The isolation of cell derived extracellular matrix constructs using sacrificial open-cell foams.
Extracellular matrix derived from human and animal tissues is being used to repair and reconstruct a variety of tissues clinically. The utility of such constructs is limited by the geometry, composition and constitutive properties of the tissue or organ from which the ECM is harvested. To address this limitation, we have developed an approach to isolate extracellular matrix in bulk from populations of living cells grown in culture on three-dimensional substrates. Human biopsy derived fibroblasts were seeded within open-cell foams and cultured in-vitro for periods up to three weeks, after which the synthetic component was removed by incubation in a water miscible solvent. After several wash steps and lyophilization, a white, lacy, multi-molecular construct was isolated. Tandem mass spectroscopy showed that it contained 22 extracellular matrix constituents, including such proteins and proteoglycans as collagen type I and type III, fibronectin, transforming growth factor beta, decorin and biglycan among others. On average 47 mg of construct was isolated for each gram of synthetic substrate initially seeded with cells. The biomaterial harvested from human tracheal fibroblasts had an elastic modulus (250 kPa) and a composition similar to that of human vocal fold tissue, and supported reseeding with human tracheal derived fibroblasts. An important finding was that the approach was useful in isolating ECM from a variety of cell lineages and developmental stages including skin fibroblasts, brain derived astrocytes and mesenchymal stem cells. The results, together with the archival literature, suggest that the approach can be used to produce a range of cell derived constructs with unique physical and chemical attributes for a variety of research and medical applications.
The Keck Center for Tissue Engineering, Department of Bioengineering, College of Engineering, University of Utah, 20 S 2030 E Building, 570 BPRB, Room 108D, Salt Lake City, UT 84112, USA.
Jeffrey C
JC
Wolchok
Patrick A
PA
Tresco
D000818:Animals; D001672:Biocompatible Materials / Q000494:pharmacology*; D001696:Biomechanical Phenomena / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects; D005452:Fluoresceins / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence; D011108:Polymers / Q000494:pharmacology*; D051381:Rats; D017550:Spectroscopy, Fourier Transform Infrared; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds / Q000737:chemistry*; D014746:Vimentin / Q000378:metabolism
D001672:Biocompatible Materials; D005452:Fluoresceins; D011108:Polymers; D014746:Vimentin; C085925:calcein AM