ABSTRACT A study with transmission electron microscopy of mycoplasma-contaminated HeLa cells using five cell donors referred to as donors A, B, C, D and E, observations are herein presented. Experiments performed with cells from donors B, C and D, revealed the presence of Mycoplasma hyorhinis after PCR and sequencing experiments. Bacteria probably originated from a cytoplasm with compacted tiny granular particles replacing the normal cytosol territories, or from the contact with the cytoplasm through a clear semi-solid material. The compact granularity (CG) of the cytoplasm was crossed by stripes of smooth and rough endoplasmic reticulum cisternae. Among apparently normal mitochondria, it was noted, in variable proportions, mitochondria with crista-delimited lucent central regions that expand to and occupied the interior of a crista-less organelle, which can undergo fission. Other components of the scenarios of mycoplasma-induced cell demolition are villus-like structures with associated 80-200 nm vesicles and a clear, flexible semi-solid, process-sensitive substance that we named jam-like material. This material coated the cytoplasmic surface, its recesses, irregular protrusions and detached cytoplasmic fragments. It also cushioned forming bacteria. Cyst-like structures were often present in the cytoplasm. Cells, mainly apoptotic, exhibiting ample cytoplasmic sectors with characteristic net-like profile due to adjoined vacuoles, as well as ovoid or elongated profiles, consistently appeared in all cells from the last four cell donors. These cells were named “modified host cells” because bacteria arose in the vacuoles. The possibility that, in some samples, there was infection and/or coinfection of the host cell by another organism(s) cannot be ruled out.

ABSTRACT OBJECTIVE: To analyze and to quantify morphological acinar postmortem changes in rat sublingual glands (SLG). MATERIAL AND METHODSs: Fifty rats were divided into two groups of 25 animals each. Group I was used for morphological and morphometric evaluations and group II for the determination of gland density and processed gland volume. Acinar autolytic changes were studied at 0 (control group), 3, 6, 12 and 24 h postmortem periods. The morphometric analysis of the volume density (Vv) and total volume (Vt) of intact (ia) and autolyzed (aa) acini was performed under light microscopy using a Zeiss II integration grid with 100 symmetrically distributed points. RESULTS: Morphologically, temporal progressive nuclear alterations and gradual loss of the structural architecture of acinar cells were found. Regarding quantitative results, both the Vvaa and the Vvia showed statistically significant differences among all postmortem periods (p<0.05). Vvaa increased from 0.42% at 0 h to 75.84% at 24 h postmortem and Vvia decreased from 71.16% to 0% over the same period. For Vtaa and Vtia, no statistically significant differences occurred between 12-24 h and 0-3 h (p>0.05), respectively. Vtaa increased from 0.18 mm³ at 0 h to 38.17 mm³ at 12 h, while Vtia showed a decrease from 33.47 mm³ to 0 mm³ between 3-24 h postmortem. Data concerning Vtaa were adjusted by two-variable linear regression, obtaining the equation: y=-3.54 + 3.38x (r²=0.90). The Vtaa growth rate calculated by this equation was 3.38 mm³/h between 0-12 h. CONCLUSION: Acinar autolysis on rat SLG demonstrated the most significant signs during the first 6 h postmortem and was widely spread through the gland at 12 h.

O objetivo deste estudo foi analisar morfometricamente a resposta tecidual a um pino obtido a partir de osso bovino desvitalizado cortical (DBCB pinos) implantado no tecido subcutâneo de ratos, bem como para avaliar o seu aspecto microestrutural por microscopia eletrônica de varredura (MEV). Os pinos foram implantados no tecido subcutâneo de 20 ratos, que foram sacrificados aos 7, 14, 28 e 60 dias (5 animais / período) após a implantação. No tecido subcutâneo, o pino DBCB promoveu a formação de uma cápsula fibrosa. Aos 7 dias, a cápsula apresentou espessura de 70 ± 3,2 μm com maior densidade de capilares neoformados e menor densidade de fibras colágenas. Entre 14 e 60 dias, a cápsula fibrosa apresentava-se mais organizada e exibiram menor espessura (53 ± 5,5 μm) com maior densidade de fibroblastos e fibras colágenas. Nesse período, foi observada uma bioreabsorção pequena e lenta dos pinos DBCB por macrófagos e raras células gigantes multinucleadas, sem dano tecidual. A espessura dos pinos DBCB reabsorvidos foi em média de apenas 9,3 µm. Durante todos os períodos experimentais não ocorreu presença de células como linfócitos e células plasmáticas. Concluiu-se que o pino derivado de osso bovino cortical foi bem tolerado pelo tecido subcutâneo de ratos e reabsorvido lentamente, sendo um potencial material alternativo para fixação da membrana nos procedimentos de regeneração tecidual guiada.

