Desenvolvimento e avaliação imunológica de Nanotubos de Carbono funcionalizados com a proteína de envelope de Dengue virus 3

• Ano de defesa: 2011
• Autor(a) principal: Ruiz Gerhardt Astigarraga
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Minas Gerais; UFMG
• Palavras-chave em Português: Microbiologia; Nanotubos de carbono; Vírus da dengue
• Link de acesso: http://hdl.handle.net/1843/BUOS-8P4GKP

In terms of epidemiologic impact, Dengue (DENV) infections represent the most important infectious disease in Brazil. Consequently, the development of an efficient vaccine is considered a high priority. Due to this fact, many studies are being developed towards this goal, but no vaccine is currently available to the population. Carbon Nanotubes (CNT) have been broadly studied due to their exceptional properties such as biocompatibility, high aspect ratio and cell internalization ability. Their use has been proposed for many applications such as a delivery system of drugs, proteins and genes to cells, and also presentation of biomolecules to biologic systems, including to the immune system. In this work, we have used the CNT technology to build an experimental immunogen against DENV. CNT functionalized with antigens have immunogenic potential, as shown in previous studies. The DENV envelope (E) glycoprotein is an immunodominant protein that generates serotype-specific protective response upon its presentation to the immune system. The E protein gene from Dengue virus 3 (DENV3E) was cloned in pQE9 vector and expressed in E. coli strain M-15. The produced protein was purified and covalently bound to Multi-Walled CNTs (MWNT) by treatment with diimide-activated amidation, in which amine radicals from the recombinant protein bind to carboxilic portions on the CNT surface. The functionalization efficacy was successfully verified by Raman Spectroscopy and the developed tool was called MWNT-DENV3E. Preliminary immunologic evaluation after 3-dose BALB/C mice immunization showed high antibody titers and antigen-specific lymphocyte proliferation. Clinical signs of infection were verified upon challenge with a mouse-adapted DENV-3 strain after mice immunization with MWNT-DENV3E, such as high viral load in spleen and blood, high MPO activity, high liver transaminase level, high TNF- production and thrombocytopenia. These results suggest an enhancement of infection, once mice separately immunized with DENV3E or MWNT showed significant levels of protection upon challenge, being able to control viral replication. Thus, more studies are needed to clarify these mechanisms and better understand the interactions between DENV immunopathogenesis and the impact of CNT application in biological systems, in order to better evaluate the full potential of this new immunogen.