Biocerâmica porosa com liberação de fármaco para o tratamento de osteoporose
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Data
2017-08-28
Autores
Costa, Taciane Pereira da [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Neste trabalho scaffolds de hidroxiapatita (HA) foram sintetizados e sua superfície modificada com uma matriz polimérica (copolímero tribloco óxido de polipropileno e óxido de polietileno - PEO-PPO Pluronic®), na qual o fármaco alendronato (AL) foi incorporado, visando à liberação controlada de fármaco e o uso deste biomaterial como substituto ósseo para pessoas com osteoporose. Em uma primeira etapa, o método da dispersão sólida foi utilizado para preparar compósitos de Pluronic® e alendronato. Por meio das análises de difratometria de raios X e calorimetria exploratória diferencial (DSC) chegou-se à conclusão que a melhor amostra foi a F108AL2, que indicava uma melhor interação do polímero com o alendronato. Na segunda etapa, um template de sacrifício de esferas de sílica foi utilizado para preparar a HA porosa. As duas amostras obtidas HA200 e HA400 com tamanho de poros médios de 170nm e 350nm respectivamente foram utilizadas para sintetizar amostras de hidroxiapatita porosa com alendronato, baseando-se nas interações intermoleculares entre os íons cálcio da hidroxiapatita com os íons fosfato e amina do fármaco. As análises de espectroscopia na região do infravermelho e DSC indicaram o sucesso na incorporação do fármaco na HA. Como última etapa, o biocompósito HA porosa - Pluronic® F108 - AL foi preparado. As análises de IV, DSC e microscopia eletrônica de varredura indicaram o sucesso na modificação da superfície da HA com o polímero, bem como a presença do alendronato. Para as amostras de todas as etapas foram realizadas curvas de liberação do AL. Nas amostras Pluronic®-AL, o máximo de liberação foi alcançado em 120 min, com a amostra F108AL2 apresentando o melhor resultado. Já nas amostras HA-AL, mais de 80% do fármaco foi incorporado nas amostras porosas e 24% foi o máximo de liberação alcançado em 72h. Nas amostras do biocompósito HA-F108-AL, o melhor resultado foi observado para a amostra F108HA400AL que apresentou um comportamento distinto das outras, alcançando um máximo de liberação de 80% em 100 minutos. Após a síntese e caracterização dos biocompósitos, a segunda parte deste trabalho foi focada nos testes in vitro. Devido à falta de estabilidade no meio de cultura não foi possível realizar os testes nas amostras HA-F108-AL. Novas amostras foram preparadas baseadas em discos de hidroxiapatita preparados com a amostra HA400. O alendronato foi incorporados nesses discos com duas concentrações diferentes, 0,1mg e 1,0mg. Para as amostras HA, HAAL-0,1 e HAAL-1,0 foram realizados ensaios com osteoclastos. Os testes de atividade da fosfatase ácida tartarato-resistente e alamar blue® mostraram uma menor presença dos osteoclastos nas amostras com maior concentração de alendronato. Com os resultados positivos, um novo biocompósito foi preparado a partir de um gel formado de Pluronic® F127 e alginato juntamente com os discos de HA, duas amostras foram sintetizadas HAGEL e HAGELAL com incorporação de alendronato. Para os ensaios dessas amostras dois tipos de células foram utilizadas, uma co-cultura de osteoclastos e fibroblastos e células mesenquimais. O teste de atividade da fosfatase alcalina com as células mesenquimais indicou que as amostras poderiam estar induzindo à diferenciação destas em osteoblastos, principalmente a amostra HAGELAL. Os resultados indicam um bom potencial dos biocompósitos para uso como substituto ósseo para pessoas com osteoporose.
