Alterations in gene expression of human vascular endothelial cells associated with nanotopographic cues

Abstract

Human cells in vivo are exposed to a topographically rich, 3-dimenisional environment which provides extracellular cues initiating a cascade of biochemical signals resulting in changes in cell behavior. One primary focus of our group is the development of biomimetic substrates with anisotropic nanoscale topography to elucidate the mechanisms by which physical surface cues are translated into biochemical signals. To investigate changes in gene expression as a result of nanotopographic cues, Human Umbilical Vein Endothelial Cells (HUVECs) were cultured on chemically identical flat and 400 nm pitch nanogrooved surfaces. After 12 hours, RNA was harvested for an Affymetrix HG U133 Plus 2.0 gene array. Of over 47,000 possible gene probes, 3171 had at least a two-fold difference in expression between the control flat and 400 nm pitch. The gene ontology groups with the most significant increase in expression are involved in protein modification and maintenance, similar to cells upregulating chaperone and protein synthesis genes in response to physical stresses. The most significant decreases in expression were observed with cell cycle proteins, including cyclins and checkpoint proteins. Extracellular matrix proteins, including integrins, collagens, and laminins, are almost uniformly down regulated on the 400 nm pitch surfaces compared to control. The down regulation of one of these genes, integrin beta 1, was confirmed via quantitative PCR. Together, these gene array data, in addition to our studies of cell behavior on nanoscale surfaces, contribute to our understanding of the signaling pathways modulated by topographical surface cues.

Comparing the endothelialisation of extracellular matrix bioscaffolds with coated synthetic vascular graft materials

“…This has lead to nanoscale engineering to influence cellular orientation, growth, attachment and migration [32,33] In vitro endothelialisation not only requires adequate cellular attachment, but also rapid proliferation. Slow proliferation increases incubation times leading to higher rates of contamination and production costs.…”

Early responses of vascular endothelial cells to topographic cues

“…A previously performed gene chip analysis showed no differences in FAK expression between HUVECs cultured on the 400-nm pitch and those cultured on planar surfaces (14). To analyze additional pitch sizes, we performed a qPCR using RNA isolated from HUVECs cultured on topographically patterned surfaces and planar control substrates.…”

Influence of Extracellular Matrix Proteins and Substratum Topography on Corneal Epithelial Cell Alignment and Migration

“…55 An earlier study from our group using human umbilical vein endothelial cells demonstrated that over 3000 genes, many associated with the ECM and cell adhesion, cytoskeletal reorganization, DNA replication and repair, and spindle organization, were significantly altered by biomimetic size scale (400-nm pitch) topographic features. 56 It is known that while nanoscale topographic cues can modulate the expression of integrins in cells, 57 there is bidirectional signaling between the cytoskeleton and integrins. 58 Additionally, the mechanical link between integrins and various cytoskeletal Tracks of fibronectin (yellow trace lines) were followed for cells migrating on FNCcoated topographies.…”