Adhesion response of filopodia to an AFM lateral detachment force and functional changes after centrifugation of cells grown on nanoporous titanium.

Cells sense and respond to mechanical cues from the surrounding substrate through filopodia. Regulation of cellular biomechanics operates at the nanoscale. Therefore, a better understanding of the relationship between filopodia and nanoscale surface features is highly relevant for the rational design of implant surfaces. The objective of this work was to determine the biomechanical contribution of filopodia and their nanoprotrusions to the adhesive interaction of cells with nanostructured surfaces. We have also analyzed the functional changes of entire cells subjected to an external force. MC3T3-E1 osteogenic cells were cultured on polished (Ti-Control) and nanotextured titanium discs (Ti-Nano). An AFM approach was used to measure the lateral detachment force of filopodia. Filopodia on Ti-Nano exhibited higher resistance to a lateral detachment force, which indicates that they adhere to the surface with more strength. SEM analysis revealed a restructuration of the cell membrane in response to centrifugation, being more evident on Ti-Nano. Fluorescence labeling also highlighted a difference in the mitochondrial footprint, a cellular compartment that provides energy for cellular processes. Together, these results show for the first time that surface topography can change the adhesive interaction of a subcellular structure that is fundamental in sensing physico-chemical surfaces features.

Effect of molecular weight on the exponential growth and morphology of hyaluronan/chitosan multilayers: a surface plasmon resonance spectroscopy and atomic force microscopy investigation.

The layer-by-layer growth of multilayer assemblies of two polysaccharides, the polyanion hyaluronan (HA) and the polycation chitosan (CH), was investigated using atomic force microscopy (AFM) and surface plasmon resonance (SPR) spectroscopy, with primary emphasis on the effect of the polysaccharide molecular weights on the film thickness and surface morphology. The HA/CH multilayers exhibit an exponential increase of the optical film thickness with the number of deposited bilayers. We show that the multilayer thickness at a given stage depends on the size of both CH, the diffusing polyelectrolyte, and HA, the non-diffusing species. Assemblies (12 bilayers) of high molecular weight polysaccharides (HA, 360,000; CH, 160,000) were twice as thick (approximately 900 nm vs approximately 450 nm) as those obtained with low molecular weight polymers (HA, 30,000; CH, 31,000), as assessed by AFM scratch tests. The exponential growth rate is the same for the high and low molecular weight pairs; the larger film thicknesses observed by SPR and by AFM arising from an earlier onset of the steep exponential growth phase in the case of the high molecular weight pair. In all cases, isolated islets form during the deposition of the first CH layer onto the underlying HA. Upon further film growth, individual islets coalesce into larger vermiculate features. The transition from distinct islands to vermiculate structures depends on the molecular weights of the polysaccharides and the lower molecular weight construct presents larger worm-like surface domains than the high molecular weight pair.

Enzymatic lithography of phospholipid bilayer films by stereoselective hydrolysis.

The stereoselective phospholipase A2-catalyzed hydrolysis of patterned phospholipid bilayers consisting of the l- and d-isomers of alpha-dilauroylphosphatidylcholine (DLPC) and alpha-dipalmitoylphosphatidylcholine (DPPC) is reported. The stereochemically directed enzyme lithography demonstrated herein allows the parallel modification of large surface areas and constitutes a potentially useful method to structure biomimetic films, given the stereospecific action of many enzymes.