Long noncoding RNA NEAT1 knockdown inhibits MPP(+)-induced apoptosis, inflammation and cytotoxicity in SK-N-SH cells by regulating miR-212-5p/RAB3IP axis.

Some long non-coding RNAs (lncRNAs) have been suggested to play critical roles in Parkinson's disease (PD) pathogenesis, including nuclear enriched abundant transcript 1 (NEAT1). The purpose of this study was to further elucidate the molecular mechanism of NEAT1 in PD. The expression levels of NEAT1, miR-212-5p and RAB3A-interacting protein (RAB3IP) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and apoptosis were evaluated by Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis, respectively. Western blot analysis was applied to detect the protein expression of IL-1β, TNF-α and RAB3IP. The LDH activity, ROS generation and SOD activity were measured by Lactate LDH activity assay kit, ROS assay kit, and SOD activity assay kit, respectively. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were performed to verify the relationship between miR-212-5p and NEAT1 or mRNA of RAB3IP. 1-methyl-4-phenylpyridinium ion (MPP+)-treated SK-N-SH cells were used as an in vitro model of PD. NEAT1 and RAB3IP were upregulated while miR-212-5p was downregulated in SK-N-SH cells treated with MPP+. NEAT1 knockdown or miR-212-5p overexpression inhibited MPP+-induced apoptosis, inflammation and cytotoxicity in SK-N-SH cells. Moreover, miR-212-5p was a direct target of NEAT1 and its downregulation reversed the eff ;ects caused by NEAT1 knockdown in MPP+-induced SK-N-SH cells. Furthermore, RAB3IP was a downstream target of miR-212-5p and its overexpression attenuated the effects of miR-212-5p restoration in MPP+-induced SK-N-SH cells. Besides, NEAT1 acted as a molecular sponge of miR-212-5p to regulate RAB3IP expression. NEAT1 knockdown suppressed MPP+-induced apoptosis, inflammation and cytotoxicity in SK-N-SH cells through regulating miR-212-5p and RAB3IP expression, providing a possible therapeutic strategy for PD patients.

Liu R ，Li F ，Zhao W 《-》

Deficiency of NEAT1 prevented MPP(+)-induced inflammatory response, oxidative stress and apoptosis in dopaminergic SK-N-SH neuroblastoma cells via miR-1277-5p/ARHGAP26 axis.

Noncoding RNAs including long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) have been documented to play prominent role in neurodegenerative diseases including Parkinson's disease (PD). This study intended to investigate the role of lncRNA nuclear enriched assembly transcript 1 (NEAT1) in MPP+-induced PD model in dopaminergic neuronblastoma SK-N-SH cells, as well as its mechanism through sponging miRNA (miR)-1277-5p. Real-time PCR and western blotting revealed that NEAT1 and ARHGAP26 were upregulated, and miR-1277-5p was downregulated in MPP+-treated SK-N-SH cells in a certain of concentration- and time- dependent manner. MPP+ induced apoptosis in SK-N-SH cells, as evidenced by decreased cell viability and Bcl-2 expression, and elevated apoptosis rate and levels of Bax and cleaved caspase-3, which were examined by MTT assay, flow cytometry and western blotting. Moreover, commercial assay kits indicated that inflammatory response and oxidative stress were provoked in response to MPP+, due to promoted contents of interleukin (IL)-6, IL-1β, tumor necrosis factor-α, malondialdehyde, and lactate dehydrogenase, accompanied with suppressed superoxide dismutase and glutathione peroxidase levels. Notably, MPP+-induced apoptosis, inflammatory response and oxidative stress in SK-N-SH cells were mitigated by NEAT1 knockdown and/or miR-1277-5p overexpression. Moreover, silencing of miR-1277-5p could abrogate the suppression of NEAT1 deficiency on MPP+-induced cell injury. Similarly, upregulating miR-1277-5p-elicited neuroprotection in MPP+-induced SK-N-SH cells was reversed by ARHGAP26 restoration. Dual-luciferase reporter assay demonstrated a direct interaction between miR-1277-5p and NEAT1 or ARHGAP26. Collectively, NEAT1 upregulation might contribute to MPP+-induced neuron injury via NEAT1-miR-1277-5p-ARHGAP26 competing endogenous RNAs (ceRNAs) pathway.

Zhou S ，Zhang D ，Guo J ，Chen Z ，Chen Y ，Zhang J ... -  《-》

LINC00943 knockdown attenuates MPP(+)-induced neuronal damage via miR-15b-5p/RAB3IP axis in SK-N-SH cells.

Meng C ，Gao J ，Ma Q ，Sun Q ，Qiao T ... -  《-》

Long non-coding RNA UCA1 regulates MPP(+)-induced neuronal damage through the miR-671-5p/KPNA4 pathway in SK-N-SH cells.

