Região metafisária da tíbia de coelhos: uma opção para estudos dos substitutos ósseos / Metaphyseal region of the tibia of rabbits: an option for bone substitutes studies

Apresenta-se um modelo experimental de ostectomia de tíbia em coelhos para estudo de biomateriais em processos de reparação óssea. Foi realizada falha segmentar de 6mm de diâmetro na região metafisária medial proximal de tíbia de 72 coelhos as quais foram preenchidas com substituto ósseo e avaliadas clinicamente, por exame radiográfico e por meio de tomografia computadorizada feixe cônico, em diferentes tempos. Conclui-se que a metáfise medial proximal de tíbia de coelhos é adequada como modelo para estudos que avaliem o comportamento de enxertos e/ou biomateriais em falhas ósseas.

Is presented an experimental model of tibial ostectomy in rabbits to study biomaterials during bone repair process. Segmental failure of 6 mm diameter was performed in the medial proximal tibial metaphyseal region of 72 rabbits, which were filled with bone substitute and evaluated by clinical exam, X-ray, and cone beam computed tomography at different times. It is concluded that the medial proximal tibial metaphysis region of rabbits is suitable as a model for studies that evaluate the behavior of grafts and/or biomaterials on bone defects.

BONEFILL ® block as alternative for bone substitute: a toxicological evaluation

ABSTRACT Bone substitutes based on hydroxyapatite (HA) and Bonefill® (BO - inorganic bovine bone) associated with poly(lactic-co-glycolic acid) (PLGA) (HA/PLGA and BO/PLGA) were evaluated concerning cytotoxicity, genotoxicity and mutagenicity as potential candidates for bone repair. The materials were developed and provided by Bionnovation Biomedical Products Ltda. Eluates from these bone substitutes were prepared for toxicity evaluations using eukaryotic cell cultures. HA/PLGA was used as a comparison for Bonefill®. Cell viability was evaluated by XTT assay and surviving fraction was calculated for clonogenic survival. Additionally, tail moment was used to assess genotoxicity (comet assay). The frequencies of binucleated cells with micronucleus (FBMN), micronucleus (FMN), nucleoplasmic bridges (NPBs), and nuclear buds (NBUDs) were analysed by cytokinesis-block micronucleus assay (CBMN assay). Results showed no statistical difference in cell viability compared with negative control (NC) The eluates did not promote delayed cytotoxicity whereas the surviving fraction rate for cultured cells was similar to NC. Furthermore, no genotoxicity or mutagenicity effects were observed for cultured cells with the Bonefill/PLGA and HA/PLGA eluates. In conclusion, the negative cytotoxicity, genotoxicity and mutagenicity results indicate that these bone substitutes presented interesting preliminary results as potential biomaterials for bone repair.

Aspecto morfológico da interface entre o compósito, constituído de quitosana e polimetilmetacrilato, e a falha óssea de tíbia de coelhos / Morphological aspect of the interface between the composite, made of chitosan and polymethylmethacrylate, and bone failure of the rabbit tibia

As afecções ortopédicas com perda de tecido ósseo são um desafio não só na medicina veterinária mas também na medicina humana. Analisou a interface entre compósito, constituído de esferas de quitosana e polimetilmetacrilato em falha óssea (leito receptor) de tíbia de coelhos, por meio de técnica radiológica, avaliação macroscópica e pela microscopia eletrônica de varredura, em diferentes tempos. Foram utilizados 12 coelhos adultos da raça Nova Zelândia, divididos em quatro Grupos Experimentais (E1, n=3; E2, n=3; E3, n=3 e E4, n=3), que tiveram as falhas ósseas das tibiais direitas preenchidas com compósito, e avaliadas no pós-operatório imediato, aos 30, 60 e 90 e 120 dias. Dos compósitos implantados nas tíbias de coelhos, apenas dois permaneceram em seus leitos receptores, enquanto que os demais se encontravam encapsulados no tecido subcutâneo. As esferas de quitosana, presentes nas superfícies dos biomateriais implantados, que mantiveram contato direto com o leito receptor de tíbias de coelhos apresentavam-se preservadas e não integraram ao tecido ósseo. Diante disso, para melhor compreensão do comportamento da quitosana como substituto ósseo, novas pesquisas serão necessárias.(AU)

