(505d) Heparin/Collagen Coatings Improve Human Mesenchymal Stromal Cell Response to Interferon Gamma

Authors: 
Almodovar, J., University of Arkansas
Castilla-Casadiego, D., University of Arkansas
García, J. R., Georgia Institute of Technology
Garcia, A., Georgia Institute of Technology
Mesenchymal stromal cells (MSC) are a promising source for cell-based therapies since they secrete a myriad of reparative factors in response to inflammatory stimuli. In this project, the influence of interferon gamma (IFN-g) in contact with multilayers of collagen/heparin (COL/HEP) was evaluated on the cell response of two human donor patients to secrete multiple human biomarkers including vascular endothelial growth factor (VEGF-A), fibroblast growth factor (FGF-2), colony stimulating factor 1 (M-CSF), interleukin 10 (IL-10) and interleukin 6 (IL-6). In addition, the proliferation potential and cellular adhesion of human mesenchymal stromal cells (h-MSCs) was evaluated. Multilayers varying the final layer between COL (6 bilayers) and HEP (6.5 bilayers) were fabricated by using Layer by Layer technique, and fully characterized by Infrared Variable Angle Spectroscopic Ellipsometry. In-vitro studies consisted on evaluating the cellular adhesion (Immunostaining), proliferation response (EDU Imaging Kit), and protein expression of h-MSCs seeded onto the multilayers (Luminex assay). Infrared Variable Angle Spectroscopic Ellipsometry confirmed the physical-chemical properties of the multilayers fabricated on the substrate. Characteristic peaks for COL and HEP could be observed in the spectrums obtained at 25 °C (temperature of fabrication of multilayers) and 37°C (temperature of incubation). IRVASE reported an average thickness of 129 nm for 6 bilayers and 178 nm for 6.5 bilayers. For roughness, 6 bilayers presented an average of 168 nm and 6.5 bilayers a value of 230 nm. An EdU incorporation assay revealed that IFN-g had an anti-proliferative effect in all conditions evaluated except when h-MSCs were cultured on HEP-ending films supplemented with IFN-g. Moreover, h-MSCs cultured on HEP-ending films supplemented with IFN-g had a higher cytokine expression as compared to cells cultured on tissue culture polystyrene, COL-ending films with and without IFN-g, and HEP-ending films without IFN-g as measured by Luminex assay. Finally, immunostaining revealed strong integrin binding and FAK phosphorylation for each condition. This study shows that HEP/COL films can modulate h-MSC response to soluble factors, which may be exploited in cell manufacturing practices.