The effect of air, SF6 and C3F8 on immortalized human corneal endothelial cells.
OBJECTIVE
While anterior chamber air bubbles aid attachment during posterior lamellar surgery only for few days, these periods can be prolonged with gases in non-expanding concentrations. To test the effects of different gas compositions on immortalized human corneal endothelial cells (HCEC-12), we utilized Transwell inserts with semipermeable membranes as an artificial anterior chamber model.


METHODS
Human corneal endothelial cells (HCEC-12) were cultured on Transwell inserts for 24 hr, then flipped, burdened and sunk with titanium rings in medium (M1), as well as filled with 2 ml of air (A), 20% sulphur hexafluoride (SF6) (S), or 12% C3F8 (C). After gas exposition for 24, 48 and 120 hr, cells were evaluated by live/dead staining, cell viability assay and Ki67 immunohistochemistry.


RESULTS
Proliferation was significantly reduced (Ki67-positive fraction; M1, 14.8 ± 2.0%; A, 7.9 ± 1.4%; S, 8.1 ± 1.3%; C, 9.9 ± 2.3%; p-values; A, S, C versus M1 < 0.01), the total cell number decreased and the percentage of dead cells increased under gas exposition, independently of the type of gas (120 hr cell count/2.25 cm2: M1 = 660.8 ± 57.0 cells; A = 125.5 ± 17.4 cells, S = 123.5 ± 17.0 cells, C = 118.8 ± 16.6 cells; p-value: M versus A/S/C < 0.001; 120 hr dead cells: M = 2.6 ± 1.0%, A = 8.4 ± 2.7%, S = 9.5 ± 3.2%, C = 11.3 ± 3.1%; p-value: M1 versus A/S/C < 0.01). Medium (M1)-control also proved significantly higher cell viability values in comparison with the gases, which did not differ significantly among them (120 hr luminescence: M1 = 1752.2 ± 91.4, A = 433.0 ± 30.3, S = 507.8 ± 23.3, C = 523.8 ± 20.3; p-value: M1 versus A/S/C < 0.01).


CONCLUSIONS
Gas exposition led to a reduction in proliferation and an increase in cell death in HCEC-12, independently of the gas composition.
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
Marina
M
Hesse
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
David
D
Kuerten
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
Peter
P
Walter
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
Niklas
N
Plange
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
Sandra
S
Johnen
Department of Ophthalmology, RWTH Aachen University, Aachen, Germany.
Matthias
M
Fuest
D000369:Aged, 80 and over; D000388:Air; D002452:Cell Count; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D055954:Corneal Endothelial Cell Loss / Q000473:pathology / Q000628:therapy*; D004728:Endothelium, Corneal / Q000187:drug effects* / Q000473:pathology; D005260:Female; D006801:Humans; D013459:Sulfur Hexafluoride / Q000008:administration & dosage*
D013459:Sulfur Hexafluoride
air; cornea; corneal endothelial cells; gas; keratoplasty