title: Rhein Elicits In Vitro Cytotoxicity in Primary Human Liver HL-7702 Cells by Inducing Apoptosis through Mitochondria-Mediated Pathway.
abstract: Objective. To study rhein-induced apoptosis signaling pathway and to investigate its molecular mechanisms in primary human hepatic cells. Results. Cell viability of HL-7702 cells treated with rhein showed significant decrease in dose-dependent manner. Following rhein treatment (25 μM, 50 μM, and 100 μM) for 12 h, the detection of apoptotic cells was significantly analyzed by flow cytometry and nuclear morphological changes by Hoechst 33258, respectively. Fatty degeneration studies showed upregulation level of the relevant hepatic markers (P < 0.01). Caspase activities expressed significant upregulation of caspase-3, caspase-9, and caspase-8. Moreover, apoptotic cells by rhein were significantly inhibited by Z-LEHD-FMK and Z-DEVD-FMK, caspase-9 inhibitor, and caspase-3 inhibitor, respectively. Overproduction of reactive oxygen species, lipid peroxidation, and loss of mitochondrial membrane potential were detected by fluorometry. Additionally, NAC, a ROS scavenger, significantly attenuated rhein-induced oxidative damage in HL-7702 cells. Furthermore, real-time qPCR results showed significant upregulation of p53, PUMA, Apaf-1, and Casp-9 and Casp-3 mRNA, with no significant changes of Fas and Cytochrome-c. Immunoblotting revealed significant Cytochrome-c release from mitochondria into cytosol and no change in Fas expression. Conclusion. Taken together, these observations suggested that rhein could induce apoptosis in HL-7702 cells via mitochondria-mediated signal pathway with involvement of oxidative stress mechanism.
authors.affiliation: Department of Clinical Pharmacy, China Pharmaceutical University, No. 24, Tong Jia Xiang, Jiangsu, Nanjing 210009, China.
authors.forename: Guy-Armel
authors.initials: GA
authors.lastname: Bounda
authors.affiliation: Department of Pharmacology, China Pharmaceutical University, No. 24, Tong Jia Xiang, Jiangsu, Nanjing 210009, China.
authors.forename: Wang
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Department of Pharmacology, China Pharmaceutical University, No. 24, Tong Jia Xiang, Jiangsu, Nanjing 210009, China.
authors.forename: Dan-Dan
authors.initials: DD
authors.lastname: Wang
authors.affiliation: Department of Clinical Pharmacy, China Pharmaceutical University, No. 24, Tong Jia Xiang, Jiangsu, Nanjing 210009, China ; Department of Pharmacology, China Pharmaceutical University, No. 24, Tong Jia Xiang, Jiangsu, Nanjing 210009, China ; Key Laboratory of Drug Quality Control and Pharmacovigilance, China Pharmaceutical University, Ministry of Education, Nanjing 210009, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Yu