Parkinson's disease (PD) is an age-related neurodegenerative disease. Long non-coding RNA urothelial carcinoma-associated 1 (UCA1) is involved in the pathogenesis of PD. However, the pathogenesis of PD regulated by UCA1 has not been fully explained. We used 1-Methyl-4-phenylpyridinium (MPP+)-induced SK-N-SH cells for functional analysis. Expression levels of UCA1, microRNA (miR)-671-5p, and KPNA4 (karyopherin subunit alpha 4) mRNA were detected using quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and apoptosis were analyzed using MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) or flow cytometry assays. Some protein levels were measured by western blotting. The levels of pro-inflammatory cytokines were tested by ELISA (enzyme-linked immunosorbent assay). The levels of LDH (lactate dehydrogenase), MDA (malondialdehyde), and SOD (superoxide dismutase) were measured using corresponding kits. The relationship between UCA1 or KPNA4 and miR-671-5p was verified by dual-luciferase reporter assay and/or RNA immunoprecipitation (RIP) assay. MPP+ induced UCA1 expression in SK-N-SH cells in a concentration-dependent manner or time-dependent manner. UCA1 knockdown reduced MPP+-induced apoptosis, inflammation, and oxidative stress in SK-N-SH cells. MiR-671-5p was downregulated while KPNA4 was upregulated in MPP+-treated SK-N-SH cells. UCA1 sponged miR-671-5p to regulate KPNA4 expression. MiR-671-5p inhibition counteracted UCA1 knockdown-mediated influence on apoptosis, inflammation, and oxidative stress of MPP+-induced SK-N-SH cells. KPNA4 overexpression offset the inhibitory influence of miR-671-5p mimic on apoptosis, inflammation, and oxidative stress of MPP+-treated SK-N-SH cells. UCA1 inhibition reduced MPP+-induced neuronal damage through the miR-671-5p/KPNA4 pathway in SK-N-SH cells, providing a novel mechanism to understand the pathogenesis of PD.

Hao Z ，Dang W ，Zhu Q ，Xu J ... -  《-》

Long Noncoding RNA SNHG1 Knockdown Ameliorates Apoptosis, Oxidative Stress and Inflammation in Models of Parkinson's Disease by Inhibiting the miR-125b-5p/MAPK1 Axis.

Parkinson's disease (PD) is a prevalent neurodegenerative disease. Long noncoding RNA small molecule RNA host gene 1 (SNHG1) has been reported to play critical roles in Parkinson's disease (PD) progression. The study aimed to further elucidate the mechanism of SNHG1 in PD pathogenesis. The levels of SNHG1, miR-125b-5p and mitogen-activated protein kinase 1 (MAPK1) were determined by quantitative real-time polymerase chain reaction (qRT-PCR) or Western blot. Cell viability and apoptosis were evaluated by Cell Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. The activity of Caspase-3 or Caspase-9 was measured using a Caspase-3 or Caspase-9 Assay Kit. The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β, lactic dehydrogenase (LDH) activity, reactive oxygen species (ROS) generation and superoxide dismutase (SOD) activity were gauged by enzyme-linked immunosorbent assay (ELISA). Dual-luciferase reporter assay was performed to identify the relationship between miR-125b-5p and SNHG1 or MAPK1. The MPTP-induced PD mouse was used as an in vivo model of PD and MPP+-treated SK-N-SH and MN9D cells were used as in vitro models of PD. SNHG1 and MAPK1 were significantly up-regulated while miR-125b-5p was down-regulated in the MPTP-induced PD mouse model and MPP+-induced PD cell models. SNHG1 silence or miR-125b-5p overexpression protected against MPP+-evoked apoptosis, oxidative stress and inflammation in SK-N-SH and MN9D cells. Moreover, SNHG1 acted as a molecular sponge of miR-125b-5p, and the protective impact of SNHG1 silence on MPP+-evoked cell damage was reversed by miR-125b-5p inhibition. Furthermore, MAPK1 was a functional target of miR-125b-5p and its overexpression attenuated the effects of miR-125b-5p restoration in MPP+-triggered cell injury. In addition, the behavioral changes in MPTP-induced PD mouse in vivo model were relieved by SNHG1 silence. SNHG1 knockdown exerted neuroprotective effects in MPP+-evoked cytotoxicity through regulating the miR-125b-5p/MAPK1 axis both in human and mouse PD cell models, highlighting a possible target for PD therapy.

Xiao X ，Tan Z ，Jia M ，Zhou X ，Wu K ，Ding Y ，Li W ... -  《Neuropsychiatric Disease and Treatment》