PATRICK JACK SPENCER

Resumo

Possui graduação em Ciências Biológicas pela Universidade Presbiteriana Mackenzie (1991), mestrado em Tecnologia Nuclear pela Universidade de São Paulo (1995) e doutorado em Tecnologia Nuclear pela Universidade de São Paulo (2000) tendo sido bolsista sandwich no US Army Medical Research Institute for Infeccious Diseases (98-99). É responsável pelo Biotério de criação e manutenção de animais de laboratório do IPEN. Tem experiência na área de Bioquímica, com ênfase em Proteínas, atuando principalmente nos seguintes temas: veneno, proteínas, bothrops, irradiação e miotoxina.(Texto extraído do Currículo Lattes em 22 dez. 2021)

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Agora exibindo 1 - 10 de 49
  • Artigo IPEN-doc 27113
    Molecular model of cytotoxin-1 from Naja mossambica mossambica venom in complex with chymotrypsin
    2015 - MUNAWAR, AISHA; AKREM, AHMED; HUSSAIN, ASHIQ; SPENCER, PATRICK; BETZEL, CHRISTIAN
    Snake venom is a myriad of biologically active proteins and peptides. Three finger toxins are highly conserved in their molecular structure, but interestingly possess diverse biological functions. During the course of evolution the introduction of subtle mutations in loop regions and slight variations in the three dimensional structure, has resulted in their functional versatility. Cytotoxin-1 (UniProt ID: P01467), isolated from Naja mossambica mossambica, showed the potential to inhibit chymotrypsin and the chymotryptic activity of the 20S proteasome. In the present work we describe a molecular model of cytotoxin- 1 in complex with chymotrypsin, prepared by the online server ClusPro. Analysis of the molecular model shows that Cytotoxin-1 (P01467) binds to chymotrypsin through its loop I located near the N-terminus. The concave side of loop I of the toxin fits well in the substrate binding pocket of the protease. We propose Phe10 as the dedicated P1 site of the ligand. Being a potent inhibitor of the 20S proteasome, cytotoxin-1 (P01467) can serve as a potential antitumor agent. Already snake venom cytotoxins have been investigated for their ability as an anticancer agent. The molecular model of cytotoxin-1 in complex with chymotrypsin provides important information towards understanding the complex formation.
  • Resumo IPEN-doc 26933
    Redução da atividade edematogênica promovida pelo muco da raia de água doce Paratrygon aiereba utilizando soro produzido contra o muco irradiado com raios gama de 60Co
    2016 - THOMAZI, G.O.C.; PREZOTTO NETO, J.P.; ALVES, G.J.; TURIBIO, T.O.; AIRES, R.S.; ROCHA, A.M.; SEIBERT, C.S.; SPENCER, P.J.; NASCIMENTO, N.
