Rotenone-induced death of RGC-5 cells is caspase independent, involves the JNK and p38 pathways and is attenuated by specific green tea flavonoids.

The aim of the present studies was to characterise cell death following inhibition of mitochondrial complex I with rotenone in a transformed cell line (RGC-5 cells) and to examine the neuroprotective properties of the flavonoids genistein, epigallocatechin gallate (EGCG), epicatechin (EC) and baicalin. Rotenone-induced cell death of RGC-5 cells results in a generation of reactive oxygen species, a breakdown of DNA, the translocation of membrane phosphatidylserine, an up-regulation of haemoxygenase-1 and is unaffected by necrostatin-1 (inhibitor of necroptosis), z-VAD-fmk (pan caspase inhibitor) or NU1025 (PARP inhibitor) but attenuated with SP600125 (JNK inhibitor). Rotenone-induced toxicity of RGC-5 cells also caused an activation of mitogen-activated kinases indicated by an up-regulation and translocation into mitochondria of p-c-Jun, pJNK and pp38. Exposure of RGC-5 cells to rotenone does not affect apoptosis inducing factor or significantly stimulate caspase-3 activity. EGCG and EC both significantly blunt rotenone toxicity of RGC-5 cells at concentrations of 50 µM while genistein and baicalin were without effect. Significantly, genistein is approximately 20 times less efficacious than EGCG (IC(50) 2.5 µM) and EC (IC(50) 1.5 µM) at inhibiting sodium nitroprusside-induced lipid peroxidation. These studies show that rotenone toxicity of RGC-5 cells is neither necroptosis nor caspase-dependent apoptosis but involves the activation of mitogen-activated kinases and is inhibited by a JNK inhibitor, EGCG and EC. Genistein attenuates lipid peroxidation less efficaciously than EC and EGCG and does not affect rotenone toxicity of RGC-5 cells.

Genistein blunts the negative effect of ischaemia to the retina caused by an elevation of intraocular pressure.

AIMS: Deduce whether the isoflavone genistein blunts the effect of ischaemia to the retina. METHODS: Ischaemia was induced in rats by raising the intraocular pressure (120 mm Hg) for 50 min. Genistein (10 mg/kg) was injected intraperitoneally 1 h before and after ischaemia. Seven days after ischaemia, the level of mRNAs for neurofilament light (NF-L), caspase 3, caspase 8, glial fibrillary acidic protein (GFAP), poly-ADP ribose polymerase (PARP), Thy-1 and proteins (GFAP, NF-L, PARP) in whole retinas were determined. NF-L and tubulin proteins in optic nerves were also determined. Retinas were also processed for the localization of choline acetyltransferase (ChAT) and GFAP immunoreactivities. RESULTS: Ischaemia caused a significant reduction in ganglion cell proteins in the optic nerve (NF-L and tubulin) and retina (NF-L). Retinal Thy-1 (mRNA and protein) and NF-L (mRNA) were also reduced while mRNAs of caspase 3, caspase 8, PARP and GFAP (also protein) were increased. Changes in the mRNAs and proteins induced by ischaemia were significantly blunted by genistein with the exception of the increase in GFAP and PARP protein/mRNA levels. Ischaemia-induced changes in the localization of ChAT were also clearly attenuated by genistein treatment. CONCLUSIONS: Genistein blunts most of the damaging effects caused to the retina by ischaemia.