Nano hydroxyapatite-blasted titanium surface creates a biointerface able to govern Src-dependent osteoblast metabolism as prerequisite to ECM remodeling

Abstract

Over the last several years, we have focused on the importance of intracellular signaling pathways in dynamically governing the biointerface between pre-osteoblast and surface of biomaterial. Thus, this study investigates the molecular hallmarks involved in the pre-osteoblast relationship with different topography considering Machined (Mc), Dual Acid-Etching (DAE), and nano hydroxyapatite-blasted (nHA) groups. There was substantial differences in topography of titanium surface, considering Atomic Force Microscopy and water contact angle (Mc = 81.41 ± 0.01; DAE = 97.18 ± 0.01; nHA = 40.95 ± 0.02). Later, to investigate their topography differences on biological responses, pre-osteoblast was seeded on the different surfaces and biological samples were collected after 24 h (to consider adhesion signaling) and 10 days (to consider differentiation signaling). Preliminary results evidenced significant differences in morphological changes of pre-osteoblasts mainly resulting from the interaction with the DAE and nHA, distinguishing cellular adaptation. These results pushed us to analyze activation of specific genes by exploring qPCR technology. In sequence, we showed that Src performs crucial roles during cell adhesion and later differentiation of the pre-osteoblast in relationship with titanium-based biomaterials, as our results confirmed strong feedback of the Src activity on the integrin-based pathway, because integrin-ß1 (∼5-fold changes), FAK (∼12-fold changes), and Src (∼3.5-fold changes) were significantly up-expressed when Src was chemically inhibited by PP1 (5 μM). Moreover, ECM-related genes were rigorously reprogrammed in response to the different surfaces, resulting on Matrix Metalloproteinase (MMP) activities concomitant to a significant decrease of MMP inhibitors. In parallel, we showed PP1-based Src inhibition promotes a significant increase of MMP activity. Taking all our results into account, we showed for the first time nano hydroxyapatite-blasted titanium surface creates a biointerface able to govern Src-dependent osteoblast metabolism as pre-requisite to ECM remodeling.

Differential inflammatory landscape stimulus during titanium surfaces obtained osteogenic phenotype

“…On this context, a draw of the molecular network of these findings was necessary, and bioinformatics hypothesizes a strong crosstalk between Shh signaling and IL10 being mediated by Src and mTor proteins. Unsurprisingly, we reported previously elsewhere the involvement of Src in the intracellular signaling in response to nano HA‐blasted titanium surfaces and this reinforces our previous hypothesis. This anti‐inflammatory signal might explain the low profile of RANKL expression by osteoblasts, guaranteeing consequently a low profile of osteoclastogenesis at this stage, as already detailed.…”

“…Although nanometric‐scaled structures have important properties considering osseointegration, detailed interfacial interactions with osteogenic cells have not been fully addressed. On this matter, we have previously mapped the cellular phenotype of osteoblast when in response to nano hydroxyapatite‐blasted surfaces (nHA), showing a decisive capacity of the nHA in stimulating osteoblast differentiation by requiring Src as prerequisite . However, in order to better comprehend the molecular mechanisms triggered by nHA, other aspects must be addressed such as inflammatory cytokines reprogramming.…”

Layered Double Hydroxides Are Promising Nanomaterials for Tissue Bioengineering Application

“…Overall, we have shown that cells actively respond to biomaterials modifying their morphology as an immediate consequence of a well‐orchestrated cytoskeleton rearrangement upon integrin activation, where the phosphorylation of cofilin drives actin filaments rearrangement and governs cell adaptation on the substrate. Additionally, on the adaptive process on the biomaterials, we have also shown that the cell growth requires intracellular cascades requiring MAP kinases (ERK, JNK, and P38) and ECM remodeling . In turn, ECM plays structural and functional roles, it being sufficient and necessary for the plasticity of the cells and their remodeling to be considered in regenerative mechanisms.…”

“…Among the biomaterials, those devoted to bioengineering protocols need to recapitulate the wound‐healing steps triggering a moderate inflammatory cascade activating hemostasis and later requiring platelets and fibrinogen. In this context, advances in physics and chemistry area allowed the application of nanoscaled structures on modifying and functionalizing molecules/materials, ameliorating their performance in medical proposes . It is not new the purpose of layered double hydroxides (LDHs) as biomaterial .…”

“…Regrettably, serious preclinical concerns remain, the signaling cross talk with cells and biomaterials proposes are still the challenge for this promising therapeutic modality . Thus, we have explored and suggested in vitro approaches to predict the quality of the cell/biomaterial interaction, suggesting the intracellular signaling as a preliminary and reliable tool on this matter . Overall, we have shown that cells actively respond to biomaterials modifying their morphology as an immediate consequence of a well‐orchestrated cytoskeleton rearrangement upon integrin activation, where the phosphorylation of cofilin drives actin filaments rearrangement and governs cell adaptation on the substrate.…”