title: In vitro changes in human tenocyte cultures obtained from proximal biceps tendon: multiple passages result in changes in routine cell markers.
abstract: OBJECTIVE
  Results of in vitro cell models are commonly used to promote new therapies (e.g., platelet-rich plasma), and clinicians have to be aware of the specific limitations of such models. To gain a sufficient and effective cell load, many current in vitro models use cells multiplied through various passages. This is especially important in tendon-like cell (TLC) models, since native tendon tissue is not available unlimited and contains limited amount of tenocytes. The purpose was to determine the occurrence of phenotypic changes following extended monolayer culture of TLCs, according to cell-passage number.
  
  
  METHODS
  Tendon samples were obtained from 15 healthy patients undergoing biceps tenodesis. Tendons were digested and cultured (monolayer) for six passages. Tendon-specific markers (collagens I and III, decorin, tenascin-C, and tenomodulin) and their histology were analyzed using gene expression and protein content assays. Native cells, the cells cultured and cells passaged one to six times were analyzed at each passage.
  
  
  RESULTS
  Gene expression of types I and III collagen of cultured TLCs significantly decreased after two passages. Gene expression of decorin, tenascin-C, and tenomodulin exhibited a trend of decreased gene expression with increased passage. Protein levels of types I and III collagen and decorin decreased after four passages.
  
  
  CONCLUSIONS
  The significant findings let conclude that tenocyte-like cells obtained from human LHB can be maintained in monolayer culture at low passages, before the signs of phenotypic drift are present. But researchers must be aware of rapid phenotypic drift at higher passage numbers. Therefore, only cells within the first 3 passages should be used as a precaution for in vitro monolayer cell models, and one has to be aware of the phenotypic changes if TLCs passaged multiple times are used. The clinical relevance of this data is that understanding of in vitro TLC models, and their limitations may finally help the clinician to judge the potential of experimental data of new biologic treatment options.
authors: Department of Orthopaedic Surgery, University of Connecticut Health Center, Farmington, CT 06030, USA. admazzocca@yahoo.com
authors: Augustus D
authors: AD
authors: Mazzocca
authors: David
authors: D
authors: Chowaniec
authors: Mary Beth
authors: MB
authors: McCarthy
authors: Knut
authors: K
authors: Beitzel
authors: Mark P
authors: MP
authors: Cote
authors: William
authors: W
authors: McKinnon
authors: Robert
authors: R
authors: Arciero
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D024042:Collagen Type I / Q000096:biosynthesis / Q000235:genetics; D024061:Collagen Type III / Q000096:biosynthesis / Q000235:genetics; D058575:Decorin / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D018482:Muscle, Skeletal; D019063:Tenascin / Q000096:biosynthesis / Q000235:genetics; D013710:Tendons / Q000166:cytology* / Q000378:metabolism* / Q000601:surgery; D053656:Tenodesis
chemical_list: D015415:Biomarkers; D024042:Collagen Type I; D024061:Collagen Type III; D058575:Decorin; D008565:Membrane Proteins; C422444:TNMD protein, human; D019063:Tenascin