Mechanosignaling-related angiocrine factors drive osteoblastic phenotype in response to zirconia.

BACKGROUND: The growing use of zirconia as a ceramic material in dentistry is attributed to its biocompatibility, mechanical properties, esthetic appearance, and reduced bacterial adhesion. These favorable properties make ceramic materials a viable alternative to commonly used titanium alloys. Mimicking the physiological properties of blood flow, particularly the mechanosignaling in endothelial cells (ECs), is crucial for enhancing our understanding of their role in the response to zirconia exposure. METHODS: In this study, EC cultures were subjected to shear stress while being exposed to zirconia for up to 3 days. The conditioned medium obtained from these cultures was then used to expose osteoblasts for a duration of 7 days. To investigate the effects of zirconia on osteoblasts, we examined the expression of genes associated with osteoblast differentiation, including Runx2, Osterix, bone sialoprotein, and osteocalcin genes. Additionally, we assessed the impact of mechanosignaling-related angiocrine factors on extracellular matrix (ECM) remodeling by measuring the activities of matrix metalloproteinases 2 and 9 (MMP2 and MMP9) during the acquisition of the osteogenic phenotype, which precedes mineralization. RESULTS: Our data revealed that mechanosignaling-related angiocrine factors play a crucial role in promoting an osteoblastic phenotype in response to zirconia exposure. Specifically, exposed osteoblasts exhibited significantly higher expression levels of genes associated with osteoblast differentiation, such as Runx2, Osterix, bone sialoprotein, and osteocalcin genes. Furthermore, the activities of MMP2 and MMP9, which are involved in ECM remodeling, were modulated by mechanosignaling-related angiocrine factors. This modulation is likely an initial event preceding the mineralization phase. CONCLUSION: Based on our findings, we propose that mechanosignaling drives the release of angiocrine factors capable of modulating the osteogenic phenotype at the biointerface with zirconia. This process creates a microenvironment that promotes wound healing and osseointegration. Moreover, these results highlight the importance of considering the mechanosignaling of endothelial cells in the modulation of bone healing and osseointegration in the context of blood vessel effects. Our data provide new insights and open avenues for further investigation into the influence of mechanosignaling on bone healing and the osseointegration of dental devices.

The Action of Angiocrine Molecules Sourced from Mechanotransduction-Related Endothelial Cell Partially Explain the Successful of Titanium in Osseointegration.

Since Branemark's findings, titanium-based alloys have been widely used in implantology. However, their success in dental implants is not known when considering the heterogenicity of housing cells surrounding the peri-implant microenvironment. Additionally, they are expected to recapitulate the physiological coupling between endothelial cells and osteoblasts during appositional bone growth during osseointegration. To investigate whether this crosstalk was happening in this context, we considered the mechanotransduction-related endothelial cell signaling underlying laminar shear stress (up to 3 days), and this angiocrine factor-enriched medium was harvested further to use exposing pre-osteoblasts (pOb) for up to 7 days in vitro. Two titanium surfaces were considered, as follows: double acid etching treatment (w_DAE) and machined surfaces (wo_DAE). These surfaces were used to conditionate the cell culture medium as recommended by ISO10993-5:2016, and this titanium-enriched medium was later used to expose ECs. First, our data showed that there is a difference between the surfaces in releasing Ti molecules to the medium, providing very dynamic surfaces, where the w_DAE was around 25% higher (4 ng/mL) in comparison to the wo_DAE (3 ng/mL). Importantly, the ECs took up some of this titanium content for up to 3 days in culture. However, when this conditioned medium was used to expose pOb for up to 7 days, considering the angiocrine factors released from ECs, the concentration of Ti was lesser than previously reported, reaching around 1 ng/mL and 2 ng/mL, respectively. Thereafter, pOb exposed to this angiocrine factor-enriched medium presented a significant difference when considering the mechanosignaling subjected to the ECs. Shear-stressed ECs showed adequate crosstalk with osteoblasts, stimulating the higher expression of the Runx2 gene and driving higher expressions of Alkaline phosphatase (ALP), bone sialoprotein (BSP), and osteocalcin. Mechanotransduction-related endothelial cell signaling as a source of angiocrine molecules also stimulated the higher expression of the Col3A1 gene in osteoblasts, which suggests it is a relevant protagonist during trabecular bone growth. In fact, we investigated ECM remodeling by first evaluating the expression of genes related to it, and our data showed a higher expression of matrix metalloproteinase (MMP) 2 and MMP9 in response to mechanosignaling-based angiocrine molecules, independent of considering w_DAE or the wo_DAE, and this profile reflected on the MMP2 and MMP9 activities evaluated via gelatin-based zymography. Complimentarily, the ECM remodeling seemed to be a very regulated mechanism in mature osteoblasts during the mineralization process once both TIMP metallopeptidase inhibitor 1 and 2 (TIMP1 and TIMP2, respectively) genes were significantly higher in response to mechanotransduction-related endothelial cell signaling as a source of angiocrine molecules. Altogether, our data show the relevance of mechanosignaling in favoring ECs' release of bioactive factors peri-implant, which is responsible for creating an osteogenic microenvironment able to drive osteoblast differentiation and modulate ECM remodeling. Taking this into account, it seems that mechanotransduction-based angiocrine molecules explain the successful use of titanium during osseointegration.

