Abstract
Abstract: :
Purpose:The accumulation of advanced glycation endproducts (AGEs) during the Maillard reaction are known to be associated with the risk of diabetic neuropathy, diabetic retinopathy, age related macular degeneration and Mb. Alzheimer. Extracellular effects of the AGEs like crosslinking of proteins are well known, whereas the effects of AGEs on the cellular function are still unclear. To show reproducible damaging effects of glyoxal and methylglyoxal (reactive intermediates of Maillard reaction leading to AGEs) and eventual protective effects of dorzolamide, a blocker of carboanhydrase. Methods:The retinal cell line E1A–NR3 was used. Cells were triggered into apoptosis by 0.8mM concentrations of glyoxal and methylglyoxal (reactive intermediates of Maillard reaction). By flow cytometry a) fragmented DNA was detected by propidium iodide staining (sub–G1 peak) and by b) annexin V phosphatidylserine in the outer membrane and c) ROS production. pH–ratio–imaging was made by fluorescence microscopy. Results:Glyoxal as well as methylglyoxal caused apoptotic events in 60%. In both cases dorzolamide could reduce apoptosis by 50%. Both AGE intermediates reduced the intracellular pH by about 0.5 which could be ameliorated to a –0.3 – reduction after dorzolamide. The amount of increased ROS production (maximum 4 – 5 hours after adding the noxious agents) was not weakened substantially by dorzolamide. Conclusions:The results show that dorzolamide has a beneficial effect rather in the later stages of apoptosis with decreased pH than in the early events which are characterized increased ROS production.
Keywords: retinal culture • neuroprotection • oxidation/oxidative or free radical damage