Engineering Cell Adhesion and Orientation via Ultrafast Laser Fabricated Microstructured Substrates

Abstract

Cells take decisions on their responses depending on the stimuli received by the surrounding extracellular environment, that provides the essential cues at the micro and the nano-lengthscales required for adhesion, orientation, proliferation and differentiation. In this study, discontinuous microcones on silicon (Si) and continuous microgrooves on polyethylene terephthalate (PET) substrates were fabricated via ultrashort-pulsed laser irradiation at various fluences, resulting in microstructures with different roughness and geometrical characteristics. The topographical models attained were specifically developed to imitate the guidance and alignment of Schwann cells for oriented axonal regrowth, towards nerve regeneration. At the same time, positive replicas of the silicon microstructures formed were successfully reproduced, via soft lithography, on the biodegradable polymer poly(lactide-co-glycolide) (PLGA). The anisotropic continuous (PET) and discontinuous (PLGA replicas) microstructured polymeric substrates were assessed in terms of their influence on the Schwann cells responses. It is shown that the developed micropatterned substrates enable control over the cellular adhesion, proliferation and orientation and are thus useful to engineer cell alignment in vitro. This property could be potentially useful in the fields of neural tissue engineering and for dynamic microenvironment systems that simulate in vivo conditions.

Combined Effect of Shear Stress and Laser-Patterned Topography on Schwann cell Outgrowth: Synergistic or Antagonistic?

“…In the present study, polymeric continuous MG (PET-MG substrates) have been prepared by ultrafast laser direct writing ( Figure 2) [18]. The width of the MG was chosen to be in the range of 2 to 30 μm, which is considered to be optimal for the alignment of Schwann cells [4].…”

“…For the dynamic cultures, cells were seeded on the planar PET coverslips or the laser-microstructured substrates, placed inside the chamber and kept at rest overnight in a 5% CO 2 incubator at 33 o C. The next day, the continuous perfusion was initiated and lasted for 2 days. Based on the results of our previous work [18], under static conditions, we determined that it would be more effective to quantify the cell length after a 3-day culture period. Moreover, parametric studies were performed both under static and dynamic conditions before selecting these optimal parameters (dimensions of the substrate, topography of the substrate, cell densities, flow rates) for investigating the effect of shear stress (section A in the Supplementary Material).…”

“…The laser-structured microgrooved substrates (PET-MG) were morphologically characterized by scanning electron microscopy (SEM). The corresponding top-view and cross-section SEM images are presented in Figure 2, showing that the width, w, of the MG was 28.68 ± 0.47 μm, the depth, d, 8.87 ± 0.44 μm, the aspect ratio, and the roughness ratio, [18].…”

“…Therefore, many groups have focused their interest on the controlled modification of materials' surfaces, as a strategy for guiding the neurite outgrowth, which is crucial for the development of functional neuronal interfaces [10][11][12][13]. Among various techniques employed, ultrashort-pulse laser structuring has proved to be important for engineering surface topography in various materials [13][14][15][16][17][18][19][20][21][22] including silicon (Si), PET, and poly(lactide-co-glycolide) (PLGA). Indeed, the laser-induced topography was found to significantly affect the neuronal adhesion, growth, and orientation.…”

Combined effect of shear stress and laser-patterned topography on Schwann cell outgrowth: synergistic or antagonistic?

“…Therefore, many groups have focused their interest on the controlled modification of materials' surfaces, as a strategy for guiding the neurite outgrowth, which is crucial for the development of functional neuronal interfaces [10][11][12][13] . Among various techniques employed, ultrashort-pulse laser structuring has proved to be important for engineering surface topography in various materials [13][14][15][16][17][18][19][20][21][22] including silicon (Si), PET, and poly(lactide-co-glycolide) (PLGA). Indeed, the laser-induced topography was found to significantly affect the neuronal adhesion, growth, and orientation.…”