Medicina del futuro en los venenos de animales. Ecuador como fuente importante de investigación

La evolución ha permitido que varios animales produzcan secreciones para su supervivencia: los venenos, aquellos que son dañinos y letales cuando ejercen sus efectos en el organismo, modifican las respuestas normales como por ejemplo la coagulación y el proceso inflamatorio. Presentamos una revisión bibliográfica actualizada de los venenos que pueden ser empleados y así explicar cómo estas sustancias pueden representar una terapéutica en el futuro; de la misma manera describimos la situación de nuestro país que puede ser fuente de investigación en este tema

Potencial terapêutico de um heparinóide isolado do invertebrado marinho Litopenaeus vannamei

The occurrence of bioactive compounds in marine organisms comes awaking the interest of the pharmaceutical industry. Heparin, a sulfated polysaccharide which presence was already identified in several marine invertebrates, is very attractive due its remarkable functional versatility. Besides to intervene in blood coagulation, this molecule has a great anti-inflammatory potential. However, its strong anticoagulant activity difficult the clinical exploitation of its anti-inflammatory properties. Thus, the aims of this work were to evaluate the effect of a heparin-like compound (heparinoid), isolated from the cephalotorax of the Litopenaeus vannamei shrimp, on the inflammatory response, hemostasia and synthesis of antithrombotic heparan sulfate by endothelial cells, besides studying some aspects concerning its structure. The purified heparinoid was structurally characterized following an analytical boarding, involving electrophoresis and chromatography. The structural analysis have shown that this compound possess a high content of glucuronic acid residues and disulfated disaccharide units. In contrast to mammalian heparin, the heparinoid was incapable to stimulate the synthesis of heparan sulfate by endothelial cells in the tested concentrations, beyond to show reduced anticoagulant activity and hemorrhagic effect. In a model of acute inflammation, the compound isolated from the shrimp reduced more than 50% of the cellular infiltration. Besides reduce the activity of MMP-9 and proMMP-2 of the peritoneal lavage of inflamed animals, the heparinoid also reduced the activity of MMP-9 secreted by activated human leukocytes. These results demonstrate the potential of heparinoid from L. vannamei to intervene in the inflammatory response. For possessing reduced anticoagulant activity and hemorrhagic effect, this compound can serve as a structural model to direct the development of more specific therapeutical agents to the treatment of inflammatory diseases

Homogalactanas sulfatadas da alga Codium isthmocladum com atividade anticoagulante

Since the first description of sulfated polysaccharides from seaweeds, the biological activities of these compounds have been evaluated under different aspects and experimental procedures. Among the broad biological activities presented by seaweed polysaccharides, anticoagulant action appears as a promising function. In this present study we have obtained sulfated polysaccharides from the green seaweed Codium isthmocladium by proteolytic digestion, followed by separation into five fractions (0.3, 0.5, 0.7, 0.9 and 1.2) by sequential acetone precipitation. The chemical analyses have demonstrated that all fractions are composed mainly by sulfated polysaccharides. The anticoagulant activity of these fractions was determined by activated partial thromboplastin time (aPTT) and prothrombin time test (PT) using citrate normal human plasma. None fraction has shown anticoagulant activity by PT test. Furthermore, all of them have shown anticoagulant activity by aPTT test. These results indicated that the molecular targets of these sulfated polysaccharides are mainly in the intrinsic via of the coagulation cascade. Agarose gel electrophoresis in 1,3-diaminopropane acetate buffer, pH 9.0, stained with 0.1% toluidine blue showed the presence of two or three bands in several fractions while the fraction 0.9 showed a single spot. By anion exchange chromatography, the acid polysaccharides from the 0.9 acetone fraction were separated into two new fractions eluted respectively with 2.0 and 3.0 M NaCl. These compounds showed a molecular weight of 6.4 and 7.4 kDa respectively. Chemical analyses and infrared spectroscopy showed that Gal 1 and Gal 2 are sulfated homogalactans and differ one from the other in degree and localization of sulfate groups. aPPT test demonstrated that fractions 2,0 and 3,0M (Gal1 and Gal 2, respectively) have anticoagulant activity. This is the first time that anticoagulant sulfated homogalatans have been isolated from green algae. To prolong the coagulation time to double the baseline value in the aPTT, the required amount of sulfated galactan 1 (6,3mg) was similar to low molecular heparin Clexane®, whereas only 0,7mg of sulfated galactan 2 was needed to obtain the same effect. Sulfated galactan 2 in high doses (250mg) induces platelet aggregation. These results suggest that these galactans from C. isthmocladum have a potential application as an anticoagulant drug

