A new hydroxyapatite-alginate-gelatin biocomposite favor bone regeneration in a critical-sized calvarial defect model

Abstract This study aimed to evaluate the osteogenic potential of hydroxyapatite (HA), Alginate (Alg), and Gelatine (Gel) composite in a critical-size defect model in rats. Twenty-four male rats were divided into three groups: a negative control with no treatment (Control group), a positive control treated with deproteinized bovine bone mineral (DBBM group), and the experimental group treated with the new HA-Alg-Gel composite (HA-Alg-Gel group). A critical size defect (8.5mm) was made in the rat's calvaria, and the bone formation was evaluated by in vivo microcomputed tomography analysis (µCT) after 1, 15, 45, and 90 days. After 90 days, the animals were euthanized and histological and histomorphometric analyses were performed. A higher proportion of mineralized tissue/biomaterial was observed in the DBBM group when compared to the HA-Alg-Gel and Control groups in the µCT analysis during all analysis periods. However, no differences were observed in the mineralized tissue/biomaterial proportion observed on day 1 (immediate postoperative) in comparison to later periods of analysis in all groups. In the histomorphometric analysis, the HA-Alg-Gel and Control groups showed higher bone formation than the DBBM group. Moreover, in histological analysis, five samples of the HA-Alg-Gal group exhibited formed bone spicules adjacent to the graft granules against only two of eight samples in the DBBM group. Both graft materials ensured the maintenance of defect bone thickness, while a tissue thickness reduction was observed in the control group. In conclusion, this study demonstrated the osteoconductive potential of HA-Alg-Gel bone graft by supporting new bone formation around its particles.

Resumo Este estudo teve como objetivo avaliar o potencial osteogênico de um compósito de hidroxiapatita (HA), alginato (Alg) e gelatina (Gel) em um modelo de defeito de tamanho crítico em ratos. Vinte e quatro ratos machos foram divididos em três grupos: um controle negativo sem tratamento (grupo controle), um controle positivo tratado com osso bovino desproteinizado (grupo DBBM) e o grupo experimental tratado com o novo compósito HA-Alg-Gel (grupo HA-Alg-Gel). Um defeito de tamanho crítico (8,5mm) foi feito na calvária dos ratos, e a formação óssea foi avaliada por análise de microtomografia computadorizada in vivo (µCT) após 1, 15, 45 e 90 dias. Após 90 dias, os animais foram eutanasiados e análises histológicas e histomorfométricas foram realizadas. Uma maior proporção de tecido mineralizado/biomaterial foi observada no grupo DBBM quando comparado aos grupos HA-Alg-Gel e controle na análise de µCT durante todos os períodos de análise. Entretanto, não foram observadas diferenças na proporção tecido mineralizado/biomaterial no dia 1 (pós-operatório imediato) em relação aos períodos posteriores de análise em todos os grupos. Na análise histomorfométrica, os grupos HA-Alg-Gel e controle apresentaram maior formação óssea do que o grupo DBBM. Além disso, na análise histológica, cinco amostras do grupo HA-Alg-Gal exibiram espículas ósseas formadas adjacentes aos grânulos do enxerto contra apenas duas das oito amostras do grupo DBBM. Ambos os materiais de enxerto garantiram a manutenção da espessura óssea do defeito, enquanto uma redução da espessura do tecido foi observada no grupo controle. Em conclusão, este estudo demonstrou o potencial osteocondutor do enxerto ósseo de HA-Alg-Gel, promovendo a formação de osso novo ao redor das suas partículas.

Influence of the geometry of nanostructured hydroxyapatite and alginate composites in the initial phase of bone repair

Abstract Purpose: To analyze, histomorphologically, the influence of the geometry of nanostructured hydroxyapatite and alginate (HAn/Alg) composites in the initial phase of the bone repair. Methods: Fifteen rats were distributed to three groups: MiHA - bone defect filled with HAn/Alg microspheres; GrHA - bone defect filled with HAn/Alg granules; and DV - empty bone defect; evaluated after 15 days postoperatively. The experimental surgical model was the critical bone defect, ≅8.5 mm, in rat calvaria. After euthanasia the specimens were embedded in paraffin and stained with hematoxylin and eosin, picrosirius and Masson-Goldner's trichrome. Results: The histomorphologic analysis showed, in the MiHA, deposition of osteoid matrix within some microspheres and circumjacent to the others, near the bone edges. In GrHA, the deposition of this matrix was scarce inside and adjacent to the granules. In these two groups, chronic granulomatous inflammation was noted, more evident in GrHA. In the DV, it was observed bone neoformation restricted to the bone edges and formation of connective tissue with reduced thickness in relation to the bone edges, throughout the defect. Conclusion: The geometry of the biomaterials was determinant in the tissue response, since the microspheres showed more favorable to the bone regeneration in relation to the granules.

Biocompatibility analysis of a novel reabsorbable alloplastic membrane composed of alginate-Capsul / Análise da biocompatibilidade de uma membrana aloplástica reabsorvível composta de alginato-capsul

Objective The aim of this study was to evaluate in vivo the biological response after implantation of a novel alginate-capsule membrane. Methods The material was implanted into subcutaneous tissue of mice (n=15) and after 1, 3 and 9 weeks, the animals were sacrificed and biopsies analyzed with light microscopy, using the stains hematoxylin-eosin, picrosirius and alcian blue pH 2.5. The parameters evaluated were: intensity and kind of inflammatory infiltrate, presence of connective tissue, foreign body reaction, vascularization and biodegradation. Results 1 week after implantation, the following was observed: mixed inflammatory infiltrate, absence of necrosis and beginnings of membrane fragmentation; after 3 weeks, discrete presence of multinuclear giant cells and beginnings of neovascularization; and after 9 weeks there was minor biodegradation associated with the presence of new connective tissue, and persistence of moderate inflammatory reaction observed from beginning to end of the experiment. Conclusion Considering the results obtained, it is possible to conclude that the novel alginate-capsule membrane is partially reabsorbable but with low biocompatibility, requiring more tests to validate its clinical use.

Objetivo Avaliar in vivo a resposta tecidual após a implantação de uma nova membrana de alginato-capsul. Métodos O material foi implantado no tecido subcutâneo de camundongos (n=15) e após 1, 3 e 9 semanas, os animais foram mortos e as biópsias analisadas à microscopia de luz, através de coloração com hematoxilina-eosina, picrosirius e azul de alcian pH 2,5. Os parâmetros avaliados foram: intensidade e tipo de infiltrado inflamatório, presença de tecido conjuntivo, reação de corpo estranho, vascularização e biodegradação. Resultados Após 1 semana da implantação, notou-se infiltrado inflamatório misto, ausência de necrose e início de fragmentação da membrana, em 3 semanas, observou-se presença discreta de células gigantes multinucleadas e início de neovascularização, e em 9 semanas houve pequena biodegradação associada com a presença de novo tecido conjuntivo e persistência de reação inflamatória moderada observada desde o início do experimento. Conclusão Considerando os resultados obtidos concluiu-se que a nova membrana de alginato-capsul é parcialmente reabsorvível, mas com baixa biocompatibilidade, necessitando de mais testes para validar seu uso clínico.