Department of Pharmacology, Medical School of Southeast University, Southeast University, Nanjing, 210009, Jiangsu, China.
Department of Physiology, Medical School of Southeast University, Southeast University, Nanjing, China.
Metab Brain Dis. 2019 Feb;34(1):61-69. doi: 10.1007/s11011-018-0319-y. Epub 2018 Sep 26.
In this study, we demonstrated that PUMA was involved in the microglial migration induced by methamphetamine. PUMA expression was examined by western blotting and immunofluorescence staining. BV2 and HAPI cells were pretreated with a sigma-1R antagonist and extracellular signal-regulated kinase (ERK), mitogen-activated protein kinase (MAPK), c-Jun N-terminal protein kinase (JNK), and phosphatidylinositol-3 kinase (PI3K)/Akt inhibitors, and PUMA expression was detected by western blotting. The cell migration in BV2 and HAPI cells transfected with a lentivirus encoding red fluorescent protein (LV-RFP) was also examined using a wound-healing assay and nested matrix model and cell migration assay respectively. The molecular mechanisms of PUMA in microglial migration were validated using a siRNA approach. The exposure of BV2 and HAPI cells to methamphetamine increased the expression of PUMA, reactive oxygen species (ROS), the MAPK and PI3K/Akt pathways and the downstream transcription factor signal transducer and activator of transcription 3 (STAT3) pathways. PUMA knockdown in microglia transfected with PUMA siRNA attenuated the increased cell migration induced by methamphetamine, thereby implicating PUMA in the migration of BV2 and HAPI cells. This study demonstrated that methamphetamine-induced microglial migration involved PUMA up-regulation. Targeting PUMA could provide insights into the development of a potential therapeutic approach for the alleviation of microglia migration induced by methamphetamine.
在这项研究中，我们证明了 PUMA 参与了甲基苯丙胺诱导的小胶质细胞迁移。通过 Western blot 和免疫荧光染色检测 PUMA 的表达。用 sigma-1R 拮抗剂和细胞外信号调节激酶（ERK）、丝裂原活化蛋白激酶（MAPK）、c-Jun N 端蛋白激酶（JNK）和磷脂酰肌醇-3 激酶（PI3K）/Akt 抑制剂预处理 BV2 和 HAPI 细胞，并通过 Western blot 检测 PUMA 的表达。通过划痕愈合试验、嵌套基质模型和细胞迁移试验分别检测转染红色荧光蛋白（LV-RFP）的 BV2 和 HAPI 细胞的细胞迁移。使用 siRNA 方法验证了 PUMA 在小胶质细胞迁移中的分子机制。BV2 和 HAPI 细胞暴露于甲基苯丙胺增加了 PUMA、活性氧（ROS）、MAPK 和 PI3K/Akt 途径以及下游转录因子信号转导和转录激活因子 3（STAT3）途径的表达。用 PUMA siRNA 转染的小胶质细胞中的 PUMA 敲低减弱了甲基苯丙胺诱导的细胞迁移增加，从而表明 PUMA 参与了 BV2 和 HAPI 细胞的迁移。这项研究表明，甲基苯丙胺诱导的小胶质细胞迁移涉及 PUMA 的上调。针对 PUMA 可能为缓解甲基苯丙胺诱导的小胶质细胞迁移提供一种潜在的治疗方法提供了新的思路。
