Time course of ischemia/reperfusion-induced oxidative modification of neural proteins in rat forebrain.

Time course of oxidative modification of forebrain neural proteins was investigated in the rat model of global and partial cerebral ischemia/reperfusion. Animals were subjected to 4-vessel occlusion for 15 min (global ischemia). After the end of ischemia and at different reperfusion times (2, 24 and 48 h), lipoperoxidation-dependent and direct oxidative modification neural protein markers were measured in the forebrain total membrane fraction (tissue homogenate). Ischemia itself causes significant changes only in levels of tryptophan and bityrosine fluorescence when compared to controls. All tested parameters of protein modification altered significantly and were maximal at later reperfusion stage. Content of carbonyl group in re-flow period steadily increased and culminated at 48 h of reperfusion. The highest increase in the fluorescence of bityrosines was detected after 24 h of reperfusion and was statistically significant to both sham operated and ischemic groups. The changes in fluorescence intensity of tryptophan decreased during a reperfusion time dependent manner. Formation of lysine conjugates with lipoperoxidation end-products significantly increased only at later stages of reperfusion. Total forebrain membranes from animals subjected to 3-vessel occlusion model to 15 min (partial ischemia) show no altered content of oxidatively modified proteins compared to controls. Restoration of blood flow for 24 h significantly decreased only fluorescence of aromatic tryptophan. Partial forebrain ischemia/reperfusion resulted in no detectable significant changes in oxidative products formation in extracerebral tissues (liver and kidney) homogenates. Our results suggest that global ischemia/reperfusion initiates both the lipoperoxidation-dependent and direct oxidative modifications of neural proteins. The findings support the view that spatial and temporal injury at later stages of ischemic insult at least partially involves oxidative stress-induced amino acid modification. The results might have important implications for the prospective post-ischemic antioxidant therapy.

Impact of Ginkgo Biloba Extract EGb 761 on ischemia/reperfusion - induced oxidative stress products formation in rat forebrain.

Dysbalance in reactive oxygen/nitrogen species is involved in the pathogenesis of cerebral ischemia/reperfusion injury (IRI). Ginkgo biloba extract (Egb 761) pre-treatment was used to observe potential antioxidant/neuroprotective effect after global ischemia/reperfusion. Egb 761 significantly decreased the level of lipoperoxidation (LPO) in rat forebrain total membrane fraction (homogenate) induced by in vitro oxidative stress (Fe(2+)+H(2)O(2)). In animals subjected to four-vessel global ischemia for 15 min and 2-24 h reperfusion the EGb pretreatment slightly decreased LPO in forebrain homogenate. However, as detected in EGb treated group, the LPO-induced lysine conjugates are attenuated in comparison to non-treated IRI animals. EGb significantly improved parameters which indicate forebrain protein oxidative damage after IRI. The intensity of tryptophane fluorescence was increased by the 18.2% comparing to non-treated IRI group and bityrosine fluorescence was significantly decreased in ischemic (21%) and 24 h reperfused (15.9%) group in comparison non-treated IRI group. In addition, the level of total free SH- groups in pre-treated animals was significantly higher comparing to non-treated animals. Our results indicate that extract of EGb 761 has potent antioxidant activity and could play a role to attenuate the IRI-induced oxidative protein modification and lipoperoxidation in the neuroprotective process.