The aim of this study was to morphometrically analyze the tissue response to a customized pin obtained from devitalized bovine cortical bone (DBCB-pin) implanted in the subcutaneous tissue of rats, as well as to assess its microstructural aspect by scanning electron microscopy (SEM). The pins were implanted in the subcutaneous tissue of 20 rats, which were killed at 7, 14, 28 and 60 days (5 rats/period) after implantation. In the subcutaneous tissue, DBCB-pin promoted the formation of a fibrous capsule. At 7 days, capsule showed thickness of 70 ± 3.2 µm with higher density of newly formed capillaries and smaller density of collagen fibers. Between 14 and 60 days, more organized fibrous capsule exhibited smaller thickness (53 ± 5.5 µm) with higher density of fibroblasts and collagen fibers. In this period, a small and slow bioresorption of the DBCB-pin by macrophages and rare multinucleated giant cells without tissue damage was observed. The thickness of DBCB-pin resorbed was in mean only of 9.3 µm. During all experimental periods not occurred presence of immune reaction cells as lymphocytes and plasma cells. It was concluded that the pin derived from cortical bovine bone was well tolerated by subcutaneous tissue of rats and slowly resorbed could be an alternative material for membrane fixation in the guided tissue regeneration procedures.

Intramucosal inserts made of zirconia ceramic have been used for the rehabilitation of edentulous patients. This study aimed to follow up on patients with complete dentures having intramucosal zirconia inserts and to perform a histological analysis of biopsies of the epithelium surrounding the inserts. Twelve 31-66-year-old subjects of both genders received complete denture treatment having the intramucosal inserts in place. Detailed clinical, prosthetic and surgical procedures were described. Clinical exams were done on postoperative days 3, 7, 15, 120 and 360. One year after denture placement, the subjects received local anesthesia and the tissues surrounding the insertion sites in the alveolar ridge were removed using a scalpel blade. Biopsies were fixed for up to 48 h in 10% phosphate-buffered formaldehyde and 5-µm-thick sections were cut and stained with hematoxylin and eosin and Gomori Trichrome. Immunohistochemistry was used to identify endothelium (anti-CD3) and T lymphocytes (anti-CD31). Removing and reinserting the denture was painful until day 15, but all patients reported a marked increase in the retention and stability of the complete denture with little or no discomfort after 30 days. The histopathological analysis showed that zirconia inserts were well tolerated by the oral mucosa, with the presence of collagen fibers in the tissue around the insert, with mild inflammatory response and allowing reepithelialization, expressed by parakeratosis, epithelial hyperplasia and presenting granular layer. In conclusion, intramucosal zirconia inserts did not affect the health of oral mucosa and provide adequate retention and stability of the complete denture and comfort to the patients.

The aim of this study was to evaluate histologically, three types of calcium sulfate - Merck (Brazil), Surgiplaster (Italy) and Capset (USA) - in surgically created defects on rabbit femurs. Twenty male New Zealand rabbits were used. Two surgical bone defects (5 mm diameter x 8 mm depth) were created on each distal epiphysis using a #3 Dentoflex trephine bur. Defects were filled with: group 1 - di-hydrated calcium sulfate (Merck); group 2 - Capset (Lifecore-USA); group 3 - Surgiplaster (Classimport-Italy); group 4 - control (blood clot). The animals were sacrificed 30, 60, 90 and 180 days postoperatively. Semi-serial 6-mm-thick sections were obtained, stained with hematoxylin and eosin and examined under light microscopy. Bone defects treated with calcium sulfate exhibited new bone formation regardless of the product trademark.

