title: Effect of coculturing canine notochordal, nucleus pulposus and mesenchymal stromal cells for intervertebral disc regeneration.
abstract: BACKGROUND
  Early degenerative changes in the nucleus pulposus (NP) are observed after the disappearance of notochordal cells (NCs). Thus, it has been suggested that NCs play an important role in maintaining the NP and may have a regenerative potential on other cells of the NP. As the number of resident NP cells (NPCs) decreases in a degenerating disc, mesenchymal stromal (stem) cells (MSCs) may be used for cell supplementation. In this study, using cells of one species, the regenerative potential of canine NCs was assessed in long-term three-dimensional coculture with canine NPCs or MSCs.
  
  
  METHODS
  Canine NCs and canine NPCs or MSCs were cocultured in alginate beads for 28 days under hypoxic and high-osmolarity conditions. Cell viability, cell morphology and DNA content, extracellular matrix production and expression of genes related to NC markers (Brachyury, KRT18) and NP matrix production (ACAN, COL2A1, COL1A1) were assessed after 1, 15 and 28 days of culture.
  
  
  RESULTS
  NCs did not completely maintain their phenotype (morphology, matrix production, gene expression) during 28 days of culture. In cocultures of NPCs and NCs, both extracellular matrix content and anabolic gene expression remained unchanged compared with monoculture groups, whereas cocultures of MSCs and NCs showed increased glycosaminoglycan/DNA. However, the deposition of these proteoglycans was observed near the NCs and not the MSCs. Brachyury expression in the MSC and NC coculture group increased in time. The latter two findings indicate a trophic effect of MSCs on NCs rather than vice versa.
  
  
  CONCLUSIONS
  No regenerative potential of canine NCs on canine NPCs or MSCs was observed in this study. However, significant changes in NC phenotype in long-term culture may have resulted in a suboptimal regenerative potential of these NCs. In this respect, NC-conditioned medium may be better than coculture for future studies of the regenerative potential of NCs.
authors.affiliation: Department of Biomedical Engineering, Eindhoven University of Technology, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands. i.t.m.arkesteijn@tue.nl.
authors.forename: Irene T M
authors.initials: IT
authors.lastname: Arkesteijn
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, PO Box 80.154, NL-3508 TD, Utrecht, The Netherlands. lsmolders@vetclinics.uzh.ch.|Clinic for Small Animal Surgery, Vetsuisse Faculty, Zurich University, Winterthurerstrasse 260, CH-8057, Zurich, Switzerland. lsmolders@vetclinics.uzh.ch.
authors.forename: Lucas A
authors.initials: LA
authors.lastname: Smolders
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, PO Box 80.154, NL-3508 TD, Utrecht, The Netherlands. s.spillekom@students.uu.nl.
authors.forename: Sandra
authors.initials: S
authors.lastname: Spillekom
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, PO Box 80.154, NL-3508 TD, Utrecht, The Netherlands. f.m.riemers@uu.nl.
authors.forename: Frank M
authors.initials: FM
authors.lastname: Riemers
authors.affiliation: Department of Biomedical Engineering, Eindhoven University of Technology, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands. esther.potier@paris7.jussieu.fr.|Laboratoire de Bioingénierie et Biomécanique Ostéo-Articulaire (B2OA), UMR CNRS 7052, Université Denis Diderot Paris 7, Sorbonne Paris Cité, 690, Paris, France. esther.potier@paris7.jussieu.fr.
authors.forename: Esther
authors.initials: E
authors.lastname: Potier
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, PO Box 80.154, NL-3508 TD, Utrecht, The Netherlands. b.p.meij@uu.nl.
authors.forename: Björn P
authors.initials: BP
authors.lastname: Meij
authors.affiliation: Department of Biomedical Engineering, Eindhoven University of Technology, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands. k.ito@tue.nl.|Department of Orthopedics, University Medical Center Utrecht, P.O. Box 85500, HP G05.228, 3508 GA, Utrecht, The Netherlands. k.ito@tue.nl.
authors.forename: Keita
authors.initials: K
authors.lastname: Ito
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, PO Box 80.154, NL-3508 TD, Utrecht, The Netherlands. m.a.tryfonidou@uu.nl.
authors.forename: Marianna A
authors.initials: MA
authors.lastname: Tryfonidou
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002470:Cell Survival; D002478:Cells, Cultured; D002574:Cervical Vertebrae; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004195:Disease Models, Animal; D004285:Dogs; D007403:Intervertebral Disc / Q000473:pathology*; D055959:Intervertebral Disc Degeneration / Q000473:pathology*; D008159:Lumbar Vertebrae; D059630:Mesenchymal Stem Cells / Q000473:pathology*; D009672:Notochord / Q000473:pathology*; D013904:Thoracic Vertebrae
chemical_list: D017077:Culture Media, Conditioned