Desenvolvimento, caracterização e avaliação in vivo de um filme de quitosana- sinvastatina sobre a superfície do titânio: avaliação do potencial de osseointegração sob a influência de nicotina
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Data
2023-08-14
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Universidade Estadual Paulista (Unesp)
Resumo
A osseointegração sob influência sistêmica de nicotina (NIC) apresenta-se como um desafio à implantodontia atual, visto que a capacidade de reparo ósseo mediado por células residentes é negativamente influenciada por esse fatore. Neste projeto de pesquisa, objetiva-se desenvolver um filme de quitosana (CH)-sinvastatina (SV) sobre a superfície do titânio de forma a modular e otimizar o processo de reparo peri-implantar em modelos de comprometimento por NIC. Diferentes técnicas de funcionalização com quitosana (direta ou por meio de aminosilano [APTMS]) e incorporação de sinvastatina nas concentrações de 1, 2,5, e 5 μM foram aplicadas para obtenção de maior estabilidade e potencial modulatório do filme. Para caracterização da superfície, espectroscopia de fotoelétrons excitados por raios X (XPS), microscopia eletrônica de varredura (MEV) e ângulo de contato foram utilizados. O Sucesso na adsorção de CH e SV foi confirmado pelas alterações elementares observadas através de XPS. Todos os filmes apresentaram estrutura nanofibrilar e super-hidrofilicidade. Células tronco da medula ósseo suína (pBMSC) foram utilizadas para mapeamento inicial. Os filmes obtidos através da adsorção de CH por meio de APTMS e incorporação de SV nas concentrações de 1 e 2,5 μM (principalmente) apresentaram a melhor capacidade de induzir a deposição de matriz mineralizada por pBMSCs. APTMS-CH_SV 2,5 μM foi testada in vitro em cultura de SAOS-2 em meio suplementado com NIC. O filme testado foi capaz de bio- estimular as células ainda que na presença de nicotina (quando comparado com controles não funcionalizados). Sendo assim, o filme foi testado em modelo in vivo de administração sistêmica de NIC. Os efeitos deletérios da NIC no processo de reparo ósseo peri-implantar ao redor de implantes tratados por jateamento e ataque ácido (convencionais; NF) foi reafirmado. Assim como o filme de CH_SV se provou eficaz em reverter o impacto negativo mediado por NIC no processo de osseointegração. O maior contato osso/implante (BIC) e área de tecido ósseo peri-implantar (BAFO) em CH_SV quando comparado com controle (NF) na presença de nicotina foi suportado pelos mecanismos de regulação positiva da proteína morfogenética óssea (BMP)-2 e BMP-7, angiogênese (regulação positiva do fator de crescimento endotelial vascular [VEGF]), renovação acelerada (osteocalcina), modulação da inflamação (expressão reduzida do fator de necrose tumoral [TNFα] e interleucina [IL] 1β) e redução da reabsorção óssea mediada por osteoclastos (expressão da fosfatase ácida resistente ao tartarato [TRAP]). Também, a maior formação óssea em período inicial (15 dias), que não diferiu com o período pós-osseointegração (45 dias) indica que o filme acelerou a formação óssea, tanto na presença quanto na ausência de NIC. O filme CH_SV é uma escolha eficaz para aumentar a taxa de sucesso de reabilitações mediadas por osseointegração sob influência sistêmica de NIC e amplia o escopo de aplicação de revestimentos bioativos na superfície de Ti.