OBJETIVO: Investigar a influência de Proteínas Morfogenéticas Ósseas de origem bovina (bBMPs) ligadas a hidroxiapatita mais colágeno na consolidação de fraturas instáveis do rádio. MÉTODOS: Em 15 coelhos com aproximadamente 5,5 meses de idade e peso médio de 3,5kg foi realizada uma fratura transversa na porção média da diáfise do rádio de ambos os membros. Na fratura do rádio direito foi aplicada mistura de bBMPs ligadas à hidroxiapatita (bBMP-HA) e colágeno bovino como aglutinante e na do rádio esquerdo, considerada controle, nenhum tratamento foi usado. Os coelhos (cinco por período) foram submetidos à eutanásia aos 30, 60 e 90 dias após a cirurgia para realização do processamento histológico e análise microscópica. RESULTADOS: A análise histológica descritiva revelou que a consolidação foi similar para os membros tratado e controle. Pela análise histomorfométrica, a área de novo osso foi em média 867442,16 mm², 938743.00 mm² e 779621,06 mm² para os membros controles e 841118,47 mm², 788038,76mm² e 618587,24 mm² para os membros tratados, aos 30, 60 e 90 dias, respectivamente. Desta forma, aos 60 dias de pós-operatório a área de novo osso foi 12.17% maior no membro tratado com bBMP-HA/colágeno em relação ao membro controle (p<0.05, teste de Tukey). Em ambos os membros a área de novo osso aumentou durante o período experimental até a total consolidação da fratura. CONCLUSÃO: Baseado nos resultados obtidos foi possível concluir que a mistura de bBMP-HA/colágeno induziu pequena, porém significante melhora na consolidação da fratura.

PURPOSE: To investigate the effect of bovine bone morphogenetic proteins (bBMPs) bound to hydroxyapatite plus collagen in the healing of unstable radius fractures. METHODS: A transverse fracture was induced at the mid of the diaphysis in both radii on 15 Norfolk rabbits with average age of 5.5 months and 3.5kg. A mixture of bBMPs bound to thin powdered hydroxyapatite (bBMP-HA) and bovine collagen as agglutinant was applied to the right radius fracture site. The left radius fracture was considered control and no treatment was used. After 30, 60 and 90 days (5 rabbits/period) the rabbits were euthanized and the radii were collected for histological analysis. RESULTS: The descriptive histological analysis revealed that repair was similar for both forelimbs. The histomorphometric analysis showed that the mean area of newly formed bone was 867442.16 mm², 938743.00 mm² and 779621.06 mm² for the control forelimbs, and 841118.47 mm², 788038.76mm² and 618587.24 mm² for the treated forelimbs at 30, 60 and 90 days, respectively. Thus the newly formed bone area was 12.17% larger in the forelimbs treated with bBMP-HA/collagen than in the control forelimbs (p<0.05, Tukey test) in the 60-day period after surgery. In both forelimbs the newly formed bone area increased throughout the experimental period until the complete fracture healing. CONCLUSION: Based on the result obtained here we concluded that bBMP-HA/collagen induced a lower but significant improvement in fracture consolidation.

Little information is available on the pathogenesis of fluorosis during the fetal and initial postnatal period. In the present study, female rats received 0 (control), 7 or 100 ppm of sodium fluoride in drinking water, one week before breeding and throughout gestation and nursing periods. The hemimandibles of the offspring were collected at 0, 7 and 14 days of postnatal life (n = 5) and processed for morphological analyses by light and electron microscopy, immunohistochemical analysis for amelogenin and morphometric study of enamel matrix and ameloblasts of incisors. The results showed a decrease in matrix production at the secretory phase at all study periods for the 100 ppm group. In this same group, the secretory ameloblasts showed reduction of enamel matrix secretion, disorganization of mitochondrial crests, large vacuoles at the apical portion of the cytoplasm, retention of intracisternal material and dilatation of some cisterns in the rough endoplasmic reticulum. In the groups of animals aged 7 and 14 days, analysis of variance showed significant reduction (p<0.05) in cytoplasmic volume of 23.80% and 24.75%, respectively, in relation to the control group. The smooth-ended maturation ameloblasts exhibited a large number of vacuoles with electron-dense endocytic matrix, suggesting a delay in the resorption process. Immunohistochemical analysis showed no difference in the intensity and labeling pattern of the enamel matrix in any study group. Interestingly, in offspring at the age of 14 days for the 7 ppm group, there was an increase in the matrix length at the secretory phase. Therefore, part of the excessive dose of sodium fluoride given to the mother in drinking water can reach the offspring through the placenta and mother's milk, causing morphological changes in ameloblasts and suggesting a reduction in secretion and a delay in matrix resorption.

