Spatially controlled adhesion, spreading, and differentiation of endothelial cells on self-assembled molecular monolayers.

Abstract

Chenaically moid glass substrates were used to denste dirental adheso, growth, and differention f endoel cells. Endothellal cells were eamine for Madei and growth on glass, glass treated with N-(2-aminethyl)-3-amnopropyl trimethoxysilane (EDA) Several approaches have also demonstrated the potential for controlling the spatial distribution of cells on the surface. Carter (8, 9) and later Harris (10), using acetate-coated glass, created palladium islands and channels for studying single fibroblast cell reactions and clone formation. Kleinfeld and coworkers (11) fabricated high-resolution patterns of selfassembled monolayers (SAMs) of amino-and perfluoroalkylsilanes, which resulted in dissociated neurons that preferentially adhered to and developed electrically active associations within the geometrically defined regions. The use of SAMs to create regions of defined chemical reactivity also allows selective attachment of fibroblasts and endothelial cells to regions modified by covalently bound peptides that contain the Arg-Gly-Asp (RGD) binding domain (12)(13)(14).In this study, a series of SAM-modified surfaces were examined for their ability to support adherence and facilitate growth and differentiation of endothelial cells. Patterned substrates, created by deep UV photolithography (15), were used to provide spatially defined regions that promoted adhesion, spreading ofendothelial cells, and ultimately spatially defined neovascular assemblies on a two-dimensional surface. MATERIALS AND METHODSReagents. Human fibronectin (hFN) was obtained from Collaborative Research. Heparin sulfate (HS) (47.5 kDa) was obtained from Sigma. N-(2-Aminoethyl)-3-aminopropyl trimethoxysilane (EDA) and (tridecafluoro-1,1,2,2-tetrahydrooctyl)-l-dimethylchlorosilane (13F) were used as procured from Hflls America (Bristol, PA).Substrate Preparation. Patterned deposition of EDA and 13F was accomplished by using the photolithography method of Dulcey et al. (15). Glass substrates were washed in methanol/HCl, 1:1 (vol/vol) for 1 hr and 18 M H2SO4 for 1 hr followed by multiple washes with deionized water (18 Mfl). After the final wash, the substrates were immersed in boiling deionized water for 20 min, removed, and immersed in 1% EDA/94% (vol/vol) methanol/i mM acetic acid/5% H20 for 15 min. The substrates were then rinsed at least three times in anhydrous methanol and baked at 1200C for 5 min.13F was deposited on either clean glass or patterned EDA surfaces by using a 1% (vol/vol) solution of the silane in anhydrous toluene for 1-3 hr, followed by at least four rinses in toluene, and baked 5 min at 120'C. EDA films were exposed to pulsed ArF excimer laser (193 nm) radiation (15 J/cm2, 30 Hz) through a lithographic mask. Deep UVirradiated regions of the substrate were resilanized with 13FAbbreviations: SAM, self-assembled monolayer; HUVE, human umbilical vein endothelial cell; EDA, N-(2-aminoethyl)-3-aminopropyl trimethoxysilane; 13F, (tridecafluoro-1,1,2,2-tetrahydrooctyl)-1-dimethylchlorosilane; bFGF, basic fibroblast growth factor; ...

Two Cell Circuits of Oriented Adult Hippocampal Neurons on Self-Assembled Monolayers for Use in the Study of Neuronal Communication in a Defined System

“…32 Its role in patterning applications as the cell-permissive substrate that is surrounded by noncell permissive SAMs is further strengthened because it is not degraded by the cells. 31,42 This characteristic of DETA allows it to form sharp patterns that do not blend with the noncell permissive PEG background and promotes pattern stability under long-term culture conditions. 25,31 PEG SAMs prevent the adsorption of proteins on glass surfaces by the entropy/hydrated surface hypothesis.…”

The synergistic effects of multivalent ligand display and nanotopography on osteogenic differentiation of rat bone marrow stem cells

“…Many reports have confirmed the effect of soluble factors and surface chemistry on the initial adhesion and proliferation of cells [18,19]. In nature, multivalent interactions are involved in cellecell communications and celleECM interactions.…”

“…Ligand clustering changes the receptor distribution in a dynamic way and regulates multipoint and strong attachment [20]. Depending on the ligand arrangement on the surface, it has been demonstrated that the surface chemistry not only plays a significant role in initial adhesion but may also control the signaling processes leading to different differentiation pathways [19]. The ligand arrangement and density along with the physical characteristics of the underlying substrate can modulate cell behavior independently as well as collectively.…”

Biology on a Chip: Microfabrication for Studying the Behavior of Cultured Cells

“…235 A range of techniques have been used to micropattern endothelial cells. [236][237][238][239][240][241][242][243] Chen et al 244 found that cell spreading acted as a regulator of proliferation or death in human and bovine capillary endothelial (CE) cells by tightly controlling cell spreading with adhesive microislands of fibronectin; fibronectin was adsorbed on a methyl-terminated alkanethiol SAM, whereas the nonadhesive background consisted of a hexaethylene-glycolterminated alkanethiol SAM (HEG-SAM) that resisted protein adsorption and cell attachment. Cells that were free to spread over large microislands displayed growth, as expected, but the cells that were constrained to limit their spreading to a small island displayed apoptosis.…”