Orthopedic diseases with bone loss are challenging in both veterinary and human medicine. The aim of this investigation was to analyze and compare the reactions at the interface between the composite, made of chitosan and polymethylmethacrylate, and the bone defect (receptor site) of the rabbits tibia through radiological and microscopic techniques and by scanning electron microscopy, in different periods. Twelve adult New Zealand rabbits were divided into four experimental groups (E1, n=3; E2, n=3; E3, n=3 and E4, n=3), which had the right tibial bone defects filled with the composite, and evaluated in the immediate postoperative, 30, 60, 90 and 120 days. Composite implanted in the tibia of rabbits, only two remained in their beds receivers, while the remaining were encapsulated in the subcutaneous tissue. Spheres of chitosan present in the biomaterial that has been deployed and were in direct contact with the bone defect, were preserved, however, were not integrated into the bone tissue. Therefore, to understand the behavior of chitosan as a bone substitute, further research is needed.(AU)

Avaliação da interação biológica entre compósito de quitosana, colágeno e hidroxiapatita e tecido ósseo ovino / Investigation of biological interactions of chitosan-collagen-hydroxyapatite composite with bone tissue in sheep

As lesões em membros de grandes animais são um desafio para médicos veterinários, uma vez que somente a osteossíntese não garante resultados satisfatórios. Muitos pesquisadores vêm se dedicando ao desenvolvimento e estudo de substitutos ósseos produzidos de materiais naturais, como quitosana, colágeno e hidroxiapatita, que auxiliam na regeneração óssea. Seis ovinos fêmeas da raça Santa Inês foram submetidos a ostectomias unicorticais de sete milímetros de diâmetro na região proximal da superfície dorsomedial dos III/IV metacarpianos. Foi implantado compósito de quitosana, colágeno e hidroxiapatita em um membro torácico para avaliação da biocompatibilidade do material ao tecido ósseo ovino, e no membro contralateral foi reproduzida a mesma técnica, porém foi mantido sem preenchimento, como controle. Após 60 dias do procedimento cirúrgico, realizou-se biópsia óssea na área de interface entre biomaterial/osso (membro com compósito) e tecido neoformado/osso (membro controle), para realização de avaliação histológica do material não descalcificado, por meio de microscopia de luz e microscopia eletrônica de varredura. Na análise histomorfométrica, mediante microscopia de luz, foi possível identificar maior porcentagem de tecido neoformado em membro controle, quando comparado ao membro com compósito (80% e 63,5%, respectivamente; P<0,05). Por meio da microscopia eletrônica de varredura, observou-se invasão da estrutura interna do compósito por tecido ósseo neoformado. Não houve formação de tecido cicatricial, reação de corpo estranho ou resposta inflamatória crônica nas amostras analisadas. Conclui-se que o compósito de quitosana, colágeno e hidroxiapatita, quando implantado em tecido ósseo ovino, apresenta biocompatibilidade e perfil osteocondutor.(AU)

