title: Serum-free medium and hydroxyethyl starch supports cell survival better than Minimal Essential Medium and dextran in organ-cultured mouse corneas.
abstract: OBJECTIVE
  In a previous study, we observed a deleterious effect of serum-supplemented Minimal Essential Medium (MEM) on human corneal endothelial cell survival in a cell culture model. Consequently, here we studied the effects of conventional, serum-supplemented MEM and a serum-free medium in combination with two different deswelling substances on cell survival in whole corneas in a mouse model.
  
  
  METHODS
  Murine corneas were cultured for 4 days in MEM+2% fetal calf serum (FCS) or serum-free Human Endothelial-SFM (SFM), both supplemented with either 6% dextran T500 or 7.5% hydroxyethyl starch (HES) 130/0.4. Cells were examined by differential interference contrast microscopy, H&E staining, immunocytochemistry for cleaved caspase-3, Bcl-2, haem oxygenase-1 and immunoblotting for cleaved caspase-3.
  
  
  RESULTS
  In MEM, endothelial cells were almost completely lost after 4 days and the number of epithelial cells was markedly reduced. The remaining cells showed fragmented nuclei and were positive for cleaved caspase-3 and strongly positive for Bcl-2. Corneas cultured in SFM retained an almost closed layer of endothelial cells. Fewer cells were positive for Bcl-2, and only a few cells were positive for cleaved caspase-3 even under staurosporine administration. HES supplementation was well tolerated by corneal cells over 4 days, while a 4-day supplementation with dextran resulted in the loss of endothelial and epithelial cells.
  
  
  CONCLUSIONS
  Serum-free medium, Human Endothelial-SFM, promoted cell survival during corneal organ culture better than MEM+2% FCS. HES 130/0.4 appeared to be tolerated better by the cells than dextran T500.
authors.affiliation: Medical Faculty, Institute of Anatomy, TU Dresden, Dresden, Germany.
authors.forename: Lilla
authors.initials: L
authors.lastname: Knels
authors.affiliation: Medical Faculty, Institute of Anatomy, TU Dresden, Dresden, Germany.
authors.forename: Doris
authors.initials: D
authors.lastname: Goetze
authors.affiliation: CRTD/DFG-Center for Regenerative Therapies Dresden-Cluster of Excellence, TU Dresden, Dresden, Germany Department of Ophthalmology, Klinikum Chemnitz gGmbH, Chemnitz, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Engelmann
authors.affiliation: Medical Faculty, Institute of Anatomy, TU Dresden, Dresden, Germany.
authors.forename: Monika
authors.initials: M
authors.lastname: Valtink
mesh_terms: D000818:Animals; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003315:Cornea / Q000166:cytology* / Q000378:metabolism; D003470:Culture Media / Q000633:toxicity; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D003911:Dextrans / Q000633:toxicity*; D004728:Endothelium, Corneal / Q000166:cytology / Q000378:metabolism; D019573:Epithelium, Corneal / Q000166:cytology / Q000378:metabolism; D005260:Female; D006895:Hydroxyethyl Starch Derivatives / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009924:Organ Culture Techniques; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism
chemical_list: D003470:Culture Media; D016895:Culture Media, Serum-Free; D003911:Dextrans; D006895:Hydroxyethyl Starch Derivatives; D019253:Proto-Oncogene Proteins c-bcl-2; D053148:Caspase 3
keywords: Apotosis; Cornea; Eye (Tissue) Banking