The regulation of focal adhesion complex formation and salivary gland epithelial cell organization by nanofibrous PLGA scaffolds

Department of Biological Sciences, University at Albany, State University of New York, 1400 Washington Ave, LSRB 1086, Albany, NY 12222, USA.
Biomaterials (Impact Factor: 8.56). 04/2012; 33(11):3175-86. DOI: 10.1016/j.biomaterials.2012.01.010
Source: PubMed


Nanofiber scaffolds have been useful for engineering tissues derived from mesenchymal cells, but few studies have investigated their applicability for epithelial cell-derived tissues. In this study, we generated nanofiber (250 nm) or microfiber (1200 nm) scaffolds via electrospinning from the polymer, poly-l-lactic-co-glycolic acid (PLGA). Cell-scaffold contacts were visualized using fluorescent immunocytochemistry and laser scanning confocal microscopy. Focal adhesion (FA) proteins, such as phosphorylated FAK (Tyr397), paxillin (Tyr118), talin and vinculin were localized to FA complexes in adult cells grown on planar surfaces but were reduced and diffusely localized in cells grown on nanofiber surfaces, similar to the pattern observed in adult mouse salivary gland tissues. Significant differences in epithelial cell morphology and cell clustering were also observed and quantified, using image segmentation and computational cell-graph analyses. No statistically significant differences in scaffold stiffness between planar PLGA film controls compared to nanofibers scaffolds were detected using nanoindentation with atomic force microscopy, indicating that scaffold topography rather than mechanical properties accounts for changes in cell attachments and cell structure. Finally, PLGA nanofiber scaffolds could support the spontaneous self-organization and branching of dissociated embryonic salivary gland cells. Nanofiber scaffolds may therefore have applicability in the future for engineering an artificial salivary gland.

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Available from: Melinda Larsen
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    • "This leads to the production of micron- and submicron-scale fibers, such that electrospun meshes are characterized by feature sizes that are much smaller than in traditionally extruded meshes. Electrospun meshes have been shown to support cell attachment, alignment, proliferation and differentiation [69,70]. In the context of tissue engineering, however, the small pores limit in vitro cell infiltration as well as in vivo tissue ingrowth and vascularization within the bulk of the electrospun scaffold (ES), as these cellular and tissue ingrowth processes require pore sizes of up to 500 µm [67,71-73]. "
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    • "glands ( SMGs ) were dissected from timed - pregnant female mice ( strain CD - 1 , Charles River Laboratories ) at embryonic day 12 ( E12 ) or 13 ( E13 ) with the day of plug discovery designated as E0 , in accordance with protocols approved by the University at Albany IACUC . The SMGs were cultured as described previously ( Daley et al . , 2009 ; Sequeira et al . , 2012"
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    • "When peptides representing the epitope were available, peptide preabsorbed antibodies were exposed to salivary gland formalin-fixed, paraffinembedded (FFPE) sections to verify disappearance of the staining pattern (data not shown). All staining patterns on FFPE sections were also verified in whole mount salivary gland tissues fixed in 4% paraformaldehyde and 5% sucrose in 16 PBS, subjected to immunocytochemistry, and imaged using laser scanning confocal microscopy (510 Meta, Zeiss or SP5, Leica) (Larsen et al., 2003; Daley et al., 2009; Daley et al., 2011; Daley et al., 2012; Sequeira et al., 2012). The antibodies used in this study are listed in Table 1, and the order in which the immunohistochemistry steps were performed is listed in supplementary material Table S1. "
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