Estudo de composto com propriedades funcionais em flor de cardo submetida a diferentes tratamentos de secagem

O cardo é uma planta que encontra-se associada ao fabrico de queijo, pois é responsável pelo fenómeno da coagulação do leite. No entanto, têm aumentado o número de estudos realizados nesta planta, relacionados com a composição química, de forma a perceber os benefícios para a saúde. O objetivo desta dissertação foi caracterizar a flor de cardo relativamente à sua atividade antioxidante, composição em compostos fenólicos e avaliar a sua estabilidade ao longo do trato digestivo. Para tal, foram estudas amostras de flor de cardo da espécie C. cardunculus spp flavescens, após liofilização e secagem em estufa com convecção forçada a diferentes temperaturas (40ºC, 50ºC e 60ºC). Após cada tratamento, efetuaram-se duas extrações sucessivas com soluções de metanol (98% v/v) e de acetona (60% v/v). Os extratos obtidos foram depois utilizados para quantificar os teores em compostos fenólicos totais, em orto-difenois e em flavonóides. A atividade antioxidante foi determinada utilizando os métodos DPPH e ABTS. Por fim, procedeu-se à avaliação da bioacessibilidade dos compostos presentes submetendo os vários extratos a condições simulantes do trato digestivo. Pela análise dos resultados obtidos foi possível constatar que a quantidade de compostos fenólicos e atividade antioxidante na flor de cardo varia consoante a temperatura de secagem, ocorrendo a diminuição à medida que se aumenta a temperatura. Os resultados obtidos para a bioacessibilidade mostraram um comportamento distinto dos extratos de metanol e de acetona. No entanto, em ambos os casos ocorreu uma diminuição na quantidade de compostos disponíveis para absorção intestinal.

Clínica de animais de companhia

O presente relatório apresenta as atividades desenvolvidas no estágio decorrido no Hospital Ars Veterinaria. A primeira parte do relatório refere-se ao relatório de casuística com apresentação de casos e procedimentos acompanhados. A segunda refere-se à monografia com o tema “Opções terapêuticas em hemangiossarcoma canino” e encontra-se ilustrada por três casos clínicos. O hemangiossarcoma é uma neoplasia descrita com origem endotelial, devido à aparência histológica, no entanto descobertas recentes sugerem que provém de células progenitoras hematopoiéticas. Os sinais clínicos são inespecíficos e variáveis. A localização mais comum é o baço, embora ocorra noutras localizações. O diagnóstico requer o estadiamento tumoral e para ser completo deve incluir hematologia e bioquímica sérica, provas de coagulação, exames imagiológicos e confirmação histopatológica. O tratamento base é cirúrgico associado a protocolos quimioterápicos baseados em doxorrubicina. Novas terapias começam a ser investigadas de forma a melhorar os tempos de sobrevivência desta neoplasia de prognóstico reservado; Abstract: Small animal medicine This report was elaborated following a traineeship at the Ars Veterinaria Hospital. The first part concerns about cases report and includes presentation of cases and followed procedures. The second part is the monography under the theme “therapeutical options in canine hemangiosarcoma” and it presents three clinical cases seen during the internship. The hemangiossarcoma is a cancer described to have endothelial origin, because of its histological appearance, but recent discoveries suggest that it originates from hematopoietic progenitor cells. The clinical signs are variable and unspecific. The most common location is the spleen, but others might exist. The diagnosis demands tumoral staging, which to be complete must include hematology and biochemical analysis, coagulation tests, imagiologic tests and histopathologic confirmation. The basic treatment is surgical, associated with chemotherapy based on doxorubicin. New therapies are beginning to be investigated, aiming improve survival times of this cancer with reserved prognosis.

Embryo transfer as a tool to disseminate k-casein BB genotype and improve milk clotting properties

Dairy industries are asked to be increasingly competitive and efficient. Despite the increasing trend in milk yield and protein content during the last decade genetic selection, milk coagulation ability has diminished and even if the absolute amount of cheese produced has increased, the relative cheese yield from a set amount of milk, has decreased. As casein content and variants, along with milk clotting properties (MCP) are determined to a large extent at DNA level, genetic selection and embryo transfer can provide efficacious tools to reverse this trend and achieve improvements. The aim of the proposed research was to determine how rapidly and to what extent milk coagulation properties could be improved by using embryo transfer (ET) as a tool to increase the frequency of k-casein BB genotype cattle and reducing A and E variants in an Italian Holstein herd with a low prevalence of the favourable genotype. In the effort to optimize superovulation protocols and results, synchronization of wave emergence was performed through manual transrectal ablation of the largest (dominant) ovarian follicle on days 7 or 8 of the cycle (estrus = day 0); different drugs and dosage for the superstimulation protocol were experimented trying to overcome the negative effects of stress and the perturbance of LH secretion in superovulated highly producing lactating cows and the use of SexedULTRA™ sex-sorted semen, for artificial insemination of superovulated cows was reported for the first time. The selection program carried out in this research, gave evidence and gathered empirical data of feasible genetic improvements in cheesemaking ability of milk by means of k-casein BB selection. In conclusion, in this project, selection of k-casein BB genotype markedly enhanced cheese-making properties of milk, providing an impetus to include milk coagulation traits in genetic selection and breeding programs for dairy cattle.

