title: RIA fractions contain mesenchymal stroma cells with high osteogenic potency.
abstract: BACKGROUND
  The gold standard for treatment of non-union is the transplantation of autologous bone from iliac crest. As an alternative, material can be harvested by femoral reaming with the Reamer-Irrigator-Aspirator(®) (RIA)-System. This material might be a source for human mesenchymal stroma cells (MSCs) with osteogenic potency. The aim of this study was the characterisation of cells harvested with the RIA system and the comparison of their properties with cells isolated from bone marrow ("BM") and fat tissue ("adipose"). The RIA material was separated into the liquid aspiration fraction ("liquid") and the solid RIA fraction. From the solid RIA fraction the cells were cultured either directly ("native") or after collagenase digestion and filtration ("filtrate"). Stem cell characteristics were analysed and the osteogenic potential was investigated in vitro and in vivo.
  
  
  METHODS
  Fat tissue and bone marrow were harvested from nine patients (three women, six males, with a mean of 48.1 years) with atrophic non-union RIA material. The cells were isolated and characterised by flow cytometry, three lineage differentiation capacities and colony-forming unit fibroblast assay. Gene expression profiles were performed and osteogenic differentiation in vivo was analysed.
  
  
  RESULTS
  All three RIA fractions contained mesenchymal stromal cells (MSCs) as demonstrated by CFU-F assay, three linage differentiation and surface marker analysis. The RIA-MSCs exhibited a significantly higher osteogenic potential in vitro compared to adipose-MSCs, whereas no difference was seen compared to BM-MSCs. Quantitative RT-PCR analysis revealed an expression of osteogenic markers in all isolated cells. The implantation of MSCs with β-TCP scaffolds into the mice muscle showed significantly higher bone formation for the filtrate RIA-MSC, native RIA-MSC and BM-MSC groups compared to the adipose-MSC group. The filtrate RIA-MSCs formed twice as much new bone in vivo compared to BM-MSCs.
  
  
  CONCLUSIONS
  The present study showed high potency of cells isolated by reaming. Even in the irrigation fluid, which is normally discarded, cells with the characteristics of stromal stem cells were isolated. In comparison to adipose-MSCs and BM-MSCs, the RIA-MSCs showed a similar or even better osteogenic potential in vitro and in vivo and this supports their usability in orthopaedic surgery.
authors.affiliation: Trauma and Reconstructive Surgery, Center for Orthopaedics, Trauma Surgery and Spinal Cord Injury, Heidelberg University Hospital, Schlierbacher Landstraβe 200a, D-69118 Heidelberg, Germany, HTRG - Heidelberg Trauma Research Group.
authors.forename: Pamela
authors.initials: P
authors.lastname: Kuehlfluck
authors.affiliation: Trauma and Reconstructive Surgery, Center for Orthopaedics, Trauma Surgery and Spinal Cord Injury, Heidelberg University Hospital, Schlierbacher Landstraβe 200a, D-69118 Heidelberg, Germany, HTRG - Heidelberg Trauma Research Group.
authors.forename: Arash
authors.initials: A
authors.lastname: Moghaddam
authors.affiliation: Trauma and Reconstructive Surgery, Center for Orthopaedics, Trauma Surgery and Spinal Cord Injury, Heidelberg University Hospital, Schlierbacher Landstraβe 200a, D-69118 Heidelberg, Germany, HTRG - Heidelberg Trauma Research Group.
authors.forename: Lars
authors.initials: L
authors.lastname: Helbig
authors.affiliation: Trauma and Reconstructive Surgery, Center for Orthopaedics, Trauma Surgery and Spinal Cord Injury, Heidelberg University Hospital, Schlierbacher Landstraβe 200a, D-69118 Heidelberg, Germany, HTRG - Heidelberg Trauma Research Group.
authors.forename: Christopher
authors.initials: C
authors.lastname: Child
authors.affiliation: Julius Wolff Institute, Berlin-Brandenburg Center for Regenerative Therapies, Center for Musculoskeletal Surgery, Charité - Universitätsmedizin Berlin, Germany.
authors.forename: Britt
authors.initials: B
authors.lastname: Wildemann
authors.affiliation: Trauma and Reconstructive Surgery, Center for Orthopaedics, Trauma Surgery and Spinal Cord Injury, Heidelberg University Hospital, Schlierbacher Landstraβe 200a, D-69118 Heidelberg, Germany, HTRG - Heidelberg Trauma Research Group. Electronic address: Gerhard.Schmidmaier@med.uni-heidelberg.de.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Schmidmaier
mesh_terms: D000273:Adipose Tissue / Q000166:cytology*; D000328:Adult; D000368:Aged; D001861:Bone Regeneration / Q000502:physiology; D016025:Bone Transplantation; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D004867:Equipment Design; D005260:Female; D005434:Flow Cytometry; D017759:Fractures, Malunited / Q000473:pathology* / Q000601:surgery; D006801:Humans; D007085:Ilium / Q000166:cytology*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008875:Middle Aged; D010012:Osteogenesis / Q000502:physiology*; D020858:Tissue and Organ Harvesting / Q000295:instrumentation*; D014182:Transplantation, Autologous
keywords: RIA fractions; Reamer-Irrigator-Aspirator; fat tissue (adipose-MSCs); human mesenchymal stroma cells (MSCs); iliac crest bone marrow aspirate (BM-MSCs); osteogenic potency