Assessment of attachment factors for primary cultured human corneal endothelial cells.
OBJECTIVE
To assess the ability of various attachment factors to promote attachment of primary cultured human corneal endothelial cells.


METHODS
Primary cultured human corneal endothelial cells (HCEC) were incubated for 2 hours in 24-well plates. Wells had been precoated with commercially available cell attachment improvement media (FNC coating mix), human collagen I, human fibronectin, fibronectin/collagen I, or poly-d-lysine. Ratios of cell count before and after rinsing with culture medium and ratios of cells showing morphological signs of spreading to total cells were calculated to measure effectiveness of attachment factors.


RESULTS
Incubation of HCEC for 2 hours in wells without precoating of attachment factors led to a rate of 41 +/- 16% (mean +/- SD) of cells showing signs of spreading. FNC coating mix, collagen I, and fibronectin/collagen I increased significantly the percentage of cells showing morphological features of attachment at 2 hours. Total cell loss was highest with poly-d-lysine and no pretreatment with attachment factor. Without the use of any attachment factor, 67 +/- 19% of cells remained after rinsing. The lowest cell loss was observed with FNC coating mix where 108 +/- 5% of cells remained after rinsing.


CONCLUSIONS
Collagen I, collagen I/fibronectin, and FNC coating mix significantly enhance the spreading of human corneal endothelial cells to tissue culture plates after 2 hours. FNC coating mix significantly reduces cell loss due to rinsing. Without the use of any attachment factor, 67% of the cells remained in situ after rinsing.
Engler
Christoph
C
Wilmer Eye Institute, The Johns Hopkins Medical Institutions, Baltimore, Maryland, USA.
Kelliher
Clare
C
Speck
Caroline L
CL
Jun
Albert S
AS
D002448:Cell Adhesion / Q000187:drug effects*; D002452:Cell Count; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D024042:Collagen Type I / Q000494:pharmacology*; D004338:Drug Combinations; D004728:Endothelium, Corneal / Q000166:cytology / Q000502:physiology*; D005353:Fibronectins / Q000494:pharmacology*; D006801:Humans; D008239:Lysine / Q000494:pharmacology
D024042:Collagen Type I; D004338:Drug Combinations; D005353:Fibronectins; D008239:Lysine