O tratamento do osso bovino com peróxidos e agentes caotrópicos visa à obtenção de uma matriz óssea acelular, com manutenção do complexo colágeno-apatita e maior resistência mecânica, denominada osso bovino misto (OBM). O objetivo desse estudo foi avaliar a citocompatitiblidade do osso bovino misto e a interação célula-OBM. Células Balb-c 3T3 foram tratadas com diferentes concentrações do extrato hidrossolúvel do OBM (0,1g por mililitro de meio de cultura sem soro). Foram avaliadas a morfologia e número de células, a capacidade de redução do metil tetrazol e a incorporação de vermelho neutro, de acordo com o padrão ISO 10993-5. A interação células-OBM foi estudada por microscopia eletrônica de varredura. Os extratos hidrosolúveis do OBM levaram à morte celular, possivelmente devido à redução da função mitocondrial e aumento da permeabilidade de membrana. Células plaqueadas diretamente sobre o OBM não sobreviveram. Entretanto, após diálise e condicionamento com DMEM +10% de soro fetal bovino, as células aderiram e proliferaram sobre o material. Com base nos resultados obtidos, concluiu-se que o OBM é citotóxico. A citotoxicidade foi revertida por diálise resultando em um material adequado para terapia celular no campo da bioengenharia.

Treatment of bovine bone with peroxides and chaotropic agents aims to obtain an acellular bone matrix that is able to maintain the collagen-apatite complex and a higher mechanical resistance, a mixed biomaterial hereby named mixed bovine bone (MBB). The purpose of this study was to evaluate the cytocompatibility of MBB and cell-MBB interaction. Cell morphology, number of viable cells, ability to reduce methyltetrazolium and to incorporate neutral red upon exposure to different concentrations of the hydrosoluble extract of MBB were assessed in Balb-c 3T3 cells according to ISO 10993-5 standard. The interaction between cells and MBB surface was evaluated by scanning electron microscopy. The water-soluble MBB extracts were cytotoxic and led to cell death possibly due to its effect on mitochondrial function and membrane permeability. Cells plated directly onto the MBB did not survive, although after dialysis and material conditioning in DMEM + 10% FCS, the cells adhered and proliferated onto the material. It may be concluded that, in vitro, water-soluble MBB extracts were cytotoxic. Nevertheless, MBB cytotoxic effect was reverted by dialysis resulting in a material that is suitable for cell based-therapy in the bioengineering field.

OBJETIVO: Avaliar a capacidade osteo-regenerativa de dois biomateriais utilizando um modelo de defeito segmentar efetuado nas diáfises do rádio de coelhos. MÉTODOS: O defeito direito foi preenchido com pool de proteínas morfogenéticas ósseas (pBMPs) e hidroxiapatita em pó ultrafina absorvível (HA) combinada com matriz óssea inorgânica desmineralizada e colágeno, derivados do osso bovino (Grupo A). O defeito esquerdo foi preenchido com matriz óssea desmineralizada bovina com pBMPs e hidroxiapatita em pó ultrafina absorvível (Grupo B). Em ambos os defeitos utilizou-se membrana reabsorvível de cortical bovina desmineralizada para reter os biomateriais no defeito ósseo e guiar a regeneração tecidual. Os coelhos foram submetidos à eutanásia aos 30, 90 e 150 dias após a cirurgia. Foram efetuados exames radiográficos, tomográficos e histológicos em todos os espécimes. RESULTADOS: Aos 30 dias de pós-cirúrgico, o osso cortical desmineralizado foi totalmente reabsorvido em ambos os grupos. A HA tinha reabsorvido nos defeitos do Grupo A, mas persistiu nos do Grupo B. Uma reação de corpo estranho foi evidente com ambos os produtos, porém mais pronunciada no Grupo B. Aos 90 dias os defeitos do grupo B tinham mais formação óssea que os do Grupo A. Entretanto, aos 150 dias após a cirurgia, nenhum tratamento havia promovido o completo reparo do defeito. CONCLUSÃO: Os biomateriais testados contribuíram pouco ou quase nada para a reconstituição do defeito segmentar.

