Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2003 May 27;100(11):6331-6.
doi: 10.1073/pnas.1031426100. Epub 2003 May 13.

Interaction of soft condensed materials with living cells: phenotype/transcriptome correlations for the hydrophobic effect

Lorcan T Allen  1 , Edward J P Fox, Irena Blute, Zoe D Kelly, Yuri Rochev, Alan K Keenan, Kenneth A Dawson, William M Gallagher
Affiliations

Interaction of soft condensed materials with living cells: phenotype/transcriptome correlations for the hydrophobic effect

Lorcan T Allen et al. Proc Natl Acad Sci U S A. .

Abstract

The assessment of biomaterial compatibility relies heavily on the analysis of macroscopic cellular responses to material interaction. However, new technologies have become available that permit a more profound understanding of the molecular basis of cell-biomaterial interaction. Here, both conventional phenotypic and contemporary transcriptomic (DNA microarray-based) analysis techniques were combined to examine the interaction of cells with a homologous series of copolymer films that subtly vary in terms of surface hydrophobicity. More specifically, we used differing combinations of N-isopropylacrylamide, which is presently used as an adaptive cell culture substrate, and the more hydrophobic, yet structurally similar, monomer N-tert-butylacrylamide. We show here that even discrete modifications with respect to the physiochemistry of soft amorphous materials can lead to significant impacts on the phenotype of interacting cells. Furthermore, we have elucidated putative links between phenotypic responses to cell-biomaterial interaction and global gene expression profile alterations. This case study indicates that high-throughput analysis of gene expression not only can greatly refine our knowledge of cell-biomaterial interaction, but also can yield novel biomarkers for potential use in biocompatibility assessment.

PubMed Disclaimer

Figures

Fig. 1.
Fig. 1.
Structure of NIPAAm and NtBAAm monomers. Variations in the ratio of NtBAAm to NIPAAm make it possible to modulate the adhesivity of the copolymer surface to living cells by altering surface hydrophobicity, as previously confirmed by microcalorimetry studies (5).
Fig. 2.
Fig. 2.
Cell adhesion on TCP or various NIPAAm:NtBAAm copolymer films. Adhesion was quantified by determining cell numbers on the different surfaces 24 h postseeding. (A) Adhesion of two transformed cell lines, HeLa (epithelial) and MCR-5 (fibroblast). (B) Adhesion of two primary cell types, 1BR3 (fibroblast) and HASMC (aortic smooth muscle). Error bars indicate SEM (n = 3). *, P < 0.05; **, P < 0.01; ***, P < 0.005, respectively, as evaluated by unpaired Student's t test with respect to difference between adhesion determined on the various copolymer films in comparison with TCP. •, ••, and ••• indicate significant differences (P < 0.05, P < 0.01, or P < 0.005, respectively; unpaired Student's t test) between individual copolymer films.
Fig. 3.
Fig. 3.
Cell growth on TCP or various NIPAAm:NtBAAm copolymer films. The extent of growth was quantified by determining cell numbers on the different surfaces 96 h postseeding. (A) Adhesion of two transformed cell lines, HeLa (epithelial) and MCR-5 (fibroblast). (B) Adhesion of two primary cell types, 1BR3 (fibroblast) and HASMC (aortic smooth muscle). Error bars indicate SEM (n = 3). *, P < 0.05; **, P < 0.01; ***, P < 0.005, respectively, as evaluated by unpaired Student's t test with respect to difference between the extent of growth determined on the various copolymer films in comparison with TCP. •, ••, and ••• indicate significant differences (P < 0.05, P < 0.01, or P < 0.005, respectively; unpaired Student's t test) between individual copolymer films.
Fig. 4.
Fig. 4.
Inhibition of cell spreading on NIPAAm:NtBAAm copolymer films. HeLa cells were allowed to adhere and spread on either copolymer film or TCP for 24 h and then fixed with 4% paraformaldehyde. Fixed cells were visualized by light microscopy. (A) The average spread area is expressed relative to the degree of cell spreading observed on TCP. Error bars indicate SEM (with numbers of cells analyzed shown under each bar). *, P < 0.05; **, P < 0.01; ***, P < 0.005, respectively, as evaluated by unpaired Student's t test with respect to difference between cell spreading determined on the various copolymer films in comparison with TCP. • and ••• indicate significant differences (P < 0.05 or P < 0.005, respectively; unpaired Student's t test) between individual copolymer films. (B) Representative images of adherent cells seeded onto either copolymer film or TCP. Cells grown on 85:15 copolymer film for >24 h displayed a stellate morphology similar to that observed with the other copolymer films at 24 h. (C) Modulation of cytoskeletal structure. HeLa cells were allowed to spread on either copolymer film-coated or uncoated Lab-TekII chamber slides for 24 h and then fixed with 4% paraformaldehyde. Fixed cells were stained with rhodamine-phalloidin to detect F-actin. (Scale bar = 50 μm.)
Fig. 5.
Fig. 5.
Differential gene expression in HeLa cells in response to interaction with copolymer films. HeLa cells were seeded onto either TCP or the various copolymer films and left for 24 h before DNA microarray-based gene expression profile analysis. (A) Venn diagram illustrating results from comparative analysis of different gene expression profiles. Each circle represents a copolymer film/TCP comparison. Figures represent the number of human transcripts differentially expressed between particular comparisons. (B) A graphical illustration of fold change values associated with the central subset of 21 differentially expressed transcripts. Transcript names were derived from GeneCards (http://bioinformatics.weizmann.ac.il/cards). As an illustration of experimental reproducibility, the altered expression levels of c-fos and FOLR1 were verified by two independent probe sets (probe sets 1 and 2). Differential gene expression of a subset of these transcripts was further confirmed by quantitative RT-PCR amplification (data not shown).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Rihova, B. (1996) Adv. Drug Delivery Rev. 21, 157–176.
    1. Dawson, K. A. (2002) Curr. Opin. Colloid Interface Sci. 7, 218–227.
    1. Tang, L. & Eaton, J. W. (1995) Am. J. Clin. Pathol. 103, 466–471. - PubMed
    1. Ito, Y. (1999) Biomaterials 20, 2333–2342. - PubMed
    1. Rochev, Y., Golubeva, T., Gorelov, A., Allen, L. T., Gallagher, W. M., Selezneva, I., Gavrilyuk, B. & Dawson, K. (2001) Prog. Colloid Polym. Sci. 118, 153–156.

Publication types

LinkOut - more resources