Background: Directing autogenous Mesenchymal Stem Cell (MSC) to defect sites has a great promise in bone regeneration. We designed a MSC specific, bone affinity peptide (E7HA7) by conjugating E7 with a polyglutamate hydroxyapatite (HA) binding motif. We sought to characterize the in-vivo releasing pattern and bioactivity of E7HA7. Methods: HA discs were coated with fluorescent labeled peptides E7HA7, E7HA2 or E7 were subcutaneously implanted in Sprague Dawley rats. In an ectopic bone formation model was used to test the in-vivo bioactivity of E7HA7 conjugated to DBM. Results: E7HA7 showed slower peptide release from scaffolds in comparison to other groups, being statistically significant at week 2 compared to E7, and to E7HA2 at week 4 and 8. In ectopic model, the medians for new bone formation in each group were: iDBM=0.041mm3, iDBM-E7=0.071mm3, aDBM=0.138mm3, and aDBM-E7=0.192mm3. Conclusions: Conjugation of E7 to polyglutamate bone binding domain showed slow releasing kinetics and osteoinductive potential.

No description available.

Demineralized bone matrix (DBM) is an allograft material used in orthopedics that promotes endochondral bone formation. While the placement of DBM on either the periosteal surface of a bone or within a skeletal muscle promotes the recruitment of stem cells that can form skeletal tissues through the temporal progression of endochondral bone development, it remains unclear to what degree these processes are different between the two sites. In this study, we utilize a comparative in vivo model of endochondral ossification by implanting the DBM on the periosteum and in the muscle. Within the muscle we further compared the effects of DBM with and without Bone morphogenetic protein-2 (BMP-2), a primary morphogenetic factor involved in the differentiation of skeletal stem cells. The mice were harvested at various time points after DBM implantation in order to analyze the development of the bone. Analysis included X-ray imaging, microCT imaging, and mRNA expression. Plain x-ray and micro-CT imaging analysis showed mineralized bone formation in the implant on the periosteum and in the muscle with BMP-2, but no growth in the muscle when BMP-2 was not added to the DBM. The mechanisms for bone development were further analyzed by qRT-PCR to determine temporal patterns and levels of expression of various stem cell and differentiated skeletal cell associated genes. The stem cell gene expression varied between implant placement locations suggesting different mechanisms for stem cell recruitment. Interesting, while DBM implants in the muscle without BMP did not induce mineralized tissue specific mRNA expression; specific stem cell and early skeletal cell lineage commitment genes were present. These results suggest that while DBM in muscle is capable of recruiting stem cells that higher BMP-2 levels are needed to promote the progression of cartilage to mineralized bone in muscle tissues.

Angiogenesis is required for endochondral ossification during development and fracture healing; however the exact mechanisms and temporal relationship between the two processes remains unclear. In this study, we utilize an in vivo model of endochondral ossification in mice by implanting demineralized bone matrix (DBM) proximal to the femur to induce ectopic bone formation. TNP-470, a drug known to be anti-angiogenic, was used to inhibit vascularization during the time course of de novo bone formation in order to define the role of angiogenesis during the chondrogenic phase of endochondral bone formation. Day 2, day 8, and day 16 post-surgery were selected time points to represent pre-chondrogenic, chondrogenic, and bone mineralization stages, respectively. Plain x-ray and micro-CT analysis showed that inhibition of angiogenesis led to decreased mineralized tissue formation. Inhibited angiogenesis was confirmed with qRT-PCR. Most striking, however, is that while stem cells are recruited and committed to the chondrogenic lineage, subsequent chondrogenesis failed to progress based on the failure of Sox5 and Sox6 expression, which directs chondrocyte commitment. This expands the role for angiogenesis to a much earlier stage than currently thought and places the necessity of angiogenesis very early in the endochondral ossification process.

A reparação de defeitos ósseos é uma preocupação constante em diversas áreas da odontologia. Vários materiais são usados com a finalidade de ajudar e acelerar esta reparação, tais como enxertos autógenos, xenógenos, membranas e alguns medicamentos, assim como a associação de alguns destes métodos. A sinvastatina, medicamento utilizado para redução de colesterol, em alguns estudos, tem demonstrado ação na estimulação de formação óssea. O objetivo do presente estudo foi avaliar a influência da associação da sinvastatina à matriz desmineralizada de osso bovino na reparação de defeitos ósseos em calvária de ratos. Foram confeccionados defeitos ósseos na calvária de 48 ratos, sendo um defeito 5mm de diâmetro em cada parietal do animal. Os ratos foram divididos em três grupos de acordo com o material utilizado: grupo controle, que não recebeu nenhum tipo de substância, grupo sinvastatina-matriz óssea desmineralizada, que recebeu uma associação de sinvastatina e matriz desmineralizada de osso bovino e grupo matriz óssea desmineralizada que recebeu somente matriz desmineralizada de osso bovino como material de enxertia. Os sacrifícios foram realizados após trinta e sessenta dias. Após o sacrifício as calvárias foram radiografadas em aparelho de raios-X digital para análise de densidade radiográfica em tons de cinza e foram submetidas à preparação histológica de rotina, para posterior análise histológica descritiva e histométrica da área de matriz óssea neoformada, utilizando-se programa computacional para análise de imagens. Os dados submetidos à analise estatística ANOVA a 5% demonstraram que os animais que receberam a associação de sinvastatina-MODB apresentaram, na análise histométrica a menor área de neoformação e na análise radiográfica a menor densidade óssea. Concluiu-se, de acordo com a metodologia utilizada, que a associação sinvastatina-MODB influenciou negativamente o reparo ósseo. / Bone defect healing is a constant concern to several areas of Dentistry. A great variety of materials is currently used to help in the healing process as well as to speed it up. Materials such as autografts, xenografts, membranes, some drugs, and the association of some of them are being used. Simvastatin, a substance used to reduce cholesterol levels, has shown, in some studies, a capacity to stimulate bone formation. The purpose of the present experimental study is to evaluate the influence of the association of sinmvastatin with bovine demineralized bone matrix on the healing of bone defects in rats calvariae. Bone defects were produced in the calvariae of 48 rats, thus each animal had a defect in each parietal bone measuring 5mm of diameter. The rats were grouped according to the graft material used: the control group which didnt receive any substance; the simvastatin-demineralized bone matrix group which received an association of simvastatin and bovine demineralized bone matrix; and the demineralized bone matrix group which received bovine demineralized bone matrix. The animals were sacrificed after thirty or sixty days. After the sacrifices, digital radiographies were taken of the calvariae in order to analyse the radiographic density in shades of gray. They were also submitted to routine histological preparation for future descriptive histological and histomorfometric analyses of the new formed bone matrix through the use of a software to analyse the images. The data submitted to statistical analysis ANOVA (5%) showed that the animals which received the association simvastatin-demineralized bone matrix presented the smallest density and area of new bone formation of the three groups. According to the methodology used, we concluded that the association simvastatin demineralized bone matrix influenced negatively the bone healing.