PURPOSE: To evaluate the osteo-regenerative capacity of two proprietary bone grafting materials, using a segmental defect model in both radial diaphyses of rabbits. METHODS: The right defect was filled with pooled bone morphogenetic proteins (pBMPs) bound to absorbable ultrathin powdered hydroxyapatite (HA) mixed with inorganic and demineralized bone matrix and bone-derived collagen, derived from bovine bone (Group A). The left defect was filled with bovine demineralized bone matrix and pBMPs bound to absorbable ultrathin powdered HA (Group B). In both groups, an absorbable membrane of demineralized bovine cortical was used to retain the biomaterials in the bone defects, and to guide the tissue regeneration. The rabbits were euthanized 30, 90 and 150 days after surgery. Radiographic, tomographic and histologic evaluations were carried out on all specimens. RESULTS: At 30 days, the demineralized cortical bone cover was totally resorbed in both groups. HA was totally resorbed from Group A defects, whereas HA persisted in Group B defects. A prominent foreign body reaction was evident with both products, more pronounced in sections from Group B. At 90 days, the defects in Group B exhibited more new bone than Group A. However, at 150 days after surgery, neither treatment had stimulated complete repair of the defect. CONCLUSION: The partial bone healing of the segmental defect occurred with low or none performance of the biomaterials tested.

O crescimento das glândulas parótidas do camundongo durante 7 e 35 dias de vida pós-natal foi estudado por métodos morfométricos. A massa glandular, o volume de cada compartimento morfológico e o número de células em cada compartimento foram avaliados. Os dados obtidos para cada dimensão avaliada foram ajustados por equação exponencial, tipo Y = a.e k.x, permitindo o cálculo do seu tempo de duplicação médio (T D), ou seja, uma estimativa da sua velocidade de crescimento. A análise dos resultados mostrou a ocorrência de um marcante aumento de massa glandular no período estudado de 1.424%, com T D = 7,10 dias. Esse crescimento glandular ocorreu por aumentos nos volumes absolutos dos ácinos, dos ductos intercalares, dos ductos estriados, dos ductos excretores e do estroma, com aumentos percentuais de, respectivamente, 3.048%, 417%, 2.662%, 2.594% e 367%, e T Ds de 5,62, 11,71, 5,55, 5,47 e 14,45 dias. A análise da evolução do número de células em cada compartimento demonstrou aumentos de, respectivamente, 1.904%, 285%, 1.228%, 1.090% e 286% e T Ds de 6,62, 20,40, 7,19, 7,26 e 14,51 dias. Baseados nos resultados aqui obtidos, concluímos que o crescimento das glândulas parótidas do camundongo entre os dias 7 e 35 de idade ocorre por intenso acúmulo de células, principalmente nos ácinos e nos ductos estriados e excretores, com uma velocidade de crescimento sensivelmente maior que nos ductos intercalares e no estroma.

The growth of the mouse parotid glands during 7 and 35 days of postnatal life was studied by morphometric methods. The mass of the gland, the volume of each morphological compartment, and the cell number in each compartment were evaluated. The data obtained for each evaluated dimension were adjusted by an exponential equation, of the type Y = a.e k.x, thus permitting the calculation of their mean duplication time (T D), i.e., an estimation of their growth rate. Analysis of the results showed a marked 1,424% increase in the gland mass during the whole studied period, with T D = 7.10 days. This growth occurred by increases in absolute volume of acini, intercalated ducts, striated ducts, excretory ducts and stroma, with percentual increases of 3,048%, 417%, 2,662%, 2,594% and 367%, respectively, and T Ds of 5.62, 11.71, 5.55, 5.47 and 14.45 days, respectively. Analysis of the cell number growth in each compartment showed increases of 1,904%, 285%, 1,228%, 1,090% and 286%, respectively, and T Ds of 6.62, 20.40, 7.19, 7.26 and 14.51 days, respectively. Based on the present results, we concluded that the growth of the mouse parotid glands from day 7 to day 35 of age occurred by intense cell accumulation, mainly in the acini, striated ducts and excretory ducts, with a growth rate sensibly higher than that of the intercalated ducts and stroma.

