Snake venomics of the South and Central American Bushmasters. Comparison of the toxin composition of Lachesis muta gathered from proteomic versus transcriptomic analysis

We report the proteomic characterization of the venoms of two closely related pit vipers of the genus Lachesis, L. muta (South American Bushmaster) and L. stenophrys (Central American Bushmaster), and compare the toxin repertoire of the former revealed through a proteomic versus a transcriptomic approach. The protein composition of the venoms of Lachesis muta and L. stenophrys were analyzed by RP-HPLC, N-terminal sequencing, MALDI-TOF peptide mass fingerprinting and CID-MS/MS. Around 30–40 proteins of molecular masses in the range of 13–110 kDa and belonging, respectively, to only 8 and 7 toxin families were identified in L. muta and L. stenophrys venoms. In addition, both venoms contained a large number of bradykinin-potentiating peptides (BPP) and a C-type natriuretic peptide (C-NP). BPPs and C-NP comprised around 15% of the total venom proteins. In both species, the most abundant proteins were Zn2+-metalloproteinases (32–38%) and serine proteinases (25–31%), followed by PLA2s (9–12%), galactose-specific C-type lectin (4–8%), l-amino acid oxidase (LAO, 3–5%), CRISP (1.8%; found in L. muta but not in L. stenophrys), and NGF (0.6%). On the other hand, only six L. muta venom-secreted proteins matched any of the previously reported 11 partial or full-length venom gland transcripts, and venom proteome and transcriptome depart in their relative abundances of different toxin families. As expected from their close phylogenetic relationship, the venoms of L. muta and L. stenophrys share (or contain highly similar) proteins, in particular BPPs, serine proteinases, a galactose-specific C-type lectin, and LAO. However, they dramatically depart in their respective PLA2 complement. Intraspecific quantitative and qualitative differences in the expression of PLA2 molecules were found when the venoms of five L. muta specimens (3 from Bolivia and 2 from Peru) and the venom of the same species purchased from Sigma were compared. These observations indicate that these class of toxins represents a rapidly-evolving gene family, and suggests that functional differences due to structural changes in PLA2s molecules among these snakes may have been a hallmark during speciation and adaptation of diverging snake populations to new ecological niches, or competition for resources in existing ones. Our data may contribute to a deeper understanding of the biology and ecology of these snakes, and may also serve as a starting point for studying structure–function correlations of individual toxins.Ministerio de Educación y Ciencia/[BFU2004-01432/BMC]//EspañaConsejo Superior de Investigaciones Científicas/[CSIC-UCR 2006CR0010]/CSIC-UCR/EspañaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

Radioprotection of Sensitive Rat Tissues by Oligoelements Se, Zn, Mn Plus Lachesis Muta Venom

In this study we first evaluated the general radioprotective efficacy of Se, Zn and Mn (4 μg/ml each) plus Lachesis muta venom (4 ng/ml) combination (O-LM) by determining survival on rats irradiated with lethal doses of gamma-rays. The aim of the second part of the study was to investigate the O-LM ability to prevent ionizing radiation-induced damage on small intestine, bone marrow and submandibular glands. Hence, histological characteristics and functional studies, together with proliferation and apoptotic marker levels on whole body irradiated rats with a 5 Gy dose were evaluated. Results show that all animals of the untreated group died after whole body irradiation with 8 and 10 Gy while 60 day-survival was more than 80% and 40% in O-LM-treated animals, respectively. Histopathological examinations revealed a high degree of small intestine and submandibular gland radioprotection 3 days post-irradiation. O-LM inhibited histological damage on small intestine, restoring the radiation-induced reduction in villous height and crypt number. O-LM prevented radiation-induced loss of salivary gland function and morphological alterations. These effects were associated to a complete inhibition of radiation-induced apoptosis. Furthermore, studies performed 30 days post-irradiation revealed that O-LM significantly improved bone marrow repopulation, increasing all medullar progenies to the extent of the non-irradiated animals, and completely prevented permanent submandibular gland alterations. Based on the present results and taking into account that O-LM is being safely administered in phase I clinical trial as an immunomodulator, we conclude that O-LM is a non-toxic promising approach to achieve radioprotection for patients undergoing radiotherapy.Fil: Crescenti, Ernesto J. V.. Instituto de Inmuno Oncología "Dr. Ernesto J. V. Crescenti"; ArgentinaFil: Medina, Vanina Araceli. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Croci, Máximo. Instituto de Inmuno Oncología "Dr. Ernesto J. V. Crescenti"; ArgentinaFil: Sambuco, Lorena Andrea. Instituto de Inmuno Oncología "Dr. Ernesto J. V. Crescenti"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Prestifilippo, Juan Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Elverdín, Juan Carlos. Universidad de Buenos Aires. Facultad de Odontología; ArgentinaFil: Bergoc, Rosa Maria. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Rivera, Elena Susana. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentin

