Comparative proteomic analyses of human adipose extracellular matrices decellularized using alternative procedures

Abstract

Decellularized human adipose tissue has potential clinical utility as a processed biological scaffold for soft tissue cosmesis, grafting, and reconstruction. Adipose tissue decellularization has been accomplished using enzymatic-, detergent-, and/or solvent-based methods. To examine the hypothesis that distinct decellularization processes may yield scaffolds with differing compositions, the current study employed mass spectrometry to compare the proteomes of human adipose-derived matrices generated through three independent methods combining enzymatic-, detergent-, and/or solvent-based steps. In addition to protein content, bioscaffolds were evaluated for deoxyribose nucleic acid depletion, extracellular matrix composition, and physical structure using optical density, histochemical staining, and scanning electron microscopy. Mass spectrometry based proteomic analyses identified 25 proteins (having at least two peptide sequences detected) in the scaffolds generated with an enzymatic approach, 143 with the detergent approach, and 102 with the solvent approach, as compared to 155 detected in unprocessed native human fat. Immunohistochemical detection confirmed the presence of the structural proteins actin, collagen type VI, fibrillin, laminin, and vimentin. Subsequent in vivo analysis of the predominantly enzymatic- and detergent-based decellularized scaffolds following subcutaneous implantation in GFP transgenic mice demonstrated that the matrices generated with both approaches supported the ingrowth of host-derived adipocyte progenitors and vasculature in a time dependent manner. Together, these results determine that decellularization methods influence the protein composition of adipose tissue-derived bioscaffolds. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A:2481-2493, 2018.

Clinical Translational Potential in Skin Wound Regeneration for Adipose-Derived, Blood-Derived, and Cellulose Materials: Cells, Exosomes, and Hydrogels

“…The methodology for decellularizing adipose tissue was pioneered by Flynn and colleagues [ 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 , 71 , 72 , 73 ] and has since been confirmed by multiple independent laboratories [ 74 , 75 , 76 , 77 , 78 , 79 , 80 , 81 , 82 ]. These methods combine biological (enzyme digestion), chemical, and physical processing steps to achieve the uniform and consistent manufacture of protein scaffolds depleted of contaminating genomic DNA and lipids.…”

“…These methods combine biological (enzyme digestion), chemical, and physical processing steps to achieve the uniform and consistent manufacture of protein scaffolds depleted of contaminating genomic DNA and lipids. Obatala scientists, in collaboration with partners at Pennsylvania State University (PSU), Tulane University, and Western Ontario University, have developed and published on decellularized human adipose tissue-derived extracellular matrix (ECM) hydrogel, commercially termed “AdipoGel™” [ 70 , 83 , 84 , 85 , 86 ]. This hydrogel or scaffold is prepared using a combination of chemical and mechanical processing followed by protease digestion as a final modification prior to sterilization.…”

“…Furthermore, the decellularized adipose-derived scaffold can be chemically modified with thiol methacrylate and cross-linked to create a cytocompatible matrix that can be “tuned” to achieve biomechanical properties appropriate for the promotion of either soft or hard tissue regeneration [ 87 ]. Mass spectrometry analyses demonstrate that the AdipoGel proteome is enriched for ECM proteins, including collagens, fibrillin, laminin, nidogen, and other adhesion molecules, while the AdipoGel proteome is depleted in cytoplasmic and nuclear protein contents relative to native tissue [ 70 , 86 ]. The decellularized tissue can be prepared as a lyophilized scaffold or as a liquid hydrogel.…”

“…The decellularized tissue can be prepared as a lyophilized scaffold or as a liquid hydrogel. Both forms are compatible with in vivo applications, including subcutaneous implantation, injection for soft tissue regeneration, or as a sheet for use in orthopedic repair procedures [ 70 , 83 ]. Likewise, independent studies have demonstrated that their decellularized adipose tissue scaffolds can be used to create adipose depots when implanted into nude mice [ 82 ].…”

Human Adipose-Derived Hydrogel Characterization Based on In Vitro ASC Biocompatibility and Differentiation

“…A hydrogel of decellularized adipose tissue (DAT) can serve as an alternative ECM for research, due to the abundant availability of source tissue [11,12] and accumulating evidence of its cytocompatibility [13]. DAT hydrogel and its composites support in vitro ASC proliferation [14] and adipogenic differentiation [15][16][17] and have proven to be adipoinductive scaffolds [9,[18][19][20]. Additionally, intact DAT scaffolds have displayed positive outcomes in wound healing [21] and nerve repair [22].…”