2,5-hexanedione Induced Apoptosis In Nerve Tissue Of Rats Via PI3K/Akt Signaling Pathway

Objective In this study, we will observe HD induces neural cell apoptosis via PI3K/Akt signaling pathway.Materials and Methods Forty adult male SD rats, weighing 180-220 g, were divided randomly into control and three experimental groups(100 mg/kg, 200 mg/kg and 400 mg/kg), each of 10. The rats in control group were treated with saline and experimental groups were treated with 2,5-HD at a dosage of 100, 200 and 400 mg/kg/day for 5 weeks by intraperitoneal injection. Injection volume was 0.3 ml/100 g. After 5 week exposure, spinal cord and sciatic nerve were extracted. Apoptosis percentage in nerve tissues were estimated by Hoechst 33342 staining and TUNEL assay. The expression of Akt, p-Akt, Bad, p-Bad and cytochrome c was analyzed by western blot. Binding of Bad with Bcl-x L in the mitochondrial fraction was examined by coimmunoprecipitation method. Moreover, the activity of caspase-3 was also measured by microplate reader. The experimental results were analyzed by using SPSS 13.0 statistical package.Results The apoptosis index in spinal cord was 5.95, 7.84 and 10.14% respectively in rats exposed to 100, 200 and 400 mg/kg HD, significantly higher than 0.11% in control group(p < 0.05). In sciatic nerve, the TUNEL-positive neuronal cells were found in experimental groups, whereas the TUNEL-positive cells were almost absent in controlgroup, the apoptosis index was 5.14%, 9.23% and 12.75% respectively in rats exposed to 100 mg/kg, 200 mg/kg and 400 mg/kg HD, significantly higher than 0.13% in control group(p < 0.05). The expression levels of Akt and Bad in spinal cord and sciatic nerve of rats showed no significant changes in experimental groups compared with controls(p > 0.05). However, the amount of phosphorylated Akt and Bad in spinal cord and sciatic nerve was significantly lower in rats exposed to HD than that in control group(p < 0.05) and decreased in a dose-dependent manner. Bad/Bcl-x L dimerization was significantly increased in the mitochondrial fraction(p < 0.05). The intensity of cytochrome c band in the mitochondrion fraction was significantly lower in spinal cord and sciatic nerve of the rats treated with HD than that in control group(p < 0.05). On the other hand, the amount of cytochrome c protein in cytosol was significantly higher in spinal cord and sciatic nerve of the rats treated with HD than that in control group(p < 0.05). The activity of caspase-3 in the spinal cord and sciatic nerve was significantly higher in the groups exposed to HD than that in control group(p < 0.05). Especially, the caspase-3 activity was the highest in group exposed to 400 mg/kg HD among the four groups.Conclusion HD exposure can induce apoptosis in rat spinal cord and sciatic nerve, inhibited the phosphorylation of Akt and Bad, increased the dimerization of Bad with Bcl-x L in the mitochondrial fraction, followed by the release of cytochrome c and activation of caspase-3 in spinal cord and sciatic nerve of the rats.Our results indicate that HD induces neural cell apoptosis via PI3K/Akt signaling pathway.