The Mechanism of Midazolam and Propofol on Mouse Leydig Cell Lines Apoptosis

• Main Authors: Shu-ChunWang, 王姝淳
• Other Authors: Bu-Miin Huang
• Format: Others
• Language: en_US
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/rn2f6z

碩士 === 國立成功大學 === 細胞生物與解剖學研究所 === 103 === The incident rate of the testicular cancer is increasing in western world, and the overall incidence rate of testicular cancer rose over time, from 3.7 (per 100,000) in 1975 to 5.9 (per 100,000) in 2007. The treatment upon malignant testicular cancer is usually combined surgery with chemical drugs. Midazolam and propofol are widely used as sedative and anesthetic induction agents by modulating the different GABA receptors in the central nervous system. In previous study, we have found that midazolam could induce MA-10 mouse Leydig tumor cell apoptosis by activating caspase cascade. In addition, study has shown that propofol could induce apoptosis in lung cancer through endoplasmic reticulum (ER) stress. However, the detail mechanisms how midazolam and propofol regulating caspase and/or ER stress pathways in Leydig tumor cells remain elusive. In the present study, MA-10 cells and TM3 cells (a mouse Leydig normal cell line) were used with the treatments of midazolam or propofol. Results showed that cell viability significantly decreased by midazolam from 30 to 150 µM in MA-10 cells and from 150 to 300 µM in TM3 cells for 12 hr, respectively, in a dose-dependent manner (p〈0.05). Cell viability of MA-10 and TM3 cells also significantly decreased as the dosage of propofol increased (300 to 600 μM) for 24 hr, respectively (p〈0.05). In flow cytometry analysis, both midazolam and propofol significantly increased the amounts of subG1 phase cell numbers in both cell lines (p〈0.05). AnnexinV/PI double staining further confirmed that two sedative agents induced apoptosis in both cell lines. Moreover, cleaved caspase-8, -9, -3 and/or PARP were significantly activated after treatment of midazolam and propofol in both cell lines. Besides, Bax translocation and cytochrome C release were also involved in midazolam-induced MA-10 cell apoptosis. In TM3 cell line, the phosphorylation of JNK, ERK1/2 and p38 were elevated by midazolam treatment. On the other hand, the expression of p-JNK, p-ERK1/2 and p-p38 were activated by propofol treatment in both cell lines, which indicated that two sedative agents could induce apoptosis through MAPK pathway. In addition, propofol diminished the phosphorylation of Akt to induce apoptosis in both cell lines. Furthermore, the expression of p-EIF2α, ATF4, ATF3 and CHOP could be induced by midazolam in both cell lines, which appeared that midazolam could induce apoptosis probably through endoplasmic reticulum (ER) stress. The expressions of cyclin A, cyclin B and CDK1 could be inhibited by midazolam through the regulation of p53 in MA-10 and TM3 cells, suggesting midazolam could regulate cell cycle to induce apoptosis in mouse Leydig cell lines. In conclusion, midazolam and propofol could induce cell apoptosis by activating caspases and MAPKs pathways, and inhibiting Akt pathway in mouse Leydig cell lines. In addition, midazolam could induce cell apoptosis through activation of ER stress and regulation of cell cycle, and midazolam might be a better anticancer therapy than propofol in Leydig cancer.