Fracture management poses a great challenge to large animal practitioners. Osteosynthesis alone is often insufficient to provide satisfactory outcomes in large animals; therefore, several research efforts have been made to investigate and develop bone substitutes capable of promoting bone regeneration. Chitosan-collagen-hydroxyapatite composites constitute a promising alternative given their similar composition to bone. Six Santa Inês ewes were submitted to the creation of experimental 7mm wide unicortical defects on the dorsomedial aspect of the proximal III/IV metacarpal bone diaphysis. Limbs were randomly selected for treatment with chitosan-collagen-hydroxyapatite composite or to serve as untreated controls. Biopsy fragments were collected from the bone/new bone or the bone/biomaterial interface (control and treated defects respectively) within 60 days of surgery; composite biocompatibility was assessed using light and scanning electron microscopy. Histomorphometric analysis under light microscopy revealed greater percentage of new bone tissue in control compared to treated defects (80% and 63.5% respectively; P<0.05). No scar tissue formation, foreign body or chronic inflammatory reactions were observed. Scanning electron microscopy revealed invasion of the composite by new bone tissue. The chitosan-collagen-hydroxyapatite composite studied is biocompatible with bone and shows osteoconductive properties in sheep.

Morphological and chemical analysis of bone substitutes by scanning electron microscopy and microanalysis by spectroscopy of dispersion energy

This study evaluated the morphological and chemical composition of the following bone substitutes: cancellous and cortical organic bovine bone with macro and microparticle size ranging from 1.0 to 2.0 mm and 0.25 to 1.0 mm, respectively; inorganic bovine bone with particle size ranging from 0.25 to 1.0 mm; hydroxyapatite with particle size ranging from 0.75 to 1.0 mm; and demineralized freeze-dried bone allograft with particle size ranging from 0.25 to 0.5 mm. The samples were sputter-coated with gold in an ion coater, the morphology was observed and particle size was measured under vacuum by scanning electron microscopy (SEM). The chemical composition was evaluated by spectroscopy of dispersion energy (EDS) microanalysis using samples without coating. SEM analysis provided visual evidence that all examined materials have irregular shape and particle sizes larger than those informed by the manufacturer. EDS microanalysis detected the presence of sodium, calcium and phosphorus that are usual elements of the bone tissue. However, mineral elements were detected in all analyzed particles of organic bovine bone except for macro cancellous organic bovine bone. These results suggest that the examined organic bovine bone cannot be considered as a pure organic material.

Neste estudo foram avaliados a morfologia, o tamanho e a composição química dos seguintes substitutos ósseos: osso bovino orgânico cortical e esponjoso com micropartículas medindo entre 0,25 e 1,0 mm e macropartículas medindo entre 1,0 e 2,0 mm; osso bovino cortical inorgânico com partículas medindo entre 0,25 e 1,0 mm; hidroxiapatita com partículas medindo entre 0,75 e 1,0 mm; e osso humano descalcificado, congelado e seco medindo entre 0,25 a 0,5 mm. Para a analise da morfologia e tamanho das partículas, as amostras foram preparadas em porta-espécime, metalizadas em ouro e analisadas a vácuo em microscopia eletrônico de varredura (MEV). Para a análise da composição química, as partículas não foram metalizadas e foram analisadas por microanálise por espectroscopia por dispersão de energia (EDS). A análise em MEV, demonstrou que as partículas substitutos ossos apresentaram formato irregular e tamanho variável, maior do que o mencionado pelo fabricante. A microanálise por EDS detectou a presença de elementos como sódio, cálcio e fósforo, que são comuns à composição do tecido ósseo, porém revelaram a presença de elementos químicos nas partículas de osso bovino orgânico, exceto para a macropartícula de osso bovino orgânico esponjoso. Esses resultados sugerem que o osso bovino orgânico não pode ser considerado um material orgânico puro.

Estudo experimental de implantes bioabsorviveis de poli(L-acido latico) / poli(oxido de etileno) associados ao cloridrato de vancomicina no reparo osseo / An experimental study of the use of bioabsorbable polyl(L-lactic acid) / poly(ethylene oxide) microspheres containing vancomycin for bone repair

