Preserving integrity: innovative in vitro methods for extracellular matrix decellularization and collagen purification

Abstract

Background: In tissue engineering and cell therapy development, synthetic biomaterials are frequently supplemented with collagen or other extracellular matrix (ECM) components to enhance biocompatibility. To support these applications, novel methods for collagen purification and ECM decellularization were developed, with a focus on preserving the structural and biochemical integrity of the final products. Results: The effectiveness of these methods was validated using solid-state NMR and fluorescence spectroscopy, bright-field and confocal microscopy, amino acid analysis, proteomics and transmission electron microscopy. Intact cells were dislodged from ECM-producing cultures through the application of cytoskeleton-targeting drugs, while the native protein composition of the ECM was maintained. In parallel, collagen purified using chymotrypsin was shown to retain its native triple-helical structure and post-translational modifications. Conclusions: Both techniques are broadly applicable to various cell types capable of producing collagen and/or ECM in vitro, thereby expanding the availability of species- and tissue-specific sources. These advances hold particular promise for human-relevant tissue engineering and drug discovery applications.