Cellular Responses Modulated by FGF-2 Adsorbed on Albumin/Heparin Layer-by-Layer Assemblies

PLoS One. 2015 May 6;10(5):e0125484. doi: 10.1371/journal.pone.0125484. eCollection 2015.

Abstract

In a typical cell culture system, growth factors immobilized on the cell culture surfaces can serve as a reservoir of bio-signaling molecules, without the need to supplement them additionally into the culture medium. In this paper, we report on the fabrication of albumin/heparin (Alb/Hep) assemblies for controlled binding of basic fibroblast growth factor (FGF-2). The surfaces were constructed by layer-by-layer adsorption of polyelectrolytes albumin and heparin and were subsequently stabilized by covalent crosslinking with glutaraldehyde. An analysis of the surface morphology by atomic force microscopy showed that two Alb/Hep bilayers are required to cover the surface of substrate. The formation of the Alb/Hep assemblies was monitored by the surface plasmon resonance (SPR), the infrared multiinternal reflection spectroscopy (FTIR MIRS) and UV/VIS spectroscopy. The adsorption of FGF-2 on the cross-linked Alb/Hep was followed by SPR. The results revealed that FGF-2 binds to the Alb/Hep assembly in a dose and time-dependent manner up to the surface concentration of 120 ng/cm(2). The bioactivity of the adsorbed FGF-2 was assessed in experiments in vitro, using calf pulmonary arterial endothelial cells (CPAE). CPAE cells could attach and proliferate on Alb/Hep surfaces. The adsorbed FGF-2 was bioactive and stimulated both the proliferation and the differentiation of CPAE cells. The improvement was more pronounced at a lower FGF-2 surface concentration (30 ng/cm(2)) than on surfaces with a higher concentration of FGF-2 (120 ng/cm(2)).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Albumins / pharmacology*
  • Animals
  • Cattle
  • Cell Adhesion / drug effects
  • Cell Culture Techniques / methods*
  • Cell Differentiation / drug effects
  • Cell Proliferation / drug effects
  • Culture Media / chemistry
  • Endothelial Cells / cytology*
  • Fibroblast Growth Factor 2 / metabolism
  • Fibroblast Growth Factor 2 / pharmacology*
  • Heparin / pharmacology*
  • Humans
  • Microscopy, Atomic Force
  • Protein Binding

Substances

  • Albumins
  • Culture Media
  • Fibroblast Growth Factor 2
  • Heparin

Grants and funding

The study was funded by the Ministry of Education, Youth and Sports, Czech Republic (grant number EE2.3.30.0029), by the Grant Agency of the Czech Republic (grant number P108/11/1857 and grant number P108/12/P624), and by the European Regional Development Fund (Project BIOCEV: Biotechnology and Biomedicine Centre of the Academy of Sciences and Charles University, grant number CZ.1.05/1.1.00/02.0109). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.