O enxerto ósseo ideal deve possuir características como biocompatibilidade, capacidade osteocondutora, osteoindutora, resistência e plasticidade. Dentre os implantes xenogênicos de origem bovina, os produzidos com o osso esponjoso revestem-se de particular interesse devido a sua arquitetura constituída de poros biologicamente desenhados que favorecem a invasão celular e vascular até o centro do defeito. O objetivo deste estudo foi avaliar a resposta tecidual ao material de osso inorgânico medular bovino macrogranular. Quarenta ratos foram divididos em 2 grupos (n=20): o grupo controle recebeu cápsulas de colágeno vazias, e o grupo experimental recebeu implante subcutâneo do material teste. As amostras foram coletadas após 10, 20, 30 e 60 dias de implantação e processadas histotecnicamente. A análise histológica mostrou aos 10 dias pós-cirúrgicos que o infiltrado inflamatório era do tipo granulomatoso rico em células gigantes multinucleadas, mas livre de linfócitos ou plasmócitos, quadro similar ao observado para aloenxertos mineralizados implantados em subcutâneo de ratos. Com o avançar do tempo experimental houve significante diminuição do infiltrado inflamatório inicial concomitantemente ao aumento no grau de fibrosamento ao redor das partículas implantadas. Concluiu-se que o material de enxerto testado em tecido conjuntivo subcutâneo de ratos induziu um granuloma tipo corpo estranho e fibrose ao redor das partículas implantadas, resposta semelhante do mesmo tecido aos aloenxertos mineralizados, e não desencadeou nenhuma resposta imune, sendo portanto biocompatível.

The ideal bone graft must present biocompatibility, osteoconductive and osteoinductive properties, resistance and plasticity. Xenogenic grafts of bovine cancellous bone origin are particularly interesting due to their biologically designed porous structure that enhance both cellular and vascular invasion. The purpose of this study was to evaluate the tissue response induced by bovine macrogranular porous anorganic bone implanted in rat subcutaneous tissue. Forty rats were assigned to 2 groups, as follows: the control group received empty collagen capsules and the test group received subcutaneous implants of the test material. Samples were collected after 10, 20, 30 and 60 days and processed histologically. Histological analysis showed at 10 days a granulomatous inflammatory infiltrate, rich in multinucleated giant cells and free of lymphocytes or plasma cells, similarly to mineralized allograft implanted in rat subcutaneous. In later periods, there was a significant decrease in the inflammatory infiltrate and an increase in fibrosis around graft particles. In conclusion, the test material induced a foreign body-type granuloma with subsequent fibrosis around the graft particles implanted in rat subcutaneous and did not elicit any immune response, thus being considered biocompatible.

A resposta tecidual ao osso inorgânico bovino medular implantado em subcutâneo de rato foi avaliada por análise subjetiva através de microscopia de luz. Quarenta ratos foram divididos em 2 grupos: controle (cápsulas vazias de colágeno) e teste (cápsulas de colágeno contendo 0,1g do biomaterial) e mortos 10, 20, 30 e 60 dias após a implantação. Histologicamente, aos 10 dias, observou-se infiltrado inflamatório crônico composto por macrófagos e Células Gigantes Multinucleadas Inflamatórias (IMGC). Neutrófilos, plasmócitos e linfócitos estavam presentes de maneira discreta, desaparecendo durante o processo de reparo tecidual. A porosidade do material foi preenchida pelo tecido conjuntivo reacional mostrando as IMGC. A fibrose foi mais intensa aos 60 dias e evidentemente superior ao grupo controle. Entretanto, o material não causou reações adversas severas, não estimulando a resposta imunológica. Baseado nos resultados encontrados, concluímos que o osso inorgânico bovino medular foi bem tolerado pelo tecido conjuntivo de rato.

The tissue response to porous bovine anorganic bone implanted in rat connective tissue was evaluated by subjective light microscopy analysis. Forty rats were divided into two groups: control (empty collagen capsules) and test (collagen capsule filled with 0.1g biomaterial) and killed 10, 20, 30 and 60 days after implantation. At 10 days, intense chronic inflammatory infiltrate consisting mainly of macrophages and inflammatory multinucleated giant cells (IMGC) was observed. Neutrophils, plasma cells and lymphocytes were present in discrete amounts and slowly disappeared along the repair process. Porosity of the material was filled by reaction connective tissue exhibiting IMGC. The fibrosis was more intense after 60 days and clearly higher than the control group. Thus, the material did not cause any severe adverse reactions and did not stimulate the immune system. Based on the results it could be concluded that deproteinized bovine cancelous bone was well tolerated by rat connective tissue.

