title: Different culture conditions affect the growth of human tendon stem/progenitor cells (TSPCs) within a mixed tendon cells (TCs) population.
abstract: BACKGROUND
  Tendon resident cells (TCs) are a mixed population made of terminally differentiated tenocytes and tendon stem/progenitor cells (TSPCs). Since the enrichment of progenitors proportion could enhance the effectiveness of treatments based on these cell populations, the interest on the effect of culture conditions on the TSPCs is growing. In this study the clonal selection and the culture in presence or absence of basic fibroblast growth factor (bFGF) were used to assess their influences on the stemness properties and phenotype specific features of tendon cells.
  
  
  METHODS
  Cells cultured with the different methods were analyzed in terms of clonogenic and differentiation abilities, stem and tendon specific genes expression and immunophenotype at passage 2 and passage 4.
  
  
  RESULTS
  The clonal selection allowed to isolate cells with a higher multi-differentiation potential, but at the same time a lower proliferation rate in comparison to the whole population. Moreover, the clones express a higher amounts of stemness marker OCT4 and tendon specific transcription factor Scleraxis (SCX) mRNA, but a lower level of decorin (DCN). On the other hand, the number of cells obtained by clonal selection was extremely low and most of the clones were unable to reach a high number of passages in cultures. The presence of bFGF influences TCs morphology, enhance their proliferation rate and reduce their clonogenic ability. Interestingly, the expression of CD54, a known mesenchymal stem cell marker, is reduced in presence of bFGF at early passages. Nevertheless, bFGF does not affect the chondrogenic and osteogenic potential of TCs and the expression of tendon specific markers, while it was able to downregulate the OCT4 expression.
  
  
  CONCLUSIONS
  This study showed that clonal selection enhance progenitors content in TCs populations, but the extremely low number of cells produced with this method could represent an insurmountable obstacle to its application in clinical approaches. We observed that the addition of bFGF to the culture medium promotes the maintenance of a higher number of differentiated cells, reducing the proportion of progenitors within the whole population. Overall our findings demonstrated the importance of the use of specific culture protocols to obtain tendon cells for possible clinical applications.
authors.affiliation: Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.|Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milan, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Viganò
authors.affiliation: Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.|Department of Drug Sciences, University of Pavia, Pavia, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Perucca Orfei
authors.affiliation: Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Colombini
authors.affiliation: Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.
authors.forename: D
authors.initials: D
authors.lastname: Stanco
authors.affiliation: IRCCS Policlinico San Donato, San Donato Milanese, Milan, Italy.|Department of Biomedical Sciences for Health, University of Milan, San Donato Milanese, Milan, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Randelli
authors.affiliation: IRCCS Galeazzi Orthopaedic Institute, Milan, Italy.|Department of Biomedical Sciences for Health, University of Milano, Milan, Italy.
authors.forename: V
authors.initials: V
authors.lastname: Sansone
authors.affiliation: Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy. laura.degirolamo@grupposandonato.it.
authors.forename: L
authors.initials: L
authors.lastname: de Girolamo
keywords: Clonal selection; FGF; Progenitor cells; Tendon; Tendon cells; Tenocytes