Inducible apoptosis-promoting activity in retinal cell-conditioned medium.

PURPOSE Apoptosis is implicated in the death of retinal cells during both retinal differentiation and degeneration. We sought to investigate potential diffusible retinal cell signalling factors which may be responsible for this phenomenon. METHODS 72 hr conditioned medium was collected from death-induced R28 retinal cells undergoing serum-starvation. This conditioned medium was filtered, titrated and added to fresh cultures of R28 cells for 24 hr. Cell death was measured by trypan blue exclusion; apoptosis was evaluated by TUNEL-in situ, DNA gel electrophoresis, and observations of cell morphology. RESULTS Apoptotic cell death was at least two-fold greater in retinal cultures which received death-induced cell-conditioned medium. This apoptotic activity in conditioned medium did not appear to be due to nutrient-depletion of the medium, as the control situation (72 hr conditioned medium from resistant, non-dying cells) did not produce the same effect. This diffusible apoptotic activity was heat-labile (56 degrees C for 45 min). CONCLUSIONS We have shown for the first time, in our system, that death-induced retinal cells release a diffusible, heat-labile apoptosis-promoting element. Further investigation will be necessary to determine whether this trigger of apoptosis is a toxic metabolite or an "apoptosis-promoting factor".