title: Matrix metalloproteinases as reagents for cell isolation.
abstract: Cell isolation methods for therapeutic purposes have seen little advancement over the years. The original methods of stem cell and islet isolation using bacterial collagenases were developed in the early 1980s and are still used today. Bacterial collagenases are subject to autodegradation, and isolates obtained with these enzymes may be contaminated with endotoxins, reducing cell viability and contributing to toxicity in downstream applications. Here we describe a novel method for isolation of mesenchymal stem cells from adipose tissue (ADSC) utilizing recombinantly produced matrix metalloproteases (MMPs). The ADSCs isolated by MMPs displayed essentially identical morphological and phenotypical characteristics to cells isolated by bacterially-derived collagenase I and Liberase™. Samples isolated with MMPs and Liberase™ had comparable levels of CD73, CD90, and CD105. The adipogenic and osteogenic potential of the ADSCs isolated by MMPs was retained as compared to cells isolated with Liberase™. However, ADSCs isolated by Liberase™ displayed 6% contamination with other cells as per negative markers revealed by PE staining, as opposed to<1% for all MMP-treated samples. MMP-based cell isolation may contribute to optimization of transplantation technology.
authors.affiliation: Florida Atlantic University, Department of Chemistry & Biochemistry, Jupiter, FL 33458, United States; Torrey Pines Institute for Molecular Studies, Departments of Chemistry and Biology, Port St. Lucie, FL 34987, United States.
authors.forename: Anna M
authors.initials: AM
authors.lastname: Knapinska
authors.affiliation: Florida Atlantic University, Department of Chemistry & Biochemistry, Jupiter, FL 33458, United States; Torrey Pines Institute for Molecular Studies, Departments of Chemistry and Biology, Port St. Lucie, FL 34987, United States.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Amar
authors.affiliation: Vaccine & Gene Therapy Institute of Florida, Port Saint Lucie, FL 34987, United States.
authors.forename: Zhong
authors.initials: Z
authors.lastname: He
authors.affiliation: Akron Biotech, Boca Raton, Florida 33487, United States.
authors.forename: Sandro
authors.initials: S
authors.lastname: Matosevic
authors.affiliation: Akron Biotech, Boca Raton, Florida 33487, United States.
authors.forename: Claudia
authors.initials: C
authors.lastname: Zylberberg
authors.affiliation: Florida Atlantic University, Department of Chemistry & Biochemistry, Jupiter, FL 33458, United States; Torrey Pines Institute for Molecular Studies, Departments of Chemistry and Biology, Port St. Lucie, FL 34987, United States; The Scripps Research Institute/Scripps Florida, Jupiter, FL 33458, United States. Electronic address: fieldsg@fau.edu.
authors.forename: Gregg B
authors.initials: GB
authors.lastname: Fields
mesh_terms: D050156:Adipogenesis; D000273:Adipose Tissue / Q000166:cytology; D002454:Cell Differentiation; D002469:Cell Separation / Q000379:methods*; D017364:Collagenases / Q000378:metabolism; D004731:Endotoxins / Q000032:analysis; D004789:Enzyme Activation; D005434:Flow Cytometry; D058851:GPI-Linked Proteins / Q000378:metabolism; D006801:Humans; D016130:Immunophenotyping; D007202:Indicators and Reagents; D053508:Matrix Metalloproteinase 12 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism; D053504:Matrix Metalloproteinases, Membrane-Associated / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology; D010012:Osteogenesis; D011994:Recombinant Proteins / Q000378:metabolism; D013820:Thermolysin / Q000378:metabolism
chemical_list: D004731:Endotoxins; D058851:GPI-Linked Proteins; D007202:Indicators and Reagents; D011994:Recombinant Proteins; D017364:Collagenases; C106635:Liberase; D020782:Matrix Metalloproteinases; D053504:Matrix Metalloproteinases, Membrane-Associated; C403928:matrix metalloproteinase 25; C511768:MMP3 protein, human; D019278:Matrix Metalloproteinase 3; D013820:Thermolysin; C507122:MMP12 protein, human; D053508:Matrix Metalloproteinase 12
keywords: Adipose tissue; Clostridium histolyticum collagenase; E. coli; Matrix metalloproteinase; Recombinant enzyme; Stem cell