PMID: 9191602Jun 1, 1997Paper

Induction of apoptosis in cultured human retinal pigment epithelial cells is counteracted by flupirtine

Investigative Ophthalmology & Visual Science
N N OsborneJ P Wood

Abstract

The aim of the study was to determine whether flupirtine can counteract the induction of apoptosis in cultured retinal pigment epithelium (RPE) cells. Confluent cultures were subjected to experimental ischemia (medium free of serum, glucose, and oxygen) with or without various substances for specific periods. The cells were then examined for breakdown of DNA by the TUNEL procedure and agarose gel electrophoresis. Moreover cells were processed for the localization of oncogene proteins (bcl-2, TIAR, ICH-1t) associated with apoptosis. The effect of flupirtine on reactive oxygen species also was determined. When RPE cells were subjected to ischemia for 72 hours approximately 65% of cells remained attached to the coverslips and approximately 65% of their nuclei showed clear fragmentation of DNA by TUNEL. Most of the cells exhibited a shrunken appearance typical of apoptosis. Fragmentation of the DNA from cells given ischemia for 72 hours was also confirmed by agarose gel electrophoresis. Inclusion of flupirtine (flupirtine gluconate, 100 microM) or 10% fetal calf serum in the medium prevented ischemia-induced apoptosis occurring after 72 hours. Neither N-methyl-D-aspartate (NMDA) (100 microM) nondeferoxamine (100 microM) nor the NMD...Continue Reading

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