O trabalho teve por objetivo avaliar a capacidade de barreira da membrana de cortical óssea desmineralizada liofilizada bovina, testando sua permeabilidade e integridade, em associação com a membrana de pericárdio bovino liofilizado. Em 15 ratos Wistar, machos, adultos, implantou-se no subcutâneo da região cérvico-torácica dorsal a combinação de duas membranas de cortical óssea desmineralizada tendo no interior a membrana de pericárdio bovino. Os animais (cinco ratos/grupo) foram submetidos à eutanásia aos 15, 30 e 60 dias de pós-operatório. A avaliação microscópica mostrou que, aos 15 dias de pós-cirúrgico, as membranas estavam íntegras, com exceção de pequenas áreas de reabsorção da membrana de cortical óssea junto aos locais referentes aos antigos canais nutritivos. Aos 30 dias, havia apenas restos da membrana de cortical óssea e, aos 60 dias, a sua total ausência. Baseados nos resultados aqui obtidos, é possível concluir que a membrana de cortical óssea bovina é rapidamente absorvida e não confere proteção à membrana de pericárdio bovino liofilizado.

The aim of this study was to evaluate the barrier capacity of the bovine demineralized lyophilized bone cortical membrane, testing its permeability and its integrity in association with lyophilized bovine pericardium membrane. Fifteen male adult Wistar rats were used. The combination of two bone cortical membranes with bovine pericardium membrane inside was implanted subcutaneously in the cervical-thoracic area. The animals (five rats per group) were submitted to euthanasia at 15, 30 and 60 days postoperatively. On day 15 postoperative, the microscopic evaluation showed integrity of the membranes, except the absorption of small areas of the bone cortical membrane in the sites of nutritive channels. On day 30, there was a remnant of bone cortical membrane, and on day 60 bone cortical membrane was completely absent. It was possible to conclude that bovine demineralized lyophilized bone cortical membrane is quickly absorbed and does not preserve the lyophilized bovine pericardium membrane.

A capacidade de um pool de proteínas morfogenéticas bovinas ligada a hidroxiapatita sintética absorvível microgranular (BMPb-HA) em estimular a reparação óssea foi avaliada em defeito de tamanho crítico em calvária de rato. Defeitos de 8 mm diâmetro foi produzido na calvária de 25 ratos, sendo 15 preenchidos com BMPb-HA homogeinizado com sangue (grupo experimental) e 10 somente com coágulo sanguíneo (grupo controle). As calvárias dos animais do grupo experimental foram coletadas 1, 3 e 6 meses após as cirurgias e as do grupo controle, ao término da cirurgia e após 6 meses. Os resultados morfométricos obtidos nas imagens radiográficas apresentou ausência de formação óssea nos períodos de 1 e 3 meses e, histologicamente os defeitos estavam preenchidos por tecido conjuntivo fibroso e inúmeros focos de reação granulomatosa, tipo corpo estranho, ao redor de aglomerados de hidroxiapatita. Ao final de 6 meses, os focos de reação granulomatosa diminuiu em número e tamanho e os defeitos diminuíram a sua área em 22% em relação ao controle 0 hora, devido a neoformação óssea nas suas bordas, mas apresentaram área semelhante ao controle 6 meses. Concluímos que a utilização de BMPb-HA no tratamento de defeitos ósseos cranianos em ratos, promove a formação de reação granulomatosa tipo corpo estranho que inibe profundamente a neoformação óssea, sugerindo que a hidroxiapatita sintética microgranular não é um bom carreador para as BMPs expressarem o seu potencial indutor.

