Prote??nas associadas ?? infectividade em leishmania (leishmania) amazonensis lainson e shaw, 1972 (kinetoplastida: trypanosomatidae)

• Autor(es): Rocha, Liliane Coelho da
• Data: 2015
• Origem: Oasisbr
• Assunto(s): Infectividade; Proteoma; Leishmania amazonensis; Leishmaniose cut??nea; Infectivity; Proteome; Leishmania amazonensis; Cutaneous leishmaniasis; CI??NCIAS BIOL??GICAS
• Tipo de Documento: Tese de doutoramento
• Idioma: Português

Descrição

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Funda????o de Amparo ?? Pesquisa do Estado do Amazonas

Variations in clinical manifestations of leishmaniasis suggest the existence of a differentiation species dependent in the ability of the Leishmania parasites cause lesions in the host. The search for factors that differentiate the species of this genus and its virulence and infectivity is needed to better understand the mechanisms by which these parasites cause damage to the reservoir hosts, thus enabling the discovery of new tools for immunological and therapeutic potential against leishmaniasis. Although many aspects of immune response to this parasite already known, there are several gaps in knowledge related to the characteristics of the parasite itself, that are related to infection. The in vitro and in vivo assays with axenic promastigotes of Leishmania (Leishmania) amazonensis in logarithmic phase, maintained for long periods in culture, macrophages incubated with strain and inoculated in susceptible mammals, were unable to infect macrophages and hamsters. The loss of infectivity and virulence of promastigotes grown continuously, probably due to selection in the sample initially stable, and infective to a population of non-infective promastigotes. The decoded genome of Leishmania (Leishmania) major and Leishmania (Leishmania) infantum and functional studies of many genes by different research groups, as well as other species of the genus, coupled with the simultaneous advance of proteomics, and accelerated favored significantly study the biology of the genus Leishmania. The use of proteomics technique for the study of proteins related to the infectivity of Leishmania, using 2-DE maps for the detection of protein expression profiles of infected and non infected forms of L. (L.) amazonensis coupled with mass spectrometry ESI-QTof (Electrospray Ionization) for identification of proteins were used in this study. Were detected 251 and 145 spots of proteins differents in promastigotes infective and non-infective, respectively. The two dimensional (2-DE) proteins of L. (L.) amazonensis promastigotes infective and non infective indicated differences in protein expression among the forms studied, revealing the absence of expression of some proteins in the non infective samples. Most of the proteins identified in this study is involved in metabolic processes related to the infectivity and virulence of Leishmania such as: heat shock - HSP83 and HSP70, enzymes - nucleoside diphosphate kinase, protein disulfide isomerase, enolase, trypanothione reductase, mitochondrial tryparedoxin peroxidase and ATPase, cytosolic tryparedoxin and the elongation factor-??. These data confirm the feasibility of doing a sweep of the protein profile of an organism related to its infectivity/virulence protein profiles based on 2-DE. Given the results presented in 2-DE maps and the identification of proteins present in the forms studied we can conclude that the infectivity and virulence of promastigotes of L. (L.) amazonensis is related to a number of factors and proteins whose expression has fundamental importance for the survival/multiplication of the parasite in the reservoir-host

Varia????es nos quadros cl??nicos da leishmaniose sugerem a exist??ncia de uma diferencia????o, dependente da esp??cie e na capacidade de parasitos do g??nero Leishmania causar les??es no hospedeiro. A busca por fatores que diferenciam as esp??cies deste g??nero quanto a sua infectividade e virul??ncia se faz necess??ria para uma melhor compreens??o dos mecanismos pelos quais esses parasitos causam danos aos seus hospedeiros, possibilitando assim a descoberta de novas ferramentas de potencial imunol??gico e terap??utico contra a leishmaniose. Embora muitos aspectos, da resposta imune a esse parasito, j?? sejam conhecidos, existem diversas lacunas no conhecimento relacionadas ??s caracter??sticas do pr??prio parasito que estejam envolvidas com a infec????o. Ensaios in vitro e in vivo com promastigotas ax??nicas de fase logar??tmica de Leishmania (Leishmania) amazonensis, mantidas por longos per??odos em cultivo, incubados com macr??fagos de linhagem e inoculados em mam??feros suscet??veis, mostraram-se incapazes de infectar macr??fagos e hamsters. A perda da infectividade e virul??ncia de formas promastigotas continuamente cultivadas provavelmente ocorreram devido ?? sele????o na amostra inicialmente est??vel e infectiva para uma popula????o de formas promastigotas n??o-infectivas. O genoma decifrado de Leishmania (Leishmania) major e Leishmania (Leishmania) infantum e os estudos funcionais de v??rios genes, por diferentes grupos de pesquisa, bem como de outras esp??cies do g??nero, aliado ao avan??o simult??neo da prote??mica, acelerou e favoreceu de forma significativa o estudo da biologia do g??nero Leishmania. O uso da t??cnica prote??mica para o estudo de prote??nas de Leishmania relacionadas ?? infectividade, atrav??s do uso de mapas 2-DE para a detec????o dos perfis de express??o prot??ica das formas infectivas e n??o infectivas de L. (L.) amazonensis aliado a espectrometria de massas por ESI-QTof (Electrospray Ionization) para identifica????o das prote??nas, foram utilizados nesse estudo. Foram detectados 251 e 145 spots prot??icos diferentes nas formas promastigotas infectiva e n??o infectivas, respectivamente. A an??lise bidimensional (2-DE) de prote??nas de promastigotas de L. (L.) amazonensis infectivas e n??o infectivas indicou diferen??as na express??o prot??ica entre as formas estudadas, revelando a aus??ncia de express??o de algumas prote??nas nas amostras n??o infectivas. A maior parte das prote??nas identificadas neste estudo est?? envolvida em processos metab??licos relacionados ?? infectividade e virul??ncia de Leishmania como as: de choque t??rmico - HSP83 e HSP70, enzimas nucleos??deos difosfato quinase, dissulfeto isomerase, enolase, tripanotiona redutase, triparedoxina peroxidase mitocondrial e ATPase, a triparedoxina citos??lica e os fatores de alongamento ??. Esses dados confirmam a exequibilidade de se fazer uma varredura do perfil prot??ico de um organismo relacionado ?? sua infectividade/virul??ncia baseada nos perfis prot??icos em 2-DE. Em vista dos resultados apresentados nos mapas 2-DE e na identifica????o das prote??nas presentes nas formas estudadas podemos concluir que a infectividade e virul??ncia de promastigotas de L. (L.) amazonensis esta relacionada a um conjunto de fatores e prote??nas cuja express??o ?? de fundamental import??ncia para a sobreviv??ncia/multiplica????o do parasito no seu hospedeiro