title: Characterization of cryopreserved primary human corneal endothelial cells cultured in human serum-supplemented media.
abstract: To compare cryopreserved human corneal endothelial cells (HCECs) grown in human serum-supplemented media (HS-SM) with cryopreserved HCECs grown in fetal bovine serum-supplemented media (FBS-SM).
  
  
  
  Three pairs of human corneas from donors aged 8, 28, and 31 years were obtained from the eye bank. From each pair, one cornea was used to start a HCEC culture using HS-SM; the other cornea was grown in FBS-SM. On reaching confluence, the six cell populations were frozen using 10% dimethyl sulfoxidecontaining medium. Thawed cells grown in HS-SM were compared with those grown in FBS-SM with respect to morphology, growth curves, immunohistochemistry, real time-reverse transcriptase polymerase chain reaction (RT-PCR) for endothelial cell markers, and detachment time.
  
  
  
  No difference in morphology was observed for cells grown in the two media before or after cryopreservation. By growth curves, cell counts after thawing were similar in both media, with a slight trend toward higher cell counts in FBS-SM. Cells grown in both the media demonstrated a similar expression of endothelial cell markers when assessed by immunohistochemistry, although HCEC marker gene expression was higher in cells grown in HS-SM than in those grown in FBS-SM as assessed by RT-PCR. With FBS-SM, there was a tendency of longer detachment time and lower cell passages.
  
  
  
  HS-SM was similar to FBS-SM for cryopreservation of cultured HCECs as assessed by analysis of cell morphology, proliferation, and protein expression, although marker gene expression was higher in cells grown in HS-SM than in those grown in FBS-SM. Detachment time was longer with FBS-SM and in lower passages.
authors: Cornea and Anterior Segment Service, The Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
authors: Lucas Monferrari Monteiro
authors: LM
authors: Vianna
authors: Cornea and Anterior Segment Service, The Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
authors: Hao-Dong
authors: HD
authors: Li
authors: Lions VisionGift, Portland, OR, USA.
authors: Jeffrey D
authors: JD
authors: Holiman
authors: Lions VisionGift, Portland, OR, USA.
authors: Christopher
authors: C
authors: Stoeger
authors: Ophhtalmology and Vision Science Department, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brazil.
authors: Rubens
authors: R
authors: Belfort
authors: Cornea and Anterior Segment Service, The Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
authors: Albert S
authors: AS
authors: Jun
mesh_terms: D000328:Adult; D000818:Animals; D002417:Cattle; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects; D002648:Child; D015925:Cryopreservation / Q000379:methods*; D017077:Culture Media, Conditioned; D042783:Endothelial Cells / Q000166:cytology*; D004728:Endothelium, Corneal / Q000166:cytology*; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044967:Serum; D018709:Statistics, Nonparametric; D013997:Time Factors
chemical_list: D017077:Culture Media, Conditioned