The ability of a pool of bovine bone morphogenetic proteins bound to synthetic microgranular hydroxyapatite (BMPb-HA) to stimulate bone repair was determined in rat critical size defects. An 8-mm diameter defect was created in the calvaria of 25 rats. In 15 rats, the defects were filled with BMPb-HA homogenized with blood (experimental group), and in 10 rats the defects were filled only with blood clots (control). The calvariae of experimental rats were collected 1, 3 and 6 months after surgery and of the control rats at the end of surgery and 6 months thereafter. The morphometric results obtained in the radiographs showed an absence of new bone formation at 1 and 3 months post-surgery and, histologically, the defects were filled with fibrous connective tissue and numerous foci of a foreign body-type granulomatous reaction around hydroxyapatite agglomerates. At the end of 6 months, the number and size of the granulomatous foci decreased and the area of the defects was reduced by 22% compared to the 0-hour control due to the formation of new bone at their borders, although the mean area was similar to the 6-month control. We conclude that the use of BMPb-HA in the treatment of critical size bone defects of the rat skull leads to the formation of a foreign body-type granulomatous reaction that markedly inhibits new bone formation, suggesting that synthetic microgranular hydroxyapatite does not represent a good carrier for BMP-induced bone formation.

Nos últimos anos, vários materiais de origem bovina para enxerto ósseo têm sido lançados no comércio para serem utilizados em cirurgias bucomaxilofaciais. Na presente pesquisa avaliamos a capacidade de um material de enxerto ósseo orgânico em bloco (Gen-ox, Baumer SA - Brasil) preparado à partir de osso medular bovino, em promover a reparação de lesões ósseas de tamanho crítico em calvária de ratos. Uma lesão trans-óssea de aproximadamente 8mm de diâmetro foi realizada com trefina cirúrgica nos ossos parietais de 25 ratos, sendo que em 15 os defeitos foram preenchidos com bloco de 8mm de diâmetro do material de enxerto, embebida com sangue do próprio animal e em 10 somente com coágulo sanguíneo. As calvárias dos ratos tratados com o material foram coletadas após 1, 3 e 6 meses após as cirurgias e dos tratados com coágulo sanguíneo, imediatamente após as cirurgias e decorridos 6 meses. Durante as cirurgias verificamos que esse material enxertado é de fácil manuseio e se adapta perfeitamente ao leito receptor após sua embebição com sangue. Os resultados obtidos mostraram que na maioria dos casos tratados, o material foi reabsorvido lentamente e serviu como material de preenchimento e mantenedora de espaço, favorecendo a angiogênese, migração e adesão celular e a neoformação óssea à partir das bordas da lesão. No entanto, em 2 casos no subgrupo tratado - 1 mês e 1 caso no subgrupo tratado - 3 meses, ocorreu a presença de reação granulomatosa tipo corpo estranho com a presença de inúmeras células gigantes, que inibiu a neoformação óssea local. Esses casos foram interpretados como decorrentes de falhas na desmineralização e na retirada de potenciais fatores antigênicos durante a produção do biomaterial. Concluímos que com um aprimoramento do controle de qualidade na linha de produção do material, a matriz orgânica medular em bloco poderá ser uma boa alternativa para ser usada no reparo de defeitos ósseos na região buco-maxilofacial, devido a sua alta capacidade osteocondutora.

Over the last few years, various bone graft materials of bovine origin to be used in oromaxillofacial surgeries have entered the market. In the present study, we determined the capacity of a block organic bone graft material (Gen-ox, Baumer SA, Brazil) prepared from bovine cancellous bone to promote the repair of critical size bone injuries in rat calvaria. A transosseous defect measuring approximately 8mm in diameter was performed with a surgical trephine in the parietal bone of 25 rats. In 15 animals, the defects were filled with a block of graft material measuring 8mm in diameter and soaked in the animal's own blood, and in the other 10 animals the defects were only filled with blood clots. The calvariae of rats receiving the material were collected 1, 3 and 6 months after surgery, and those of animals receiving the blood clots were collected immediately and 6 months after surgery. During surgery, the graft material was found to be of easy handling and to adapt perfectly to the receptor bed after soaking in blood. The results showed that, in most animals treated, the material was slowly resorbed and served as a space filling and maintenance material, favoring angiogenesis, cell migration and adhesion, and bone neoformation from the borders of the lesion. However, a foreign body-type granulomatous reaction, with the presence of numerous giant cells preventing local bone neoformation, was observed in two animals of the 1-month subgroup and in one animal of the 3-month subgroup. These cases were interpreted as resulting from the absence of demineralization and the lack of removal of potential antigen factors during production of the biomaterial. We conclude that, with improvement in the quality control of the material production, block organic bone matrix will become a good alternative for bone defect repair in the oromaxillofacial region due to its high osteoconductive capacity.