Oxidative stress gradient in a medium during human corneal organ culture.
OBJECTIVE
Lipid peroxidation content was measured in an organ culture medium after one-week storage of human donor corneas. Moreover, the effects of the medium on oxidative stress, antioxidant capacity, and the proliferation of cultured human corneal cells were studied.


METHODS
The medium was sampled from the upper and lower halves of storage vials and from controls (n=42). Malondialdehyde (MDA) was measured by high pressure liquid chromatography (HPLC). Cultured human corneal epithelium (CRL-11515) was exposed to different medium samples and monitored for changes in MDA (enzyme-linked immunosorbent assay [ELISA]), total antioxidant capacity (antioxidant assay kit), and proliferation (Ki-67).


RESULTS
A significant increase in MDA was observed in the organ culture medium in the lower level of storage vials. The addition of this fraction to cultured cells increased MDA significantly after 3 days, and the medium from both levels significantly increased MDA after 7 days. The medium from both levels significantly decreased the total antioxidant capacity of the cells but did not affect proliferative activity.


CONCLUSIONS
An oxidative gradient with an evident biologic effect is established in the medium in vials during organ culture of human donor corneas. Donor tissue stored at the bottom or in lower levels of such vials is exposed to a significant amount of oxidative stress.
Fundación Oftalmológica del Mediterráneo, Valencia, Spain. johnsen_siv@gva.es
Siv
S
Johnsen-Soriano
Kristiane
K
Haug
Emma
E
Arnal
Cristina
C
Peris-Martinez
Morten C
MC
Moe
Francisco Javier
FJ
Romero
Bjørn
B
Nicolaissen
D000906:Antibodies / Q000494:pharmacology; D000975:Antioxidants / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D017077:Culture Media, Conditioned / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D019394:Ki-67 Antigen / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D008315:Malondialdehyde / Q000032:analysis; D018384:Oxidative Stress / Q000187:drug effects; D014015:Tissue Banks; D046509:Tissue Culture Techniques
D000906:Antibodies; D000975:Antioxidants; D017077:Culture Media, Conditioned; D019394:Ki-67 Antigen; D008315:Malondialdehyde