    Introdução: As raias são peixes peçonhentos com ampla distribuição geográfica nos principais rios brasileiros e merecem destaque por estarem frequentemente associadas a acidentes em seres humanos. Estes agravos são frequentes na região Norte do país e favorecidos pelo hábito desses peixes de permanecerem em repouso no fundo arenoso ou lamacento de águas rasas e pela frequente utilização humana dos rios, seja por lazer ou atividades ocupacionais. Os ferimentos provocados pelos ferrões das raias são dolorosos, de difícil cicatrização, causam necroses extensas e fenômenos sistêmicos. O muco que recobre toda a extensão do corpo desses peixes pode aumentar a gravidade desses ferimentos. A escassez de estudos voltados para o tratamento específico das lesões oriundas dos agravos por esses peixes, a ausência de estudos com outros gêneros de raias dulcícolas e a possibilidade de produção de soro contra o veneno de raias estimularam o desenvolvimento deste estudo utilizando a radiação ionizante que tem se mostrado vantajosa na atenuação de toxinas animais, resultando na obtenção de melhores imunógenos para a produção de soros. Objetivo: Avaliar e comparar o potencial neutralizante dos soros anti-mucoda raia Paratrygonaierebanativo ou irradiado (2 kGy-60Co) contra a ação edematogênicado muco nativo. Método: Mediante aprovação da CEUA/IPEN/SP n°126/2013 e do ICMBion.º 45407-1/2014 foi desenvolvido este trabalho. O muco (50μg/mL) de P. aierebafoi incubado em banho-maria com os soros anti-muconativo ou soro anti-mucoirradiado diluídos de 1:100 ou 1:1.000 (soros de coelhos imunizados contra o muco nativo ou irradiado por 60Co de P. aiereba). Após a incubação, as amostras foram injetadas (30μL) no coxim plantar da pata posterior de camundongos Swissmachos, 18 a 20g, (n=30). A interferência na atividade edematogênicafoi verificada nos tempos de 1h, 2h, 4h e 24h após a inoculação. Em todos os animais foi verificado o volume inicial individual de cada pata antes da injeção das amostras. Os volumes foram mensurados em pletismômetroe os resultados expressos como a variação do volume em relação ao volume basal em μL por período (ExpBioMed. 239:601, 2014). A avaliação estatística foi realizada pela análise de variância com auxílio do softwareGraphPadPrism5.0. Resultados: O muco de P. aierebafoi capaz de induzir edema de 1h a 4h após a inoculação, com declínio de 4 a 24h (p<0,01 em relação aos controles). O soro anti-muconativo não foi capaz de inibir a formação do edema nas diluições testadas, sem diferença estatística com o edema induzido pelo muco (p>0,05). O soro anti-mucoirradiado diluído 1:100 apresentou interferência significativa na atividade edematogênicanas primeiras quatro horas (p<0,01). O soro anti-mucoirradiado 1:1000 foi capaz de diminuir a formação de edema nas 1ª e 4ª horas (p<0,01). O edema foi reduzido pelo soro anti-mucoirradiado pré-incubado com o muco não irradiado (nativo). Conclusão: Esse resultado mostra que além do muco irradiado ser capaz de estimular a proliferação de células de memória, ou seja, a produção de anticorpos IgGespecíficos, estas imunoglobulinas são capazes de reconhecer a fração responsável pela atividade edematogênica. Estes resultados nos permite concluir que o processo de irradiação tornou o muco da raia P. aierebamais antigênico.
  • Livro IPEN-doc 26714
    O IPEN e a saúde
    2019 - LINARDI, MARCELO; NETTO, ANA P.F.A.; LUGAO, ADEMAR B.; FREITAS, ANDERSON Z. de; CARBONARI, ARTUR W.; ZEITUNI, CARLOS A.; SOARES, CARLOS R.J.; SILVA, CECILIA C.G. e; ZAMBONI, CIBELE B.; PERONI, CIBELE N.; VIEIRA, DANIEL P.; ANDRADE, DELVONEI A. de; ZEZELL, DENISE M.; LAZAR, DOLORES R.R.; PARRA, DUCLERC F.; MOREIRA, EDSON G.; LANDULFO, EDUARDO; FONSECA, EDVALDO R.P. da; PERINI, EFRAIN A.; ARAUJO, ELAINE B. de; MUCCILLO, ELIANA N. dos S.; CARVALHO, ELITA F.U. de; BERNARDES, EMERSON S.; MOURA, ESPERIDIANA A.B. de; SILVA, FABIANA M. da; MOREIRA, FERNANDO J.F.; SILVA, FLAVIA R. de O.; GENEZINI, FREDERICO A.; ALVES, GLAUCIE J.; YORIYAZ, HELIO; COSTA, ISOLDA; MENGATTI, JAIR; ROSSI, JESUALDO L.; OLIVEIRA, JOAO