The Impact of Bioactive Surfaces in the Early Stages of Osseointegration: An In Vitro Comparative Study Evaluating the HAnano® and SLActive® Super Hydrophilic Surfaces.

There is an increased effort on developing novel and active surfaces in order to accelerate their osteointegration, such as nanosized crystalline hydroxyapatite coating (HAnano®). To better understand the biological behavior of osteoblasts grown on HAnano® surface, the set of data was compared with SLActive®, a hydrophilic sandblasted titanium surface. Methodologically, osteoblasts were seeded on both surfaces up to 72 hours, to allow evaluating cell adhesion, viability, and set of genes encoding proteins related with adhesion, proliferation, and differentiation. Our data shows HAnano® displays an interesting substrate to support cell adhesion with typical spread morphologic cells, while SLActive®-adhering cells presented fusiform morphology. Our data shows that the cellular adhesion mechanism was accompanied with upexpression of integrin ß1, Fak, and Src, favoring the assembling of focal adhesion platforms and coupling cell cycle progression (upmodulating of Cdk2, Cdk4, and Cdk6 genes) in response to HAnano®. Additionally, both bioactive surfaces promoted osteoblast differentiation stimulus, by activating Runx2, Osterix, and Alp genes. Although both surfaces promoted Rankl gene expression, Opg gene expression was higher in SLActive® and this difference reflected on the Rankl/Opg ratio. Finally, Caspase1 gene was significantly upmodulated in response to HAnano® and it suggests an involvement of the inflammasome complex. Collectively, this study provides enough evidences to support that the nanohydroxyapatite-coated surface provides the necessary microenvironment to drive osteoblast performance on dental implants and these stages of osteogenesis are expected during the early stages of osseointegration.

Utilização de RHBMP-2 em reconstrução de maxila: relato de caso clínico / Use of rhBMP-2 in jaw reconstruction: Case report

Os enxertos ósseos utilizados em reconstruções são chamados de autógenos quando são retirados da própria pessoa que recebe o implante. Os enxertos autógenos ainda são utilizados pela maioria dos cirurgiões na Odontologia, mas sabe-se que essa opção apresenta algumas desvantagens, como necessidade de segunda área cirúrgica para coletar o osso, maior tempo cirúrgico, maior incidência de complicações e quantidade limitada de tecido. Devido a essas restrições, alternativas têm sido propostas e uma delas é o uso da proteina óssea morfogenética. Esta é capaz de induzir células tronco do local que foi aplicada, a diferenciarem-se em osteoblastos, promovendo assim a formação de tecido ósseo. Este trabalho demonstrou um caso clinico de reconstrucção da região anterior de uma maxilla extremamente atrófica com uso apenas de BMP e tela de titânio como arcabouço. Clínica, histológica e radiograficamente foi demonstrado um grande crescimento ósseo, possibilitando a instalação de implantes dentários.(AU)

Bone grafts used in reconstructions can be taken from the same person, when called autogenous. Autografts are still used by most surgeons in dentistry, but it is known that this option has some disadvantages, such as requiring a second surgical area to collect the bone, surgery time, higher incidence of complications and limited amount of tissue. Because of these constraints, alternatives have been proposed, one of which is the use of bone morphogenetic protein. This protein is capable of inducing stem cells has been applied, to differentiate in osteoblasts thereby promoting the formation of bone tissue. This report demonstrated a clinical case of reconstruction of extremely atrophic maxilla using only the BMP and titanium mesh scaffold. Clinically, histological and radiographically demonstrated a great bone growth, enabling the installation of dental implants. (AU)

Alterações das membranas paranasais de ratos submetidos à ação da fumaça do cigarro / Changes in the paranasal mucous membranes of rats exposed to cigarette smoke

Objetivo: reportar a relação do fumo com a perfuração da mucosa paranasal de ratos submetidos à ação intermitente da fumaça de cigarro. Material e métodos: foram utilizados nesta pesquisa 32 ratos machos e adultos Wistar (Rattus norvegicus, peso médio 400 g), divididos aleatoriamente em dois grupos (n=16), sendo: controle (não expostos à fumaça) e experimental (expostos). O grupo experimental era exposto à fumaça produzida por dez cigarros durante dez minutos, duas vezes ao dia. O nível de carboxihemoglobina (HbCO) no sangue arterial de cada rato foi coletado e analisado no espectrofotômetro. Depois, as mucosas sinusais foram retiradas cirurgicamente, embutidas em parafina, seccionadas no sentido anteroposterior, coradas e analisadas sob microscopia óptica nos períodos de 45 e 60 dias. O teste t pareado de Student foi usado para comparar os níveis de HbCO, e o teste exato de Fischer para possíveis correlações entre os achados histológicos e a exposição à fumaça (nível de significância 5%). Resultados: os níveis de HbCO no sangue dos ratos expostos foi significativamente maior (faixa de 1% - 20%, p < 0,0001) logo aos 45 dias. Os resultados histológicos demonstraram alterações significativas nas células caliciformes (60 dias, p=0,01) e na integridade ciliar (60 dias, p=0,001). Entretanto, metaplasias foram significativamente expressas nos períodos de 45 e 60 dias (p=0,03). Conclusão: nos períodos de 45 e 60 dias, alterações histológicas de grande relevância foram vistas nas mucosas sinusais expostas à fumaça do cigarro.