Adhesion formation of primary human osteoblasts and the functional response of mesenchymal stem cells to 330 nm deep microgrooves

Abstract

The surface microtexture of an orthopaedic device can regulate cellular adhesion, a process fundamental in the initiation of osteoinduction and osteogenesis. Advances in fabrication techniques have evolved to include the field of surface modification; in particular, nanotechnology has allowed for the development of experimental nanoscale substrates for investigation into cell nanofeature interactions. Here primary human osteoblasts (HOBs) were cultured on ordered nanoscale groove/ridge arrays fabricated by photolithography. Grooves were 330 nm deep and either 10, 25 or 100 mm in width. Adhesion subtypes in HOBs were quantified by immunofluorescent microscopy and cell-substrate interactions were investigated via immunocytochemistry with scanning electron microscopy. To further investigate the effects of these substrates on cellular function, 1.7 K gene microarray analysis was used to establish gene regulation profiles of mesenchymal stem cells cultured on these nanotopographies. Nanotopographies significantly affected the formation of focal complexes (FXs), focal adhesions (FAs) and supermature adhesions (SMAs). Planar control substrates induced widespread adhesion formation; 100 mm wide groove/ridge arrays did not significantly affect adhesion formation yet induced upregulation of genes involved in skeletal development and increased osteospecific function; 25 mm wide groove/ridge arrays were associated with a reduction in SMA and an increase in FX formation; and 10 mm wide groove/ridge arrays significantly reduced osteoblast adhesion and induced an interplay of up -and downregulation of gene expression. This study indicates that groove/ridge topographies are important modulators of both cellular adhesion and osteospecific function and, critically, that groove/ridge width is important in determining cellular response.

Skeletal stem cell and bone implant interactions are enhanced by LASER titanium modification

“…In contrast, Takeuchi et al showed that a modified surface can reduce cell proliferation whilst initially driving the expression of specific cell markers except for ALP 6 [27]. The cellbiomaterial interface functions not only to define the boundary between tissue and implant, but also to act as a mediator of first stage protein interactions as well as later stage cell adhesion and orientation [28]. When blood cells arrive at the implant Ti surface, the blood cells express a variety of integrins, resulting in cytoskeletal changes.…”

Substrate topography: A valuable in vitro tool, but a clinical red herring for in vivo tenogenesis

“…The groove dimensions were selected on the basis that closely represent the topographical cues (dimensionality of collagen fibrils is in the region of 10 to 1,000 nm, whilst the dimensionality of collagen fibres is in the region of 1,000 to 20,000 nm [12]) that tenocytes are exposed to in vivo. The micro-scale groove depth was selected based on previous publications, where authors supported that pitch dimensionality smaller than 4000 nm induces efficient cellular contact guidance [40,42,52].…”

“…electrospun polymeric fibres, extruded collagen fibres and isoelectrically focused collagen fibres) have been shown to maintain tenocyte phenotype and to differentiate stem cells towards tenogenic lineage in vitro and to induce acceptable regeneration in preclinical models, none of these technologies offers precise control over the spatial distribution of the fibres. Imprinting technologies, on the other hand, have demonstrated a diverse effect on a range of permanently differentiated and stem cell functions, including adhesion, orientation, secretome expression and lineage commitment [41][42][43][44][45][46][47][48] and offer significantly greater control over feature dimension and spacing. Specifically to tendon repair, such technologies have been shown to maintain tenocyte phenotype [38]; to promote aligned tendon-specific ECM deposition [39]; and to differentiate stem cells towards tenogenic lineage [40].…”

3D printed PLA-based scaffolds

“…Results revealed that although both materials displayed similar cell viability results, noteworthy differences were observed in on cell response. 26 Surface nanoporosity not only increases the contact surface area between the material and biological entities but also generates nanoscale topographical cues affecting cell behavior. In fact, enhancement of cell interaction due to nanotopography has been reported.…”