In this work, hydroxyapatite (HA) scaffolds with surface modified by a triblock copolymer polypropylene oxide and polyethylene oxide (PEO-PPO - Pluronic®) matrix were synthesized for use as alendronate (AL) releaser, with aim to use this biomaterial as a bone graft for people with osteoporosis. In a first step, the solid dispersion method was used to prepare Pluronic® and AL composites and, through X-ray diffractometry and differential scanning calorimetry (DSC) analyzes, F108AL2 was found as the best sample with better interaction between the polymer and AL. In the second step, the porous HA was prepared by a silica spheres sacrificial template. The two samples obtained HA200 and HA400 with average pore size of 170nm and 350nm respectively were used to synthesize HA-AL samples, based on the intermolecular interactions between the HA calcium ions with the AL phosphate and amine ions. Infrared spectroscopy (IR) and DSC analyzes indicated success in drug incorporation. As the last step, the porous HA-Pluronic® F108-AL biocomposite was prepared. Scanning electron microscopy, IV and DSC analyzes indicated the success in the HA's surface modification with the polymer as well as the presence of AL. After the samples characterization of all steps, it was performed AL release curves. In the Pluronic®-AL samples, maximum release was achieved in 120 min, with the F108AL2 sample showing the best result. In the HA-AL samples, more than 80% of the drug was incorporated in the porous samples and 24% was the maximum release reached in 72h. In the HA-F108-AL biocomposite samples, F108HA400AL showed the best result, it was observed a distinct behavior, reached a maximum release of 80% in 100 minutes and remained constant. After the synthesis and characterization of the biocomposites, the second part of this work was focused on the in vitro tests. Due to the lack of stability in the culture medium, the HA-F108-AL samples could not be tested. New samples were prepared based on hydroxyapatite disks prepared with the HA400 sample. Alendronate was incorporated into these disks with two different concentrations, 0.1mg and 1.0mg. For the HA, HAAL-0.1 and HAAL-1.0 samples, osteoclast assays were performed, tartarato-resistant acid phosphatase activity and alamar blue tests showed a lower presence of osteoclasts in the samples with higher concentration of alendronate. From the positive results, a new biocomposite was prepared based in a gel formed of Pluronic® F127 and alginate with the HA disks, two samples were synthesized HAGEL and HAGELAL with incorporation of alendronate. Two cells types were used for the assays, a co-culture of osteoclasts and fibroblasts and mesenchymal cells. The alkaline phosphatase activity test with the mesenchymal cells indicated that the samples could be inducing the differentiation of these into osteoblasts, especially the sample HAGELAL (containing AL). The result indicate a good potential of the biocomposites as a bone graft for people with osteoporosis.
In this work, hydroxyapatite (HA) scaffolds with surface modified by a triblock copolymer polypropylene oxide and polyethylene oxide (PEO-PPO - Pluronic®) matrix were synthesized for use as alendronate (AL) releaser, with aim to use this biomaterial as a bone graft for people with osteoporosis. In a first step, the solid dispersion method was used to prepare Pluronic® and AL composites and, through X-ray diffractometry and differential scanning calorimetry (DSC) analyzes, F108AL2 was found as the best sample with better interaction between the polymer and AL. In the second step, the porous HA was prepared by a silica spheres sacrificial template. The two samples obtained HA200 and HA400 with average pore size of 170nm and 350nm respectively were used to synthesize HA-AL samples, based on the intermolecular interactions between the HA calcium ions with the AL phosphate and amine ions. Infrared spectroscopy (IR) and DSC analyzes indicated success in drug incorporation. As the last step, the porous HA-Pluronic® F108-AL biocomposite was prepared. Scanning electron microscopy, IV and DSC analyzes indicated the success in the HA's surface modification with the polymer as well as the presence of AL. After the samples characterization of all steps, it was performed AL release curves. In the Pluronic®-AL samples, maximum release was achieved in 120 min, with the F108AL2 sample showing the best result. In the HA-AL samples, more than 80% of the drug was incorporated in the porous samples and 24% was the maximum release reached in 72h. In the HA-F108-AL biocomposite samples, F108HA400AL showed the best result, it was observed a distinct behavior, reached a maximum release of 80% in 100 minutes and remained constant. After the synthesis and characterization of the biocomposites, the second part of this work was focused on the in vitro tests. Due to the lack of stability in the culture medium, the HA-F108-AL samples could not be tested. New samples were prepared based on hydroxyapatite disks prepared with the HA400 sample. Alendronate was incorporated into these disks with two different concentrations, 0.1mg and 1.0mg. For the HA, HAAL-0.1 and HAAL-1.0 samples, osteoclast assays were performed, tartarato-resistant acid phosphatase activity and alamar blue tests showed a lower presence of osteoclasts in the samples with higher concentration of alendronate. From the positive results, a new biocomposite was prepared based in a gel formed of Pluronic® F127 and alginate with the HA disks, two samples were synthesized HAGEL and HAGELAL with incorporation of alendronate. Two cells types were used for the assays, a co-culture of osteoclasts and fibroblasts and mesenchymal cells. The alkaline phosphatase activity test with the mesenchymal cells indicated that the samples could be inducing the differentiation of these into osteoblasts, especially the sample HAGELAL (containing AL). The result indicate a good potential of the biocomposites as a bone graft for people with osteoporosis.
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Palavras-chave
Hidroxiapatita, Alendronato, Tribloco PEO-PPO-PEO, Liberação de fármaco, Hydroxyapatite, Alendronate, PEO-PPO-PEO triblock, Drug release