The osseointegration under systemic modification by nicotine (NIC) remains a great challenge to implantology since the bone reparative potential of resident cells is negatively influenced in the presence of this factor. In this research project, the objective is to develop a chitosan (CH)-simvastatin (SV) film on the surface of titanium in order to modulate and optimize the peri-implant repair process in models of systemic compromise by NIC. Distinct technique for immobilization of CH (directly os by means of an aminosilane [APTMS]) and incorporation of SV at the concentrations of 1, 2.5, and 5 μM were used to obtain greater stability and modulatory capacity. For surface characterization, X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and contact angle measurement were applied. The success in CH and SV adsorption were confirmed by the elemental changes observed through XPS. All films exhibited nanofibrillar super hydrophilic characteristics. Porcine bone marrow stem cells (pBMSCs) were used for initial screening. The films obtained by adsorption of CH through APTM and incorporation of SV at 1 and e 2,5 μM (specially) showed the best capacity to induce mineralized matrix deposition by pBMSCs. APTMS-CH_SV 2.5 μM was tested in vitro in SAOS-2 cultures in medium supplemented with NIC. The tested film exerted a bio-stimulatory potential in the presence of NIC (as compared in no-functionalized controls). Therefore, the model was tested in an in vivo model of systemic delivery of NIC. The hazardous effects of NIC on peri-implant bone healing around sandblasted acid etched implants (conventional, NF) were reaffirmed. The CH_SV film proved to be effective in reverting the impact of NIC on peri-implant bone healing. The greater bone-to-implant contact (BIC) and bone area fraction occupancy (BAFO) in CH_SV, when compared with control (NF) in the presence of NIC was underlined by the mechanisms of upregulation of bone morphogenetic protein (BMP)-2 and BMP-7, coupled angiogenesis (upregulation of vascular endothelial growth factor [VEGF]), accelerated turnover (osteocalcin), modulation of inflammation (reduced expression of tumor necrosis factor [TNFα] and interleukin [IL] 1β), and reduced osteoclast-mediated bone resorption (tartrateresistant acid phosphatase [TRAP] expression). Additionally, the increased bone formation in initial stages (15 days) did not differ from post-osseointegration phases (45 days), thus indicating that the osseointegration process was accelerated by CH_SV in the presence or absence of NIC. CH_SV film is an effective choice for widening success rate of osseointegration-mediated rehabilitations under systemic influence of NIC and enlarges the application scope of bioactive coatings on the surface of Ti.
The osseointegration under systemic modification by nicotine (NIC) remains a great challenge to implantology since the bone reparative potential of resident cells is negatively influenced in the presence of this factor. In this research project, the objective is to develop a chitosan (CH)-simvastatin (SV) film on the surface of titanium in order to modulate and optimize the peri-implant repair process in models of systemic compromise by NIC. Distinct technique for immobilization of CH (directly os by means of an aminosilane [APTMS]) and incorporation of SV at the concentrations of 1, 2.5, and 5 μM were used to obtain greater stability and modulatory capacity. For surface characterization, X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and contact angle measurement were applied. The success in CH and SV adsorption were confirmed by the elemental changes observed through XPS. All films exhibited nanofibrillar super hydrophilic characteristics. Porcine bone marrow stem cells (pBMSCs) were used for initial screening. The films obtained by adsorption of CH through APTM and incorporation of SV at 1 and e 2,5 μM (specially) showed the best capacity to induce mineralized matrix deposition by pBMSCs. APTMS-CH_SV 2.5 μM was tested in vitro in SAOS-2 cultures in medium supplemented with NIC. The tested film exerted a bio-stimulatory potential in the presence of NIC (as compared in no-functionalized controls). Therefore, the model was tested in an in vivo model of systemic delivery of NIC. The hazardous effects of NIC on peri-implant bone healing around sandblasted acid etched implants (conventional, NF) were reaffirmed. The CH_SV film proved to be effective in reverting the impact of NIC on peri-implant bone healing. The greater bone-to-implant contact (BIC) and bone area fraction occupancy (BAFO) in CH_SV, when compared with control (NF) in the presence of NIC was underlined by the mechanisms of upregulation of bone morphogenetic protein (BMP)-2 and BMP-7, coupled angiogenesis (upregulation of vascular endothelial growth factor [VEGF]), accelerated turnover (osteocalcin), modulation of inflammation (reduced expression of tumor necrosis factor [TNFα] and interleukin [IL] 1β), and reduced osteoclast-mediated bone resorption (tartrateresistant acid phosphatase [TRAP] expression). Additionally, the increased bone formation in initial stages (15 days) did not differ from post-osseointegration phases (45 days), thus indicating that the osseointegration process was accelerated by CH_SV in the presence or absence of NIC. CH_SV film is an effective choice for widening success rate of osseointegration-mediated rehabilitations under systemic influence of NIC and enlarges the application scope of bioactive coatings on the surface of Ti.
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Palavras-chave
Nicotina, Osseointegração, Estatinas (Agentes cardiovasculares), Osseointegration