A nanoporous surface is essential for glomerular podocyte differentiation in three-dimensional culture.

Although it is well recognized that cell-matrix interactions are based on both molecular and geometrical characteristics, the relationship between specific cell types and the three-dimensional morphology of the surface to which they are attached is poorly understood. This is particularly true for glomerular podocytes - the gatekeepers of glomerular filtration - which completely enwrap the glomerular basement membrane with their primary and secondary ramifications. Nanotechnologies produce biocompatible materials which offer the possibility to build substrates which differ only by topology in order to mimic the spatial organization of diverse basement membranes. With this in mind, we produced and utilized rough and porous surfaces obtained from silicon to analyze the behavior of two diverse ramified cells: glomerular podocytes and a neuronal cell line used as a control. Proper differentiation and development of ramifications of both cell types was largely influenced by topographical characteristics. Confirming previous data, the neuronal cell line acquired features of maturation on rough nanosurfaces. In contrast, podocytes developed and matured preferentially on nanoporous surfaces provided with grooves, as shown by the organization of the actin cytoskeleton stress fibers and the proper development of vinculin-positive focal adhesions. On the basis of these findings, we suggest that in vitro studies regarding podocyte attachment to the glomerular basement membrane should take into account the geometrical properties of the surface on which the tests are conducted because physiological cellular activity depends on the three-dimensional microenvironment

Effects of substrates nanopattering on osteosarcoma cell behaviour.

Engineering of the cellular microenvironment has become an attractive strategy to guide cellular activities such as spreading, motility, proliferation and differentiation. From a technological perspective, the physical crosstalk between the cell and its surroundings represents a design parameter that may be modulated to achieve desired physiological outcome. In this study we present a surface engineering approach to tap into the physical crosstalk between the cell and its surroundings in order to modulate osteogenic anchorage-dependent differentiation and bone formation. The effectiveness of this approach was studied by comparing the cellular behavior of human SOAS sarcoma cells on nanostructured silicon substrates with distinct nanoscale patterns. Random nano-islands were realized by controlled deposition of tin on the polished side of silicon wafers by thermal evaporation. Four different shaped surfaces of nano structured substrates were used in this study. Silicon substrates present surface islands with diameters ranging from 10 to 35 nm and inter-island distances of 41 (B), 51 (E) or 80 (F) nm respectively. Substrate A is planar silicon used as contro

A nanoporous surface is essential for glomerular podocyte differentiation in three-dimensional culture

Cristina Zennaro,1 Maria Pia Rastaldi,2 Gerald James Bakeine,3 Riccarda Delfino,1 Federica Tonon,1 Rossella Farra,4 Gabriele Grassi,1,5 Mary Artero,6 Massimo Tormen,7 Michele Carraro1 1Department of Medical, Surgical and Health Sciences, University of Trieste, Trieste, 2Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, 3Department of Radiology, San Martino University Hospital, University of Genoa, Genoa, 4Department of Engineering and Architecture, University of Trieste, 5Department of Life Sciences, Cattinara University Hospital, University of Trieste, 6Azienda Sanitaria Universitaria Integrata di Trieste, Trieste, 7IOM-CNR Area Science Park, Trieste, Italy Abstract: Although it is well recognized that cell–matrix interactions are based on both molecular and geometrical characteristics, the relationship between specific cell types and the three-dimensional morphology of the surface to which they are attached is poorly understood. This is particularly true for glomerular podocytes – the gatekeepers of glomerular filtration – which completely enwrap the glomerular basement membrane with their primary and secondary ramifications. Nanotechnologies produce biocompatible materials which offer the possibility to build substrates which differ only by topology in order to mimic the spatial organization of diverse basement membranes. With this in mind, we produced and utilized rough and porous surfaces obtained from silicon to analyze the behavior of two diverse ramified cells: glomerular podocytes and a neuronal cell line used as a control. Proper differentiation and development of ramifications of both cell types was largely influenced by topographical characteristics. Confirming previous data, the neuronal cell line acquired features of maturation on rough nanosurfaces. In contrast, podocytes developed and matured preferentially on nanoporous surfaces provided with grooves, as shown by the organization of the actin cytoskeleton stress fibers and the proper development of vinculin-positive focal adhesions. On the basis of these findings, we suggest that in vitro studies regarding podocyte attachment to the glomerular basement membrane should take into account the geometrical properties of the surface on which the tests are conducted because physiological cellular activity depends on the three-dimensional microenvironment. Keywords: podocytes, neurons, nanotechnology, basement membrane, differentiation, adhesio

Bottom-up approach to construct microfabricated multi-layer scaffolds for bone tissue engineering

The use of bottom-up approaches in tissue engineering applications is advantageous since they enable the combination of various layers that could be made from different materials and/or incorporate different biochemical cues. Regarding the complex structure and the vascular system of the bone tissue, the aim of this study was to develop an innovative bottom-up approach that allows the construction of 3D biodegradable scaffolds from 2D microfabricated membranes with precise shape, pore size and porosity. For that purpose, poly (caprolactone) (PCL) and starch – poly (caprolactone) (SPCL (30% starch)) blended sheets were used as substrates to produce the microfabricated membranes using micro hot-embossing. The use of this micro fabrication process allowed accurately imprinting micropillars and microholes in reproducible way. The assembling of the microfabricated membranes was performed using an easy, highly reproducible and inexpensive approach based on its successive stacking. Additionaly, the suitability of the microfabricated membranes to support the attachment and the cytoskeletal organization of human bone marrow stem cells (hBMSCs), macrovascular endothelial cells and osteoblasts derived from hBMSCs was demonstrated. Furthermore, hBMSCs proliferated and maintained the expression of the stromal progenitor marker STRO-1 when cultured on both PCL and SPCL microfabricated membranes. The proposed methodology constitutes a promising alternative to the traditional processing methods used to prepare tissue engineering scaffolds.This study was financed by Portuguese Foundation for Science and Technology (FCT) through the project MicroNanoScaff (PTDC/CTM/108209/2008)