E. de; SARKIS, JORGE E. de S.; PERROTTA, JOSE A.; ROGERO, JOSE R.; SHORTO, JULIAN M.B.; SILVA, LEONARDO G. de A. e; MOLNARY, LESLIE de; RODRIGUES, LETICIA L.C.; DIAS, LIGIA E.M.F.; CALDAS, LINDA V.E.; POZZO, LORENA; GENOVA, LUIS A.; MATSUDA, MARGARETH M.N.; HAMADA, MARGARIDA M.; FELINTO, MARIA C.F. da C.; POTIENS, MARIA da P.A.; ROSTELATO, MARIA E.C.M.; MARUMO, MARIA H.B.; VASCONCELLOS, MARINA B.A.; RIBEIRO, MARTHA S.; COTRIM, MARYCEL E.B.; NEVES, MAURICIO D.M. das; MORALLES, MAURICIO; DIAS, MAURO da S.; IGAMI, MERY P.Z.; DURAZZO, MICHELANGELO; SUZUKI, MIRIAM F.; SAIKI, MITIKO; MATHOR, MONICA B.; VIEIRA JUNIOR, NILSON D.; BARTOLINI, PAOLO; SPENCER, PATRICK J.; SILVA, PAULO S.C. da; AFFONSO, REGINA; CARNEIRO, REGINA C.G.; ROGERO, SIZUE O.; SAKATA, SOLANGE K.; CASTANHO, SONIA R.H. de M.; MAIHARA, VERA A.; ROSSI, WAGNER de; CALVO, WILSON A.P.; KODAMA, YASKO
  • Artigo IPEN-doc 26676
    Perfil clínico e epidemiológico da Leishmaniose Tegumentar Americana no Hospital de Doenças Tropicais da Universidade Federal do Tocantins
    2019 - SILVEIRA, SILVESTRE J.S. da; SPENCER, PATRICK J.
    O objetivo principal deste artigo é caracterizar o Perfil Clínico e Epidemiológico da Leishmaniose Tegumentar Americana, no período de 2010 a 2016, do Hospital de Doenças Tropicais (HDT) da Universidade Federal do Tocantins no município de Araguaína-TO, área considerada endêmica para a referida doença. Como objetivos específicos, pretende-se contribuir para o conhecimento da doença nos aspectos de: a) mostrar a distribuição geográfica regional da LTA; b) caracterizar o perfil epidemiológico regional da LTA; c) retratar as formas clínicas e sua frequência na região; d) descrever os critérios diagnósticos utilizados no HDT; e d) descrever as terapêuticas empregadas para LTA no Serviço e discutir sobre os dados de acordo com a literatura atual, visto que a doença que faz parte da lista de doenças negligenciada, e as mesmas são as principais causas de morbidade e mortalidade em todo o mundo.
  • Resumo IPEN-doc 26600
    Isolamento e caracterização de peptídases do veneno de Pseudechis Australis
    2019 - FREITAS, GUSTAVO G.; SPENCER, PATRICK J.
  • Artigo IPEN-doc 26265
    Characterization and evaluation of the enzymatic activity of tetanus toxin submitted to gamma radiation by Cobalt 60
    2019 - SARTORI, GISELLE P.; COSTA, ANDREA; MACARINI, FERNANDA L.S.; MARIANO, DOUGLAS O.C.; PIMENTA, DANIEL C.; SPENCER, PATRICK J.; GALISTEO JUNIOR, ANDRES J.
    Tetanus is a neurological disease which blocks the inhibitory neurotransmitters liberation. Also treatment does not cure the disease, and its main form of prevention is through vaccination. The vaccine is produced by the inactivation of tetanic toxin (TeNT) with formaldehyde, which may cause side effects. An alternative way is the use of ionizing radiation for inactivation of the toxin and also to improve the potential immunogenic response and to reduce the pos-vaccination side effects. Therefore, the aim of this study was to characterize the TeNT structure after different doses of ionizing radiation of Cobalt 60, and also to assess the enzymatic activity after the radiation. SDS PAGE and MALDI-TOF analysis revealed gradual modification on the TeNT structure according to doses increase. Also, fragmentation and possible aggregations of the protein fragments were observed in higher doses. In the analysis of peptide preservation by enzymatic digestion and mass spectrometry, there was a slight modification in the recognition up to the dose of 4 kGy at subsequent doses, recognition was minimal. The analysis of the enzymatic activity by fluorescence showed a 35 % attenuation in the activity even at higher doses. In the antigenic evaluation performed by ELISA and Western Blot, anti-TeNT antibodies were detected against the irradiated toxins at the different doses, with a gradual decrease as the dose increased, but remaining at satisfactory levels, indicating the possibility of possible use as an immunogen, however, studies of enzymatic activity on higher should be further analyzed.
