LncRNA NEAT1 promotes airway smooth muscle cell inflammation by activating the JAK3/STAT5 pathway through targeting of miR-139.

Background Asthma is a chronic inflammatory heterogeneous respiratory disease. Previous studies showed that the lncRNA NEAT1 (nuclear paraspeckle assembly transcript 1) might play an important role in the pathogenesis of asthma, but its potential mechanism in airway smooth muscle cell (ASMC) inflammation remains largely unknown and needs further investigation.Methods We performed cellular immunofluorescence to identify the features of ASMCs and detected the expression levels of lncRNA NEAT1, miR-139, TNF-α, IL-6, IL-8 and IL-1ß by quantitative real-time PCR (Q-PCR) and ELISA. Western blotting (WB) was used to measure the protein expression of the related genes, and bioinformatics as well as dual luciferase assays were used to validate the interaction between lncRNA NEAT1 and miR-139 and the interaction between miR-139 and the 3'-UTR of JAK3.Results The expression of lncRNA NEAT1 was increased in the ASMCs of asthma patients, but miR-139 was decreased. Overexpression of lncRNA NEAT1 promoted the expression of the inflammatory cytokines such as TNF-α, IL-6, IL-8 and IL-1ß in ASMCs. LncRNA NEAT1 was able to target miR-139 to activate the JAK3/STAT5 signaling pathway and induced the expression of these inflammatory cytokines in ASMCs. Overexpression of miR-139 or suppression of the JAK3/STAT5 signaling pathway reversed the inflammatory effect of lncRNA NEAT1.Conclusion LncRNA NEAT1 played a pivotal role in ASMC inflammation and exerted its function through the miR-139/JAK3/STAT5 signaling network.

[Effect of Nrf2 and TrxR on proliferation of chronic myeloid leukemia cell and its mechanism].

OBJECTIVE: To explore the effect of nuclear factor erythroid-2 related factor 2 (Nrf2) and thioredoxin reductase (TrxR) gene on proliferation of chronic myeloid leukemia (CML) line cells and its mechanism. METHODS: Four interfering sequences of Nrf2 and one negative control sequence were designed and synthesised based on the principle of target sequence of siRNA, then constructed lentivirus vectors, which were transfected into K562 cell lines. The transfection effect was observed by laser scanning confocal microscope (LSCM) and flow cytometer (FCM); The depressing effect of siRNA was analyzed by real-time PCR. The cell proliferation inhibiting rate was measured with CCK-8 assay, the apoptotic rate by Annexin V-PE/PI with FCM and the apoptotic morphology of cells by LSCM. RESULTS: The transfection efficiency of lentivirus was 65%. One cell line K562-C3 which significantly inhibited Nrf2 mRNA was obtained by real-time PCR, Nrf2 relative quantitation (RQ) expressions were 1.003±0.093 and 0.344±0.032 in the control group and K562-C3 respectively; TrxR expression also decreased with RQ as 1.090±0.549 and 0.395±0.029 respectively. The cellular proliferation inhibition rates of K562-C3 were (4.74±0.39)%, (6.13±1.78)% and (25.36±3.77)%, respectively at 24, 48 and 72 h. The apoptotic rate induced by K562-C3 (29.9%) at 72 hours was obviously higher than in the control group (7.9%). The Annexin V-PE positive K562-C3 cells presented the following apoptotic characteristics, such as karyopyknosis, nuclear fragmentation and apoptotic bodies observed by LSCM. CONCLUSION: Nrf2 specific siRNA could repress its expression at the cellular level and down-regulate the expression of its downstream antioxidant enzyme, such as TrxR, which lead to increased apoptotic rate and decreased cell proliferation.