Tahvilian Reza, Amini Komail, Zhaleh Hossein
Pharmaceutical Sciences Research Center, School of Pharmacy, Kermanshah University of Medical Sciences, Kermanshah, Iran.
Department of Biology, Faculty of Sciences, Razi University, Kermanshah, Iran.
Asian Pac J Cancer Prev. 2019 Sep 1;20(9):2763-2774. doi: 10.31557/APJCP.2019.20.9.2763.
Objective: Interaction of methamphetamine and sigma (σ) receptors lead to up-regulation and activation of these receptors. The σ receptors induced apoptosis in some parts of the brain by increasing calcium, dopamine, ROS, mitochondrial pores and caspase activity. Ibudilast is a phosphodiesterase inhibitor and anti-inflammatory drug, which can decrease the inflammatory cytokines. Also, it has a neuroprotective effect. It seems that ibudilast can reduce the methamphetamine-induced cell death due to inhibition of σ receptors. Materials and Methods: There were seven treatments including; control: culture medium, Treatment 1: 1mM methamphetamine, Treatment 2: 1mM methamphetamine and 1nM ibudilast, Treatment 3: 1mM methamphetamine and 10nM ibudilast, Treatment 4: 1mM methamphetamine and 100nM ibudilast, Treatment 5: 1mM methamphetamine and 1uM ibudilast, Treatment 6: 1mM methamphetamine and 10uM ibudilast, and Treatment 7: 1mM methamphetamine and 100uM ibudilast. Finally, for inhibition of PKA, CREB, IP3 receptor, NMDA receptor, Sigma receptor antagonist, sigma receptor agonist, cells were preincubated with adding H89 dihydrochloride, 666-15, Heparin, Ketamine, BMY 14802, and Pentazocine. MTT and LDH tests were performed for cell viability and cytotoxicity measurement, respectively. In continuing, the caspase activity colorimetric assay kit used for caspase 3 activity diagnosis. Rhodamine-123 performed to detection of mitochondrial membrane potential. TUNEL test used to DNA fragmentation and apoptosis, Fura-2 used to Measurement of (Ca2+) ic and (Ca2+) m, and fluorescence microscope used to Measurement of antioxidant enzyme activities. Results: Ibudilast increased the cell viability and the rhodamine-123 absorbance in methamphetamin-treated PC12 cells. It reduced cell cytotoxicity, caspase 3 activity, ic and m Ca2+ concentration, (OH) generation and DNA fragmentation in all concentrations of 1 nM t0 100 μM (p<0.05) by the optimal concentration of 100 μM, between our tested treatments. Conclusion: Ibudilast as a phosphodiesterase inhibitor can reduce the methamphetamine-induced cell death due to inhibition of σ receptors through cAMP production.
甲基苯丙胺与σ受体相互作用会导致这些受体上调和激活。σ受体通过增加钙、多巴胺、活性氧、线粒体孔和半胱天冬酶活性，在大脑某些部位诱导细胞凋亡。异丁司特是一种磷酸二酯酶抑制剂和抗炎药，可减少炎性细胞因子。此外，它还具有神经保护作用。似乎异丁司特可通过抑制σ受体来减少甲基苯丙胺诱导的细胞死亡。材料与方法：有七种处理，包括：对照：培养基；处理1：1mM甲基苯丙胺；处理2：1mM甲基苯丙胺和1nM异丁司特；处理3：1mM甲基苯丙胺和10nM异丁司特；处理4：1mM甲基苯丙胺和100nM异丁司特；处理5：1mM甲基苯丙胺和1μM异丁司特；处理6：1mM甲基苯丙胺和10μM异丁司特；处理7：1mM甲基苯丙胺和100μM异丁司特。最后，为抑制蛋白激酶A、环磷腺苷反应元件结合蛋白、三磷酸肌醇受体、N-甲基-D-天冬氨酸受体、σ受体拮抗剂、σ受体激动剂，细胞在添加盐酸H89、666-15、肝素、氯胺酮、BMY 14802和喷他佐辛的情况下进行预孵育。分别进行MTT和LDH试验以测量细胞活力和细胞毒性。接着，使用半胱天冬酶活性比色测定试剂盒诊断半胱天冬酶3活性。使用罗丹明-123检测线粒体膜电位。使用TUNEL试验检测DNA片段化和细胞凋亡，使用Fura-2测量细胞内钙离子浓度([Ca2+]ic)和线粒体钙离子浓度([Ca2+]m)，并使用荧光显微镜测量抗氧化酶活性。结果：在甲基苯丙胺处理的PC12细胞中，异丁司特增加了细胞活力和罗丹明-123吸光度。在我们测试的处理中，它在1 nM至100 μM的所有浓度下均降低了细胞毒性、半胱天冬酶3活性、[Ca2+]ic和[Ca2+]m浓度、(OH)生成和DNA片段化（p<0.05），最佳浓度为100 μM。结论：异丁司特作为一种磷酸二酯酶抑制剂，可通过抑制σ受体，减少甲基苯丙胺诱导的细胞死亡。