  • Artigo IPEN-doc 26495
    Proteomic analysis of soluble proteins retrieved from Duttaphrynus melanostictus skin secretion by IEx-batch sample preparation
    2019 - MARIANO, DOUGLAS O.C.; PREZOTTO-NETO, JOSE P.; SPENCER, PATRICK J.; SCIANI, JULIANA M.; PIMENTA, DANIEL C.
    Amphibians display a toxic secretion that works as chemical defenses against predators and/or microorganisms that is stored in specialized glands located in the tegument. For some animals, such glands have accumulated in specific regions of the body and formed prominent structures known as macroglands. The Bufonidae family displays conspicuous macroglands in a post-orbital position, termed parotoids, which secretions are known to be extremely viscous and rich in alkaloids and steroids. Few proteins have been described in this material, though. Mainly, because of the difficulties to handle such biological matrix. In this context, we have performed a proteomic study on the parotoid macrogland secretion of the Asian bufonid Duttaphrynus melanostictus. By employing the Ion-Exchange (IEx)-batch chromatography (anionic, cationic and both) we obtained six fractions - bound and unbound – that were submitted to an in solution-trypsin digestion followed by LC-MS/MS. Proteins related to: antioxidant activity, binding processes (carbohydrate/lipid/protein), energy metabolism, hydrolases, lipid metabolism and membrane traffic were identified. Moreover, IEx was able to preserve the biological activity of the retrieved proteins (peptidasic). The current study increases the knowledge on the proteins present in the bufonids parotoid macrogland secretion, providing a better understanding of the physiological role played by such molecules. Significance: The current approach allowed a detailed proteomic analysis of the several proteins synthesized in the D. melanostictus parotoid macrogland (Bufonidae) that are secreted into the skins, but embedded within a complex viscous biological matrix. Moreover, our results aim to increase the knowledge on the biological role played by such proteins at the skin.
  • Artigo IPEN-doc 26494
    Protein identification from the parotoid macrogland secretion of Duttaphrynus melanostictus
    2019 - MARIANO, DOUGLAS O.C.; MESSIAS, MARCELA D.G.; SPENCER, PATRICK J.; PIMENTA, DANIEL C.
    Background: Bufonid parotoid macrogland secretion contains several low molecular mass molecules, such as alkaloids and steroids. Nevertheless, its protein content is poorly understood. Herein, we applied a sample preparation methodology that allows the analysis of viscous matrices in order to examine its proteins. Methods: Duttaphrynus melanostictus parotoid macrogland secretion was submitted to ion-exchange batch sample preparation, yielding two fractions: salt-displaced fraction and acid-displaced fraction. Each sample was then fractionated by anionic-exchange chromatography, followed by in-solution proteomic analysis. Results: Forty-two proteins could be identified, such as acyl-CoA-binding protein, alcohol dehydrogenase, calmodulin, galectin and histone. Moreover, de novo analyses yielded 153 peptides, whereas BLAST analyses corroborated some of the proteomicidentified proteins. Furthermore, the de novo peptide analyses indicate the presence of proteins related to apoptosis, cellular structure, catalysis and transport processes. Conclusions: Proper sample preparation allowed the proteomic and de novo identification of different proteins in the D. melanostictus parotoid macrogland secretion. These results may increase the knowledge about the universe of molecules that compose amphibian skin secretion, as well as to understand their biological/physiological role in the granular gland.
