Imbalance between pro-apoptotic and pro-survival factors in human retinal pericytes in diabetic-like conditions

Abstract

Purpose: Loss of pericytes is one the key events in the pathogenesis of diabetic retinopathy. We have previously demonstrated that human retinal pericytes (HRP) are more vulnerable to intermittent than stable high glucose concentrations, with an increase in apoptosis. Our aim was to explore the expression of molecules involved in pro-apoptotic and survival pathways in pericytes cultured in stable/intermittent high glucose and/or hypoxia, to clarify the mechanisms of action of these diabetic-like stressing stimuli.

Methods: Human retinal pericytes (HRP) were exposed intermittently at 48-hr intervals to high/physiological glucose for 8 days (intHG) and/or hypoxia over the last 48 hr. Control cells were kept in stable physiological and high glucose. Cell proliferation and apoptosis were assessed. The expression of pro-apoptotic and pro-survival molecules was evaluated by Western blotting. Caspase-8 translocation from the cytoplasm into the nucleus was checked by Western blotting of nuclear versus cytoplasmic fractions and immunofluorescence.

Results: Hypoxia, alone and combined with intHG, increased HRP apoptosis and decreased proliferation. Pro-apoptotic molecules increased in HRP cultured in these conditions, while some survival markers decreased. Conversely, in stable HG, pro-apoptotic molecules were stable or even decreased, and survival factors increased. Translocation of caspase-8 from cytoplasm into nucleus indicates a primary role for this molecule in inducing apoptosis.

Conclusion: Diabetic-like conditions are able to stimulate pericyte apoptosis through activation of pro-apoptotic molecules, leading to an imbalance between pro-apoptotic and survival signalling pathways, with caspase-8 playing a pivotal role. Our identification of such intermediates could help finding new therapeutic approaches for the prevention of diabetic retinopathy.

Keywords: apoptosis; apoptotic pathways; caspase-8; diabetic retinopathy; pericyte(s).