Development and optimization of techniques and design parameters for the engineering of atmospheric pressure plasma devices for biomedical applications and plasma medicine

In the last decades, the possibility to generate plasma at atmospheric pressure gave rise to a new emerging field called plasma medicine; it deals with the application of cold atmospheric pressure plasmas (CAPs) or plasma-activated solutions on or in the human body for therapeutic effects. Thanks to a blend of synergic biologically active agents and biocompatible temperatures, different CAP sources were successfully employed in many different biomedical applications such as dentistry, dermatology, wound healing, cancer treatment, blood coagulation, etc.… Despite their effectiveness has been verified in the above-mentioned biomedical applications, over the years, researchers throughout the world described numerous CAP sources which are still laboratory devices not optimized for the specific application. In this perspective, the aim of this dissertation was the development and the optimization of techniques and design parameters for the engineering of CAP sources for different biomedical applications and plasma medicine among which cancer treatment, dentistry and bioaerosol decontamination. In the first section, the discharge electrical parameters, the behavior of the plasma streamers and the liquid and the gas phase chemistry of a multiwire device for the treatment of liquids were performed. Moreover, two different plasma-activated liquids were used for the treatment of Epithelial Ovarian Cancer cells and fibroblasts to assess their selectivity. In the second section, in accordance with the most important standard regulations for medical devices, were reported the realization steps of a Plasma Gun device easy to handle and expected to be mounted on a tabletop device that could be used for dental clinical applications. In the third section, in relation to the current COVID-19 pandemic, were reported the first steps for the design, realization, and optimization of a dielectric barrier discharge source suitable for the treatment of different types of bioaerosol.

Policitemia Vera: scoperta di saggi di biopsia liquida finalizzata a identificare i biomarcatori correlati al rischio trombotico

La Policitemia Vera (PV) è una neoplasia mieloproliferativa con un aumentato rischio di trombosi e di progressione verso la Mielofibrosi. L'infiammazione cronica è comunemente osservata nelle neoplasie mieloproliferative, compresa la PV. La rete infiammatoria, tra le varie componenti, comprende le vescicole extracellulari (EVs), che svolgono un ruolo nella comunicazione cellula-cellula. Inoltre, le componenti microbiche circolanti sono state recentemente indicate come potenziali modificatori dell'infiammazione, della coagulazione e dell’emopoiesi in generale. Qui abbiamo studiato il DNA microbico delle EVs circolanti attraverso. Sangue periferico e feci sono stati raccolti da pazienti con PV (n=38) e da donatori sani (n=30). Le EVs circolanti derivate da megacariociti (MK) e piastrine (PLT) sono state analizzate mediante citometria a flusso. Dopo l'estrazione del DNA microbico dalle feci e dalle EV isolate, è stata sequenziata la regione V3-V4 del 16S rDNA. La percentuale di EVs di MK era ridotta nei pazienti con PV rispetto ai donatori sani. Al contrario, la proporzione di EVs di PLT era aumentata. La PV è stata associata anche a una firma del DNA microbico delle EVs isolate con una maggiore diversità e una composizione microbica distinta rispetto alla controparte sana. Nei pazienti con PV c’è una maggiore proporzione di EVs associate al lipopolisaccaride. Il profilo del microbioma intestinale non differiva tra PV e doantori. Inoltre, l'aumento della proporzione di EVs di MK e la riduzione di EVs di piastre identificavano i pazienti con pregressa trombosi. Le EVs dei pazienti con trombosi erano impoverite di DNA di Staphylococcus ma arricchite di DNA di Actinobacteria e Anaerococcus. Inoltre, questi pazienti avevano livelli più bassi di EVs associate al lipopolisaccaride. I pazienti con fibrosi midollare avevano una maggiore proporzione di PE-EV ed erano arricchite in DNA di Collinsella e Flavobacterium. Questi dati possono contribuire a perfezionare la prognosi della PV e a identificare nuovi bersagli farmacologici.