  • Artigo IPEN-doc 26413
    Biochemical and biological characterization of the Hypanus americanus mucus
    2019 - COELHO, GUILHERME R.; PREZOTTO NETO, PEDRO; BARBOSA, FERNANDA C.; SANTOS, RAFAEL S. dos; BRIGATTE, PATRICIA; SPENCER, PATRICK J.; SAMPAIO, SANDRA C.; D’AMELIO, FERNANDA; PIMENTA, DANIEL C.; SCIANI, JULIANA M.
    Stingrays skin secretions are largely studied due to the human envenoming medical relevance of the sting puncture that evolves to inflammatory events, including necrosis. Such toxic effects can be correlated to the biochemical composition of the sting mucus, according to the literature. Fish skin plays important biological roles, such as the control of the osmotic pressure gradient, protection against mechanical forces and microorganism infections. The mucus, on the other hand, is a rich and complex fluid, acting on swimming, nutrition and the innate immune system. The elasmobranch's epidermis is a tissue composed mainly by mucus secretory cells, and marine stingrays have already been described to present secretory glands spread throughout the body. Little is known about the biochemical composition of the stingray mucus, but recent studies have corroborated the importance of mucus in the envenomation process. Aiming to assess the mucus composition, a new noninvasive mucus collection method was developed that focused on peptides and proteins, and biological assays were performed to analyze the toxic and immune activities of the Hypanus americanus mucus. Pathophysiological characterization showed the presence of peptidases on the mucus, as well as the induction of edema and leukocyte recruitment in mice. The fractionated mucus improved phagocytosis on macrophages and showed antimicrobial activity against T. rubrumç. neoformans and C. albicans in vitro. The proteomic analyses showed the presence of immune-related proteins like actin, histones, hemoglobin, and ribosomal proteins. This protein pattern is similar to those reported for other fish mucus and stingray venoms. This is the first report depicting the Hypanus stingray mucus composition, highlighting its biochemical composition and importance for the stingray immune system and the possible role on the envenomation process.
  • Artigo IPEN-doc 25830
    The amphibian diacylglycerol O-acyltransferase 2 (DGAT2)
    2019 - SCIANI, JULIANA M.; NEVES, ADRIANA; VASSÃO, RUTH C.; SPENCER, PATRICK; ANTONIAZZI, MARTA M.; JARED, CARLOS; PIMENTA, DANIEL C.
    Amphibians are, currently, considered the first vertebrates that had performed the aquatic to terrestrial transition during evolution; therefore, water balance and dehydration control were prerequisites for such environment conquering. Among anurans, Phyllomedusa is a well-studied genus, due to its peptide-rich skin secretion. Here, we have analyzed the skin secretion of Phyllomedusa distincta targeting the proteins present in the skin secretion. The major soluble protein was chromatographically isolated and utilized to immunize rabbits. Through proteomics approaches, we were able to identify such protein as being the diacylglycerol O-acyltransferase 2 (DGAT2), a crucial enzyme involved in lipid synthesis and in the skin water balance. Immunohistochemistry assays revealed the protein tissular distribution for different animal species, belonging to different branches of the phylogenetic tree. Specifically, there was positivity to the anti-DGAT2 on Amphibians’ skin, and no antibody recognition on fish and mammals’ skins. The DGAT2 multiple sequence alignment reveals some degree of conservation throughout the genera; however, there is a different cysteine pattern among them. Molecular modeling analyses corroborate that the different cysteine pattern leads to distinct 3D structures, explaining the different antibody recognition. Moreover, the protein phylogenetic analyses place the Xenopus DGAT2 (the available amphibian representative) next to the Coelacanthus enzyme, which have led the authors to term this a ‘paleo-protein’. DGAT2 would be, therefore, an ancient protein, crucial to the terrestrial environment conquest, with a unique folding—as indicated by the molecular models and immunohistochemistry analyses—a consequence of the different cysteine pattern but with conserved biological function.