title: Cross-linked bioreducible layer-by-layer films for increased cell adhesion and transgene expression.
abstract: The effect of cross-linking layer-by-layer (LbL) films consisting of bioreducible poly(2-dimethylaminoethyl methacrylate) (rPDMAEMA) and DNA is examined with regard to rigidity, biodegradability, cell adhesion, and transfection activity using 1,5-diiodopentane (DIP) cross-linker. DIP chemically reacts with the tertiary amines of rPDMAEMA, altering the chemical composition of these LbL films. The result is a change in surface morphology, film swelling behavior, and film rigidity, measured with AFM and ellipsometry. It is found that the apparent Young's modulus is increased more than 4 times its original value upon cross-linking. Cross-linking mass is additionally confirmed with a quartz crystal microbalance with dissipation (QCM-D). Comprehensive analyses of these experimental values were investigated to calculate the degree of cross-linking using the rubber elasticity theory and the Flory-Rehner theory. Additionally, the Flory-Huggins parameter, chi, was calculated. Good agreement in the two methods yields a cross-linking density of approximately 0.82 mmol/cm(3). The Flory-Huggins parameter increased upon cross-linking from 1.07 to 1.2, indicating increased hydrophobicity of the network and formation of bulk water droplets within the films. In addition, the effects of cross-linking on film disassembly by 1,4-dithiothreitol (DTT) are found to be insignificant despite the alteration in film rigidity. Mouse fibroblast cells and smooth muscle cells are used to study the effect of cross-linking on cell adhesion and cell transfection activity. In vitro transfection activity up to seven days is quantified using secreted alkaline phosphatase (SEAP) DNA. Film cross-linking is found to enhance cell adhesion and prolong the duration of cellular transfection. These results contribute to the development of bioreducible polymer coatings for localized gene delivery.
authors.affiliation: Department of Biomedical Engineering, Wayne State University, Detroit, Michigan 48202, USA. blacklock@mpikg.mpg.de
authors.forename: Jenifer
authors.initials: J
authors.lastname: Blacklock
authors.forename: Torsten K
authors.initials: TK
authors.lastname: Sievers
authors.forename: Hitesh
authors.initials: H
authors.lastname: Handa
authors.forename: Ye-Zi
authors.initials: YZ
authors.lastname: You
authors.forename: David
authors.initials: D
authors.lastname: Oupický
authors.forename: Guangzhao
authors.initials: G
authors.lastname: Mao
authors.forename: Helmuth
authors.initials: H
authors.lastname: Möhwald
mesh_terms: D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D004247:DNA / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D015870:Gene Expression / Q000187:drug effects*; D007700:Kinetics; D008689:Methacrylates / Q000737:chemistry* / Q000494:pharmacology* / Q000633:toxicity; D051379:Mice; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D041681:NIH 3T3 Cells; D009757:Nylons / Q000737:chemistry* / Q000494:pharmacology* / Q000633:toxicity; D010420:Pentanes / Q000737:chemistry; D014162:Transfection; D019076:Transgenes / Q000235:genetics*
chemical_list: D004220:Disulfides; D008689:Methacrylates; D009757:Nylons; D010420:Pentanes; C407037:poly(2-(dimethylamino)ethyl methacrylate); D004247:DNA