Histamine prevents radiation-induced mesenchymal changes in breast cancer cells

Radiotherapy is a prime option for treatment of solid tumors including breast cancer though side effects are usually present. Experimental evidence shows an increase in invasiveness of several neoplastic cell types through conventional tumor irradiation. The induction of epithelial to mesenchymal transition is proposed as an underlying cause of metastasis triggered by gamma irradiation. Experiments were conducted to investigate the role of histamine on the ionizing radiation-induced epithelial to mesenchymal transition events in breast cancer cells with different invasive phenotype. We also evaluated the potential involvement of Src phosphorylation in the migratory capability of irradiated cells upon histamine treatment. MCF-7 and MDA-MB-231 mammary tumor cells were exposed to a single dose of 2 Gy of gamma radiation and five days after irradiation mesenchymal-like phenotypic changes were observed by optical microscope. The expression and subcellular localization of E-cadherin, β-catenin, vimentin and Slug were determined by immunoblot and indirect immunofluorescence. There was a decrease in the epithelial marker E-cadherin expression and an increase in the mesenchymal marker vimentin after irradiation. E-cadherin and β-catenin were mainly localized in cytoplasm. Slug positive nuclei, matrix metalloproteinase-2 activity and cell migration and invasion were significantly increased. In addition, a significant enhancement in Src phosphorylation/activation could be determined by immunoblot in irradiated cells. MCF-7 and MDA-MB-231 cells also received 1 or 20 μM histamine during 24 h previous to be irradiated. Notably, pre-treatment of breast cancer cells with 20 μM histamine prevented the mesenchymal changes induced by ionizing radiation and also reduced the migratory behavior of irradiated cells decreasing phospho-Src levels. Collectively, our results suggest that histamine may block events related to epithelial to mesenchymal transition in irradiated mammary cancer cells and open a perspective for the potential use of histamine to improve radiotherapy efficacy.Fil: Galarza, Tamara Ester. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Mohamad, Nora Alicia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Táquez Delgado, Mónica Alejandra. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Vedoya, Guadalupe M.. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Crescenti, Ernesto J.. Instituto de Inmunooncología; ArgentinaFil: Bergoc, Rosa Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Martin, Gabriela Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Cricco, Graciela Patricia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentin

Therapeutic potential of histamine H4 receptor agonists in triple-negative human breast cancer experimental model

The presence of the histamine H₄ receptor (H₄R) was previously reported in benign and malignant lesions and cell lines derived from the human mammary gland. The aim of this work was to evaluate the effects of H₄R ligands on the survival, tumour growth rate and metastatic capacity of breast cancer in an experimental model.Fil: Martinel Lamas, Diego José. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Croci, Máximo. Instituto de Inmunooncología; ArgentinaFil: Carabajal, Eliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; ArgentinaFil: Crescenti, Ernesto J. V.. Instituto de Inmunooncología; ArgentinaFil: Sambuco, Lorena Andrea. Instituto de Inmunooncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Massari, Noelia Andrea. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Bergoc, Rosa Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; ArgentinaFil: Rivera, Elena Susana. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; ArgentinaFil: Medina, Vanina Araceli. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Ingeniería. Departamento de Física. Laboratorio de Radioisótopos; Argentin

Effects of oligoelements Se, Zn, and Mn plus Lachesis muta venom in experimental scleroderma

Abstract: Scleroderma, sclerosis of the skin, is a severe autoimmune disease refractant to all kind of treatments. To study the in vivo effects of a combination of three oligoelements selenium (Se), zinc (Zn), and manganese (Mn) plus Lachesis muta venom (O-LM) on the bleomycin (BLM)-induced scleroderma mouse experimental model. C3H mice were randomly divided into four groups: control (phosphate-buffered saline (PBS)), O-LM, BLM, and BLM + O-LM. All administrations were performed subcutaneously into the back of mice. BLM was injected 5 days per week for three consecutive weeks and O-LM was administered simultaneously with BLM from the beginning of the experiments and lasted for 3 weeks after the final BLM or PBS injection (for O-LM and BLM + O-LM groups), when animals were sacrificed and histopathological, immunohistochemical, thiobarbituric acid reactive species (TBARS) evaluation, and autoantibodies detection were determined. O-LM significantly reduced BLM-induced enhanced dermal thickness (605 ± 47 vs. 956 ± 59 μm, P < 0.01), collagen deposition, and mast cells infiltration (43.1 ± 1.0 vs. 102 ± 14.1 mast cells, P < 0.05). O-LM administration significantly blocked BLM-induced oxidative damage and the enhanced immunoreactive fibroblasts for α-smooth muscle actin while reduced BLM-induced autoantibodies that strongly react mainly with skin and spleen. O-LM significantly reduced BLM-induced scleroderma through the modulation of antioxidant and immunological pathways