A necessidade da utilização de enxertos naturais e/ou materiais sintéticos para auxiliar no reparo ósseo é diretamente proporcional a perda tecidual nas lesões. A administração de substâncias antibióticas é necessária para prevenir, ou mesmo combater a ação de agentes bacterianos que possam vir a retardar o reparo tecidual. Este estudo teve como objetivo avaliar a aplicação de um polímero bioabsorvível associado à uma droga antobiótica na regenereção óssea. Foram realizados implantes ósseos de microesferas da blenda de poli(L-ácido lático)PLLA/poli(óxido de etileno)PEO na composição 80:20 associadas ao cloridrato de vancomicina e não associados ao cloridrato de vancomicina em ratos. Os implantes foram colocados em cavidades de 3 mm de diâmetro realizadas em tíbias de ratos da linhagem Wistar. Grupos com 5 animais cada. foram submetidos a períodos experimentais de 2 e 4 dias e 1, 2, 4, 8, 16 e 32 semanas. Os achados morfológicos foram semelhantes em ambos grupos. Houve primeiramente a formação de malha de fibrina e hemorragia ao redor das microesferas, as quais foram gradualmente sendo substituídas por tecido de granulação. A partir do quarto dia, houve a formação inicial de matriz óssea envolvendo as microesferas centripetamente, tornando-se mais evidente e madura da primeira até a trigésima segunda semana de implantação. A comparação entre os achados histomorfométricos...

The use of natural graft and synthetic materials to help bone regeneration is directly relative to cause of bone injury and bone requirement to compose a graft. The antibiotics drugs management is necessary to prevent and combat bacterial agents that could retard the tissue repair The aim of this study was to evaluate the bioabsorbable polymeric implants antibiotic associated behavior during the bone healing. Poly(L-lactic acid)PLLA/poly(ethylene oxide)PEO microspheres blends 80:20 vancomicyn associated bone implants, was compared with PLLA/PEO blend without vancomycin. The implants were inserted in a 3 mm proximal tibiae defect in adult Wistar rats. Periods from 2 and 4 days and 1. 2, 4, 8 16 and 32 weeks were evaluated in 5 animals per group. The histological findings were similar among groups. A fibrin net and hemorrhage were observed primarily around the microspheres and both were progressively replaced by granulation tissue. In four-day implant, the initial bone formation around microspheres was noted. The growth of bone tissue was initially characterized by wolven bone with progressive maturation to lamellar bone, centripetally to microspheres group. The quantity of new bone growth, measured by histomorphometric method and semi-quantitative analysis showed no differences between groups in each experimental interval. Therefore we conclude that mixing vancomycin chloridrate into PLLA/PEO microspheres did not affect...

Sterilisation effect study on granular hydroxyapatite (HA)

Hydroxyapatite is a calcium phosphate bioceramic that has been shown by many authors to be biocompatible with bioactive properties. It is widely accepted as the best synthetic material available for surgical use as a bone graft substitute. HA granules produced by AMREC-SIRIM from local materials underwent 5 types of sterilisation techniques with different ageing periods. Samples were tested for chemical and phase composition and microbial contamination before and after being sterilised. From the microbiological tests done, none of the unsterilised positive control yielded a positive culture. Results from X-Ray diffraction studies found that all the sterilisation techniques did not chemically degrade or structurally change the HA granules significantly.

Quantitative analysis of the selection of materials for surgical implants

Surgical implant materials are used in repair of many anatomical systems in the body. Replacement of large portions of bones and a few other organs with metallic, ceramic, peramic, polymeric or composite prostheses is common. Mechanically, implants are relatively simple; however, biocompatibility requirements are difficult to meet. In the present study, a new quantitative concept for materials selection will be applied to the selection of materials for hip joints. The study starts with an analyses for the implant material requirements followed by a classification of implant materials id common use, Weighing factors are then allocated to the different materials requirement using a digital logic approach. The implant are evaluated on the basis of the above analysis. The results show that a titanium alloy of composition T[1] 6-A1 - 4v is most suitable followed by Co-Cr alloy and then cold worked 136 stainless steel The titanium alloy has good mechanical properties, excellent corrosion resistance and light weight. If the implant 316 stainless materials is intended for temporary use, the analysis 31b stainless shows that steels are most suitable