title: Isolation and characterization of mesenchymal stem cells from the fat layer on the density gradient separated bone marrow.
abstract: The density gradient centrifugation method was originally designed for the isolation of mononuclear peripheral blood cells and rapidly adapted to fractionate bone marrow (BM) cells. This method involves the use of gradient density solutions with low viscosity and low osmotic pressure that allows erythrocytes and more mature cells gravitate to the bottom at a density fraction superior to 1.080 g/dL; mononuclear cells (MNCs) held in the plasma-solution to interphase at a density between 1.053 and 1.073 g/dL; plasma, dilution medium and anticoagulant to occupy a density less than 1.050 g/dL and the fat cells to float due to their very low density. BM-mesenchymal stem cells (MSCs) are usually obtained after the separation and cultures of BM-MNCs from the plasma-solution interphase, which is traditionally considered the only source of progenitor cells (hematopoietic and nonhematopoietic). In this study evidences that MSCs could be isolated from the very low-density cells of the fat layer are presented. In addition, we demonstrated that the MSCs obtained from these cells have similar immunophenotypic characteristics, and similar proliferative and differentiation potential to those obtained from the MNCs at plasma-solution interphase. The method represents a simple and cost effective way to increase the MSCs yield from each BM donor, without the need to look for other sources, additional manipulation of cells, and risks of contamination or disturbances of the potential of differentiation. These cells might serve as a complementary source of MSCs to facilitate preclinical and clinical application in tissue engineering and cell therapy.
authors.affiliation: Unidad de Terapia Celular, Servicio de Hematología, Hospital Universitario Virgen de la Arrixaca, El Palmar, Murcia, Spain. caymed@gmail.com
authors.forename: Carmen L
authors.initials: CL
authors.lastname: Insausti
authors.forename: Miguel Blanquer
authors.initials: MB
authors.lastname: Blanquer
authors.forename: Luis Meseguer
authors.initials: LM
authors.lastname: Olmo
authors.forename: María C
authors.initials: MC
authors.lastname: López-Martínez
authors.forename: Xavier Férez
authors.initials: XF
authors.lastname: Ruiz
authors.forename: Francisco J Rodríguez
authors.initials: FJ
authors.lastname: Lozano
authors.forename: Valentín Cabañas
authors.initials: VC
authors.lastname: Perianes
authors.forename: Consuelo
authors.initials: C
authors.lastname: Funes
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Nicolás
authors.forename: María J
authors.initials: MJ
authors.lastname: Majado
authors.forename: José M Moraleda
authors.initials: JM
authors.lastname: Jiménez
mesh_terms: D000293:Adolescent; D000328:Adult; D001854:Bone Marrow Cells / Q000166:cytology* / Q000502:physiology; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods*; D002499:Centrifugation, Density Gradient; D002648:Child; D005260:Female; D006801:Humans; D016130:Immunophenotyping; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000502:physiology; D008875:Middle Aged