PD is the commonest severe neurodegenerative disease of the central nervous system. Although the pathogenesis of PD is unclear, the crucial role of apoptosis has been demonstrated in PD models. Recently, it has been revealed that lncRNAs are used as a precise biomarker for PD[24–26]. In this study, MPP+ were used to establish PD models in vitro and we focused on the role and relationship of NEAT1, miR-132-3p and apoptotic factors in PD.
SH-SY5Y cells could express the tyrosine hydroxylase, dopamine transporter and dopamine 2B hydroxylase which produced by catecholaminergic neurons, it has been frequently used in neuroscientific research as an in vitro model for the investigation of pathogenesis of PD. Neurotoxin MPTP is a neurotoxin which could destroy dopamine neurons selectively through causing oxidative stress-mediated apoptotic death to mimic PD. In the study, SH-SY5Y cells were treated with MPP+ at varying concentrations to determine the suitable concentration to construct PD cell models, and we selected 1mM MPP+ for further experiment.
Growing studies demonstrated that lncRNA played an essential role in PD development. It has been reported that NEAT1 was upregulated and downregulation of NEAT1 could effectively suppress apoptosis and autophagy in PD mouse model[12, 13]. Apoptosis is indispensable for building an effective neural network in the developing brain. However, excessive apoptosis could accelerate disease progression, including PD. Proteins of the Bcl-2 family are known as crucial regulators of apoptosis, which includes anti-apoptotic Bcl-2 and pro-apoptotic Bax members. The significant increase of Bax/Bcl-2 could result in the collapse of the mitochondrial membrane potential and caspase-3 activation, ultimately leading to cell death[29, 30]. Previous studies have showed that Bax expression upregulates and Bcl-2 expression downregulation in the SNpc and striatum of mice of PD. Lacking of Bax could protect against MPP+-induced neuronal damage[31, 32]. In our study, the expression of Bcl-2 and Bax is consistent with previous studies. Knockdown the expression of NEAT1 significantly prevented the increase in Bax expression and the decrease in Bcl-2 expression, restoring the Bax/Bcl-2 ratio in MPP+-treated cells. Interestingly, we also found the increase of Bim/caspase 3 and decrease of p-Foxo3a expression in cells after MPP + treatment. Decrease NEAT1 expression could also downregulate the Bim/caspase 3 and upregulate p-Foxo3a expression. FoxO3a is highly expressed in the brain and mediates a range of pro-apoptotic genes related to neuronal cell death as an important transcription factor. It has been reported that activated FoxO3a could induce pro-apoptotic protein such as Bim in variety of neuron death paradigms. P-FoxO3a could suppress FoxO3a activity via docking with 14-3-3 and excluding FoxO3a from the nucleus[34, 35]. Caspase-3 is classically known executioner of apoptosis and activated by Bim in response to related stimuli. A study showed amyloid-β induced astrocytosis and astrocyte death via FoxO3a-Bim-caspase3 death signaling. Based on these results, we speculate NEAT1 acts at apoptosis in PD pathogenesis through FOXO3a/Bim/caspase 3 and Bax/Bcl-2/caspase 3 signal pathway.
LncRNAs could reverse regulate miRNA-mediated suppression of target mRNA through competing with other genes for combining with miRNAs. The lncRNA-miRNA-mRNA network has been shown to exert crucial influence in PD. NEAT1 could prompt autophagy and apoptosis through mediating miR-374c-5p in MPTP-induced PD. Knockdown of NEAT1 inhibited PD development via regulating miR-212-3p/AXIN1 pathway. In this study, we hypothesized that NEAT1 also acts as a miRNA sponge to regulate PD progression. TargetScan6.2 analysis and dual-luciferase reporter assay were preformed, and results showed NEAT1 contained binding sites for miR-132-3p. Recently, emerging studies indicated that miR-132-3p regulated apoptosis in PD. For instance, MIAT suppressed MPP+-induced apoptosis by regulating miR-132/SIRT1 axis in PC12 cells. Initially, significant high expression of miR-132-3p was found in midbrain tissues in PD patients. MiR-132-3p may serve as the potential biomarkers for early diagnosis of PD. A previous study revealed downregulation of miR-132-3p expression could dramatically attenuate neuroinflammation and degeneration of dopaminergic neurons induced by MPTP in PD mouse models. Knockdown of NEAT1 inhibits glioma cell migration and invasion through modulation of SOX2 targeted by miR-132. However, there is no evidence to support the regulatory network NEAT1/miR-132-3p/apoptotic factors in PD, and the underlying mechanism needs to be clarified. In this study, we revealed that NEAT1 could regulate apoptosis in SH-SY5Y cells treated by MPP+. Further studies indicated NEAT1 could bind directly to miR-132-3p. These results strongly demonstrated that knockdown the level of NEAT1 could reduce cell apoptosis through FOXO3a/Bim/caspase 3 and Bax/Bcl-2/caspase 3 pathways. Therefore, we speculated that NEAT1 exerted effect by regulating the miR-132-3p/FOXO3a/Bim/caspase 3 and miR-132-3p/Bax/Bcl-2/ caspase 3 pathways.
In conclusion, the study suggested MPP+ treatment could upregulate the levels of NEAT and apoptotic related factors, decreasing the expression of miR-132-3p and anti-apoptotic Bcl-2. Knockdown of NEAT1 could restrain apoptosis process of PD via mediating the miR-132-3p/FOXO3a /Bim/caspase 3 and miR-132-3p /Bax/Bcl-2/caspase 3 signal pathways. which might offer innovative ideas for alleviating the PD progression.