Pan Tao, Zhu Qiu-Jiao, Xu Li-Xiao, Ding Xin, Li Jian-Qin, Sun Bin, Hua Jun, Feng Xing
Department of Neonatology, Children's Hospital Affiliated to Suzhou University, Suzhou, Jiangsu Province, China.
Department of Critical Care Medicine, Children's Hospital Affiliated to Suzhou University, Suzhou, Jiangsu Province, China.
Neural Regen Res. 2020 Nov;15(11):2154-2161. doi: 10.4103/1673-5374.282271.
Transient receptor potential melastatin 2 (TRPM2) is an important ion channel that represents a potential target for treating injury caused by cerebral ischemia. However, it is unclear whether reducing TRPM2 expression can help repair cerebral injury, and if so what the mechanism underlying this process involves. This study investigated the protective effect of reducing TRPM2 expression on pheochromocytoma (PC12) cells injured by oxygen-glucose deprivation (OGD). PC12 cells were transfected with plasmid encoding TRPM2 shRNAS, then subjected to OGD by incubation in glucose-free medium under hypoxic conditions for 8 hours, after which the cells were allowed to reoxygenate for 24 hours. Apoptotic cells, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels were detected using flow cytometry. The relative expression of C-X-C motif chemokine ligand 2 (CXCL2), NACHT, LRR, and PYD domain-containing protein 3 (NALP3), and caspase-1 were detected using fluorescence-based quantitative reverse transcription-polymerase chain reaction and western blotting. The rates of apoptosis, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels in the TRPM2-shRNA + OGD group were lower than those observed in the OGD group. Taken together, these results suggest that TRPM2 knockdown reduces OGD-induced neuronal injury, potentially by inhibiting apoptosis and reducing oxidative stress levels, mitochondrial membrane potentials, intracellular calcium concentrations, and NLRP3 inflammasome activation.
瞬时受体电位褪黑素2(TRPM2)是一种重要的离子通道,是治疗脑缺血损伤的潜在靶点。然而,尚不清楚降低TRPM2表达是否有助于修复脑损伤,如果是,这一过程的潜在机制是什么。本研究探讨了降低TRPM2表达对氧糖剥夺(OGD)损伤的嗜铬细胞瘤(PC12)细胞的保护作用。将编码TRPM2 shRNAs的质粒转染PC12细胞,然后在缺氧条件下于无葡萄糖培养基中孵育8小时进行OGD处理,之后使细胞复氧24小时。使用流式细胞术检测凋亡细胞、线粒体膜电位、活性氧水平和细胞钙水平。使用基于荧光的定量逆转录-聚合酶链反应和蛋白质印迹法检测C-X-C基序趋化因子配体2(CXCL2)、含NACHT、LRR和PYD结构域蛋白3(NALP3)以及半胱天冬酶-1的相对表达。TRPM2-shRNA + OGD组的凋亡率、线粒体膜电位、活性氧水平和细胞钙水平低于OGD组。综上所述,这些结果表明,敲低TRPM2可减轻OGD诱导的神经元损伤,可能是通过抑制凋亡以及降低氧化应激水平、线粒体膜电位、细胞内钙浓度和NLRP3炎性小体激活来实现的。