Estudo in vitro e in vivo da superfície de implantes modificados por feixe de laser com e sem deposição de hidroxiapatita : caracterização topográfica, cultura de células e análise histométrica
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Data
2023-06-15
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Universidade Estadual Paulista (Unesp)
Resumo
A proposição deste trabalho foi avaliar o comportamento biológico in vitro e in vivo do tecido ósseo diante das superfícies usinada (MS), modificada por jateamento seguido de condicionamento ácido, disponível comercialmente (SBAS), modificada por feixe de laser (LS) e modificada por feixe de laser com recobrimento de hidroxiapatita (HA) pelo método biomimético sem tratamento térmico (LHS). Foi realizada previamente a microscopia eletrônica de varredura acoplada ao sistema de espectrometria de energia dispersiva de raios X (MEV/EDX) das superfícies e estudo in vitro com o objetivo de avaliar o comportamento da linhagem celular osteogênica murina (MC3T3- E1) e células estromais derivadas da medula óssea (ST2), mediante exposição às superfícies experimentais. Para o estudo in vivo, foram utilizados 20 coelhos Albinus os quais receberam 40 implantes de 3,75x10mm em leitos cirúrgicos fresados na porção medial das tíbias direita e esquerda, sendo um implante de cada superfície em cada tíbia. Nos 10 animais pertencentes ao período de eutanásia de 4 semanas foi realizada a aplicação intramuscular dos fluorocromos calceína e alizarina. As tíbias com os implantes foram submetidas à análise microtomográfica seguido do processamento de inclusão em resina para obtenção de cortes calcificados e posterior análise por microscopia confocal a laser. Foi mensurada a extensão linear de contato entre tecido ósseo mineralizado e superfície do implante (ELCOMI) e área óssea neoformada (AON). Os valores obtidos foram levados à análise de variância e teste t de Tukey, adotando o nível de significância de 5%. O MEV mostrou diferenças entre as superfícies. No estudo in vitro, observou-se que o revestimento de HA não apenas reduziu a viabilidade das células MC3T3-E1 e ST2, como provocou a expressão e liberação de interleucina-6 (IL6) pró-inflamatória. A análise microtomográfica evidenciou diferença estatística na porcentagem de volume ósseo (BV/TV), na qual a superfície LHS foi estatisticamente superior à MS e SBAS no período de 2 semanas e a superfície LS estatisticamente superior à MS no período de 2 semanas. A ELCOMI dos implantes LHS e LS foram estatisticamente superiores a MS e SBAS no período de 2 semanas. No período de 4 semanas, houve diferença estatística entre as superfícies LHS e LS em relação à superfície MS. A análise estatística da AON dos implantes mostrou diferença entre a superfície LHS e MS no período de 2 semanas. Deste modo, conclui-se que a adição de HA na superfície modificada por laser pode provocar uma maior resposta inflamatória e que as modificações experimentais LS e LHS favorecem a deposição de tecido ósseo periimplantar.
The purpose of this study was to evaluate the in vitro and in vivo biological behavior of bone tissue on surfaces machined (MS), modified by blasting followed by acid etching, commercially available (SBAS), modified by laser beam (LS) and modified by laser beam coated with hydroxyapatite (HA) by the biomimetic method without heat treatment (LHS). Scanning electron microscopy coupled to the energy dispersive Xray spectrometry system (SEM/EDX) of the surfaces and an in vitro study were performed in order to evaluate the behavior of the murine osteogenic cell line (MC3T3- E1) and stromal cells derived from bone marrow (ST2), upon exposure to experimental surfaces. For the in vivo study, 20 Albinus rabbits were used, which received 40 implants of 3.75x10mm in surgical beds milled in the medial portion of the right and left tibias, with one implant on each surface in each tibia. In the 10 animals belonging to the 4-week euthanasia period, intramuscular application of the fluorochromes calcein and alizarin was performed. The tibias with the implants were submitted to microtomographic analysis followed by resin inclusion processing to obtain calcified sections and subsequent analysis by confocal laser microscopy. The linear extension of contact between mineralized bone tissue and implant surface (BIC) and newly formed bone area (NBA) were measured. The values obtained were taken to analysis of variance and Tukey's t test, adopting a significance level of 5%. The SEM showed differences between the surfaces. In the in vitro study, it was observed that the HA coating not only reduced the viability of MC3T3-E1 and ST2 cells, but also provoked the expression and release of pro-inflammatory interleukin-6 (IL6). The microtomographic analysis showed a statistical difference in the percentage of bone volume (BV/TV), in which the LHS surface was statistically superior to the MS and SBAS in the 2-week period and the LS surface statistically superior to the MS in the 2- week period. BIC of the LHS and LS implants were statistically superior to MS and SBAS at the 2-week period. In the period of 4 weeks, there was a statistical difference between the LHS and LS surfaces in relation to the MS surface. Statistical analysis of the NBA of the implants showed difference between the LHS and MS surface in the period of 2 weeks. Thus, it is concluded that the addition of HA to the surface modified by laser can provoke a greater inflammatory response and that the experimental modifications LS and LHS favor the deposition of peri-implant bone tissue.
The purpose of this study was to evaluate the in vitro and in vivo biological behavior of bone tissue on surfaces machined (MS), modified by blasting followed by acid etching, commercially available (SBAS), modified by laser beam (LS) and modified by laser beam coated with hydroxyapatite (HA) by the biomimetic method without heat treatment (LHS). Scanning electron microscopy coupled to the energy dispersive Xray spectrometry system (SEM/EDX) of the surfaces and an in vitro study were performed in order to evaluate the behavior of the murine osteogenic cell line (MC3T3- E1) and stromal cells derived from bone marrow (ST2), upon exposure to experimental surfaces. For the in vivo study, 20 Albinus rabbits were used, which received 40 implants of 3.75x10mm in surgical beds milled in the medial portion of the right and left tibias, with one implant on each surface in each tibia. In the 10 animals belonging to the 4-week euthanasia period, intramuscular application of the fluorochromes calcein and alizarin was performed. The tibias with the implants were submitted to microtomographic analysis followed by resin inclusion processing to obtain calcified sections and subsequent analysis by confocal laser microscopy. The linear extension of contact between mineralized bone tissue and implant surface (BIC) and newly formed bone area (NBA) were measured. The values obtained were taken to analysis of variance and Tukey's t test, adopting a significance level of 5%. The SEM showed differences between the surfaces. In the in vitro study, it was observed that the HA coating not only reduced the viability of MC3T3-E1 and ST2 cells, but also provoked the expression and release of pro-inflammatory interleukin-6 (IL6). The microtomographic analysis showed a statistical difference in the percentage of bone volume (BV/TV), in which the LHS surface was statistically superior to the MS and SBAS in the 2-week period and the LS surface statistically superior to the MS in the 2- week period. BIC of the LHS and LS implants were statistically superior to MS and SBAS at the 2-week period. In the period of 4 weeks, there was a statistical difference between the LHS and LS surfaces in relation to the MS surface. Statistical analysis of the NBA of the implants showed difference between the LHS and MS surface in the period of 2 weeks. Thus, it is concluded that the addition of HA to the surface modified by laser can provoke a greater inflammatory response and that the experimental modifications LS and LHS favor the deposition of peri-implant bone tissue.
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Palavras-chave
Implantes dentários, Cicatrização, Nd-YAG lasers, Osseointegração, Osseointegration