title: DiI labeling of human adipose-derived stem cells: evaluation of DNA damage, toxicity and functional impairment.
abstract: BACKGROUND
  Adipose tissue-derived stem cells (ASCs) have become the primary focus of tissue engineering research. To understand their functions and behavior in in vitro and in vivo models, it is mandatory to track the implanted cells and distinguish them from the resident or host cells. A common labeling method is the use of fluorescent dyes, e.g. the lipophilic carbocyanine dye, DiI. This study aimed to analyze potential DNA damage, toxicity and impairment of the functional properties of human ASCs after labeling with DiI.
  
  
  METHODS
  Cytotoxicity was measured using the MTT assay and DNA damage was determined by means of the comet assay. Potential apoptotic effects were determined using the annexin V-propidium iodide test. Differentiation potential was evaluated by trilineage differentiation procedures in labeled and unlabeled ASCs. Proliferation as well as migration capability was analyzed, and the duration and stability of DiI labeling in ASCs during in vitro expansion was observed over a period of 35 days.
  
  
  RESULTS
  DiI labeling did not cause genotoxic effects 15, or 30 min or 24 h after the labeling procedure, and there were no cytotoxic effects until 72 h afterwards. No impairment of proliferation or migration capability or differentiation potential could be determined. However, after 35 days, only 37% of labeled cells could be detected using the fluorescence microscope, which indicates a decrease in staining stability during in vitro expansion.
  
  
  CONCLUSIONS
  DiI is a convenient method for ASCs labeling which causes no toxic effects and does not impair the proliferation, migration or differentiation potential of ASCs after the labeling procedure.
authors.affiliation: Department of Otorhinolaryngology, Plastic, Aesthetic and Reconstructive Head and Neck Surgery, University of Wuerzburg, Wuerzburg, Germany. froelich_k@klinik.uni-wuerzburg.de
authors.forename: K
authors.initials: K
authors.lastname: Froelich
authors.forename: G
authors.initials: G
authors.lastname: Steussloff
authors.forename: K
authors.initials: K
authors.lastname: Schmidt
authors.forename: M
authors.initials: M
authors.lastname: Ramos Tirado
authors.forename: A
authors.initials: A
authors.lastname: Technau
authors.forename: A
authors.initials: A
authors.lastname: Scherzed
authors.forename: S
authors.initials: S
authors.lastname: Hackenberg
authors.forename: A
authors.initials: A
authors.lastname: Radeloff
authors.forename: R
authors.initials: R
authors.lastname: Hagen
authors.forename: N
authors.initials: N
authors.lastname: Kleinsasser
mesh_terms: D000273:Adipose Tissue / Q000166:cytology*; D017304:Annexin A5 / Q000378:metabolism; D002232:Carbocyanines / Q000378:metabolism* / Q000633:toxicity*; D016923:Cell Death / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D019070:Cell Lineage / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D020552:Comet Assay; D004249:DNA Damage; D006801:Humans; D010641:Phenotype; D011419:Propidium / Q000378:metabolism; D013194:Staining and Labeling; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D014343:Trypan Blue / Q000378:metabolism
chemical_list: D017304:Annexin A5; D002232:Carbocyanines; D011419:Propidium; C024286:3,3'-dioctadecylindocarbocyanine; D014343:Trypan Blue