Anti-apoptotic effect of fermentedCitrus sunkipeel extract on chemical hypoxia-induced neuronal injury

Purpose: Neuronal apoptotic events induced by aging and hypoxic/ischemic conditions is an important risk factor in neurodegenerative diseases such as ischemia stroke and Alzheimers disease. The peel of Citrus sunki Hort. ex Tanaka has long been used as a traditional medicine, based on multiple biological activities including anti-oxidant, anti-inflammation, and anti-obesity. In the current study, we examined the actions of fermented C. sunki peel extract against cobalt chloride (CoCl₂)-mediated hypoxic death in human neuroblastoma SH-SY5Y cells. Methods: Cell viability was measured by trypan blue exclusion. Expression of apoptosis related proteins and release of cytochrome c were detected by western blot. Production of intracellular reactive oxygen species (ROS) and apoptotic morphology were examined using 2,7- dichlorofluorescin diacetate (DCF-DA) and 4,6-diamidino-2-phenylindole (DAPI) staining. Results: Exposure to CoCl₂, a well-known mimetic agent of hypoxic/ischemic condition, resulted in neuronal cell death via caspase-3 dependent pathway. Extract of fermented C. sunki peel significantly rescued the CoCl₂-induced neuronal toxicity with the cell viability and appearance of apoptotic morphology. Cytoprotection with fermented C. sunki peel extract was associated with a decrease in activities of caspase-3 and cleavage of poly (ADP ribose) polymerase (PARP). In addition, increase in the intracellular ROS and release of cytochrome c from mitochondria to the cytosol were inhibited by treatment with extract of fermented C. sunki peel. Conclusion: Based on these data, fermented C. sunki peel extract might have a protective effect against CoCl₂-induced neuronal injury partly through generation of ROS and effectors involved in mitochondrial mediated apoptosis.