from brain lipids have not shown noticeable differences of plastic properties compared to the control. Giant unilamellar vesicles (GUVs) with a mean diameter of 30µm have a minimum curvature and mimic cell membranes in this respect. Snake venom was added to the sample chamber before the vesicles were formed. The membrane fluorescence probes, ANS and pyrene, were used to assess the state of the membrane and specifically mark the phospholipid domains. Fluorescent spectra were acquired on a Varian fluorometer instrument. ANS and pyrene allow us to quantify the fluidity changes in the membrane by measuring of the fluorescence intensity. The presence of viper venom in GUVs media reveals a noticeable decreasing of membrane fluidity compared to the control, while the binding of fluorophores with GUVs modified by venom leads to the appearance of channel activity. These studies also emphasize the importance of a membrane surface curvature for its interaction with enzymatic components of venom.