ABSTRACT
After discussing the reason for insufficiency of committee member in current ethical review , and considering the requirement for improving the function of Ethics Review Committee , this paper proposes that Ethics Review Committee should appoint more non -affiliated scientific members to ensure the independence of ethical re-view , thus protecting the rights and beneficence of subjects , and to secure the justice in ethical review of protocols , the paper also discussed the resolutions to address the challenge brought by appointing non -affiliated members .
ABSTRACT
AIM: To investigate the molecular mechanisms of apoptosis and to elucidate the apoptosis signaling pathway triggered by etoposide in Jurkat human leukemia cells. METHODS: Apoptosis was detected using annexin V - FITC and propidium iodide (PI) staining, respectively, and annexin V - FITC positive cells and hypodiploid cells were analyzed by flow cytometry. Mitochondrial membrane potential (△Ψm) was detected using 3,3' - dihexyloxycarbocyanine iodide [ DiOC6 (3)] staining and △Ψm low cells were analyzed by flow cytometry. Preparation of cytosolic extracts and isolation of mitochondria were completed by centrifugation. Western blotting analysis was used to evaluate the level of cytochrome c, caspase - 3, and poly ( ADP - ribose) polymerase (PARP) expression. RESULTS: Etoposide induced apoptosis showing phosphatidylserine externalization and DNA fragmentation in a time - dependent manner and the apoptosis could be inhibited by a broad caspase inhibitor benzyloxycarbonyl - Val - Ala - Asp - fluoromethylketone ( zVAD. fmk). Collapse of △Ψm induced by etoposide preceded DNA fragmentation and phosphatidylserine externalization. In contrast, it was not blocked by zVAD. fmk. Etoposide caused cytochrome c release from mitochondria into cytosol, subsequent activation of caspase-3 (32 kD) presented with an intermediate (20 kD) and its active product (17kD), and cleavage of full- length PARP (116 kD) into the so- called apoptotic 85 kD fragment. CONCLUSION:Etoposide - induced Jurkat cell apoptosis is initiated through mitochondria signaling pathway with cytochrome c release into cytoplasm and caspase is the ultimate executioner of cell apoptosis.