Y. Liang et al., Bcl-2, Bax, and c-Fos expression correlates to RPE cell apoptosis induced by UV-light and daunorubicin, CURR EYE R, 20(1), 2000, pp. 25-34
Purpose. The aim of this study was to determine the role of Bcl-2, Bcl-X-L,
Bax, and c-Fos in regulation of apoptosis. induced by ultraviolet-light A
(UV-A) and daunorubicin (DNR), in retinal pigment epithelium (RPE) cells gr
own on bovine extracellular matrix (ECM)-coated or uncoated plastic dishes.
Methods. Apoptosis in confluent RPE cells cultured on ECM-coated or uncoate
d dishes was induced by UV-A or DNR. Apoptosis was detected by 7-amino-acti
nomycin D labeling followed by flow cytometry and by terminal deoxy-transfe
rase mediated X-dUTP nick end labeling (TUNEL). Cellular expression of Bcl-
2, Bcl-X-L, Bax and c-Fos was determined by the use of antibodies and flow
cytometry, Western blot analysis, and immunocytochemical staining.
Results. Both UV-A and DNR induce apoptosis in human RPE cells in vitro. Hu
man fetal RPE cells grown on ECM-coated dishes were significantly more resi
stant to UV-A or DNR induced apoptosis than cells grown on uncoated dishes.
RPE cells grown on ECM-coated dishes expressed higher Bcl-2 levels and low
er Bax levels compared to cells grown on uncoated dishes. However, Bcl-X-L
and c-Fos levels were comparable in the two cultures. After UV-A or DNR tre
atment, Bcl-2, Bcl-X-L, Bax, and c-Fos levels were differently regulated in
cells grown on ECM-coated dishes compared to cells grown on uncoated dishe
s.
Conclusion. A significant protection against apoptosis of RPE cells grown o
n ECM compared to cells grown on uncoated plastic dishes was found after ex
posure to W-A or DNR.
This protection was found to be proportionally correlated to the anti-apopt
otic protein Bcl-2 and inversely correlated to the expression of Bax. Furth
ermore a sustained induction and expression of c-Fos was found to correlate
to a higher percentage of apoptotic cells of RPE cells grown on plastic. T
hese findings demonstrate that ECM is of great importance for RPE cell surv
ival during noxious stimuli and points out the essential role for a healthy
Bruch's Membrane (BM) for RPE survival.