Real-Time Quantification of Proteins Secreted by Artificial Connective Tissue Made from Uni- Or Multidirectional Collagen I Scaffolds and Oral Mucosa Fibroblasts

• Published in: Analytical chemistry (Washington) 2014-03, Vol.86 (5), p.2421-2428
• Main Authors: Bustos, Rosa Helena, Suesca, Edward, Millán, Diana, González, José Manuel, Fontanilla, Marta R.
• Format: Article
• Language: English
• Subjects: Cellular biology; Collagen; Collagen Type I - chemistry; Collagens; Connective tissue; Connective Tissue - chemistry; Culture Media, Conditioned; Enzyme-Linked Immunosorbent Assay; Fiber orientation; Fibroblasts; Fibroblasts - chemistry; Microstructure; Morphology; Mouth Mucosa - chemistry; Mouth Mucosa - cytology; Pharmaceuticals; Proteins; Proteins - analysis; Scaffolds; Secretions; Surface Plasmon Resonance; Tissues
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac4033164

Previously, we found that oral autologous artificial connective tissue (AACT) had a different protein secretion profile to that of clot-embedded AACT. Other oral mucosa substitutes, having different cell types and scaffolds, had dissimilar secretion profiles of proteins (including that for AACT) that influence healing outcome; thus, to ascertain the profiles of factors secreted by artificial tissue and whether they are influenced by their microstructure might help in understanding their bioactivity. An important component of tissue microstructure is the fiber orientation of the scaffold used for manufacturing it. This work developed a surface plasmon resonance (SPR) methodology to quantify factors secreted by oral artificial connective tissue (ACT) in culture medium, and a method to manufacture unidirectional laminar collagen I scaffolds. The SPR methodology was used for assessing differences in the protein secretion profile of ACT made with collagen scaffolds having different fiber orientation (unidirectional vs multidirectional). Oral fibroblasts seeded onto unidirectional scaffolds increased the secretion of six factors involved in modulating healing compared to those seeded onto multidirectional scaffolds. Histological analysis of uni- and multidirectional ACT showed that cells differ in their alignment and morphology. This SPR-methodology led to nanoscale detection of paracrine factors and might be useful to study biomarkers of three-dimensional cell growth, cell differentiation, and wound-healing progression.