title: The use of the reamer-irrigator-aspirator to harvest mesenchymal stem cells.
abstract: The scarcity of mesenchymal stem cells (MSCs) in iliac crest bone marrow aspirate (ICBMA), and the expense and time in culturing cells, has led to the search for alternative harvest sites. The reamer-irrigation-aspirator (RIA) provides continuous irrigation and suction during reaming of long bones. The aspirated contents pass via a filter, trapping bony fragments, before moving into a 'waste' bag from which MSCs have been previously isolated. We examined the liquid and solid phases, performed a novel digestion of the solid phase, and made a comparative assessment in terms of number, phenotype and differentiation capacity with matched ICBMA. The solid fraction from the filtrate was digested for 60 minutes at 37° C with collagenase. Enumeration was performed via the colony-forming unit fibroblast (CFU-F) assay. Passage (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages, and their phenotypes assessed using flow cytometry (CD33, CD34, CD45, CD73, CD90, and CD105). MSCs from the RIA phases were able to differentiate at least as well as those from ICBMA, and all fractions had phenotypes consistent with other established sources. The median number of colonies for the three groups was: ICBMA = 8.5 (2 to 86), RIA-liquid = 19.5 (4 to 90), RIA-solid = 109 (67 to 200) per 200 μl. The mean total yield of cells for the three groups was: ICBMA = 920 (0 to 4275), RIA-liquid = 114,983 (16,500 to 477,750), RIA-solid = 12,785 (7210 to 28 475). The RIA filtrate contains large numbers of MSCs that could potentially be extracted without enzymatic digestion and used for bone repair without prior cell expansion.
authors.affiliation: Department of Trauma and Orthopaedics, Academic Unit, Clarendon Wing, Leeds Teaching Hospitals NHS Trust, Great George Street, Leeds LS1 3EX, UK.
authors.forename: G
authors.initials: G
authors.lastname: Cox
authors.forename: D
authors.initials: D
authors.lastname: McGonagle
authors.forename: S A
authors.initials: SA
authors.lastname: Boxall
authors.forename: C T
authors.initials: CT
authors.lastname: Buckley
authors.forename: E
authors.initials: E
authors.lastname: Jones
authors.forename: P V
authors.initials: PV
authors.lastname: Giannoudis
mesh_terms: D000328:Adult; D001854:Bone Marrow Cells / Q000166:cytology*; D001861:Bone Regeneration / Q000502:physiology; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000502:physiology; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D004867:Equipment Design; D005260:Female; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008875:Middle Aged