Fabrication of Micro Patterned Fibronectin for Studying Adhesion and Alignment Behavior of Human Dermal Fibroblasts

摘要

The aim of this study was to fabricate a submicro-and micro-patterned fibronectin coated wafer for a cell culture,which allows the positions and dimensions of the attached cells to be controlled.A replica molding was made into silicon via a photomask in quartz,using E-beam lithography,and then fabricated a polydimethylsiloxane stamp using the designed silicon mold.Hexadecanethiol [HS(CH2)15CH3],adsorbed on the raised plateau of the surface of polydimethylsiloxane stamp,was contact-printed to form self-assembled monolayers (SAMs) of hexade-canethiolate on the surface of an Au-coated glass wafer.In order to form another SAM for control of the surface wafer properties,a hydrophilic hexa (ethylene glycol) terminated alkanethiol [HS(CH2)_(11)(OCH2CH2)6OH] was also synthesized.The structural changes were confirmed using UV and ~1H-NMR spectroscopies.A SAM terminated in the hexa(ethylene glycol) groups was subsequently formed on the bare gold remaining on the surface of the Au-coated glass wafer.In order to aid the attachment of cells,fibronectin was adsorbed onto the resulting wafer,with the pattern formed on the gold-coated wafer confirmed using immunofluorescence staining against fibronectin.Fibronectin was adsorbed only onto the SAMs terminated in the methyl groups of the substrate.The hexa (ethylene glycol)-terminated regions resisted the adsorption of protein.Human dermal fibroblasts (P=4),obtained from newborn foreskin,only attached to the fibronectin-coated,methyl-terminated hydrophobic regions of the patterned SAMs.N-HDFs were more actively adhered,and spread in a pattern spacing below 14 mu m,rather than above 17 mu m,could easily migrate on the substrate containing spacing of 10 /an or less between the strip lines.