| Autor: |
Frauchiger DA; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland., Tekari A; Laboratory of Molecular and Cellular Screening Processes, Centre of Biotechnology of Sfax, University of Sfax, Sfax, Tunisia., May RD; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland., Džafo E; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland., Chan SCW; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland., Stoyanov J; Swiss Paraplegic Research, Nottwil, Switzerland., Bertolo A; Swiss Paraplegic Research, Nottwil, Switzerland., Zhang X; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland., Guerrero J; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland.; Department of Orthopaedic Surgery and Traumatology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland., Sakai D; Department for Orthopaedic Surgery, Tokai University School of Medicine, Isehara, Japan., Schol J; Department for Orthopaedic Surgery, Tokai University School of Medicine, Isehara, Japan., Grad S; AO Research Institute, Davos, Switzerland., Tryfonidou M; Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands., Benneker LM; Department of Orthopaedic Surgery and Traumatology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland., Gantenbein B; Tissue Engineering, Orthopeadic Research & Mechanobiology, Department for BioMedical Research (DBMR), Medical Faculty, University of Bern, Bern, Switzerland.; Department of Orthopaedic Surgery and Traumatology, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. |
| Abstrakt: |
Low back pain related to intervertebral disk (IVD) degeneration has a major socioeconomic impact on our aging society. Therefore, stem cell therapy to activate self-repair of the IVD remains an exciting treatment strategy. In this respect, tissue-specific progenitors may play a crucial role in IVD regeneration, as these cells are perfectly adapted to this niche. Such a rare progenitor cell population residing in the nucleus pulposus (NP) (NP progenitor cells [NPPCs]) was found positive for the angiopoietin-1 receptor (Tie2+), and was demonstrated to possess self-renewal capacity and in vitro multipotency. Here, we compared three sorting protocols; that is, fluorescence-activated cell sorting (FACS), magnetic-activated cell sorting (MACS), and a mesh-based label-free cell sorting system (pluriSelect), with respect to cell yield, potential to form colonies (colony-forming units), and in vitro functional differentiation assays for tripotency. The aim of this study was to demonstrate the efficiency of three widespread cell sorting methods for picking rare cells (<5%) and how these isolated cells then behave in downstream functional differentiation in adipogenesis, osteogenesis, and chondrogenesis. The cell yields among the isolation methods differed widely, with FACS presenting the highest yield (5.0% ± 4.0%), followed by MACS (1.6% ± 2.9%) and pluriSelect (1.1% ± 1.0%). The number of colonies formed was not significantly different between Tie2+ and Tie2- NPPCs. Only FACS was able to separate into two functionally different populations that showed trilineage multipotency, while MACS and pluriSelect failed to maintain a clear separation between Tie2+ and Tie2- populations in differentiation assays. To conclude, the isolation of NPPCs was possible with all three sorting methods, while FACS was the preferred technique for separation of functional Tie2+ cells. Impact Statement Tissue-specific progenitor cells such as nucleus pulposus progenitor cells of the IVD could become an ultimate cell source for tissue engineering strategies as these cells are presumably best adapted to the tissue's microenvironment. Fluorescence-activated cell sorting seemed to outcompete magnetic-activated cell sorting and pluriSelect concerning selecting a rare cell population from IVD tissue as could be demonstrated by improved cell yield and functional differentiation assays. |
