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LncRNA MEG3 Participates in Caerulein-Induced Inflammatory Injury in Human Pancreatic Cells via Regulating miR-195-5p/FGFR2 Axis and Inactivating NF-κB Pathway

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Abstract

Acute pancreatitis (AP) is a dysfunctional pancreas disease marked by severe inflammation. Long non-coding RNAs (lncRNAs) involving in the regulation of inflammatory responses have been frequently mentioned. The purpose of this study was to ensure the function and action mode of lncRNA maternally expressed gene 3 (MEG3) in caerulein-induced AP cell model. HPDE cells were treated with caerulein to establish an AP model in vitro. The expression of MEG3, miR-195-5p, and fibroblast growth factor receptor 2 (FGFR2) was measured using quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation and apoptosis were detected by 3-(4, 5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and flow cytometry assay, respectively. The expression of CyclinD1, B cell lymphoma/leukemia-2 (Bcl-2), Bcl-2-associated X protein (Bax), FGFR2, P65, phosphorylated P65 (p-P65), alpha inhibitor of nuclear factor kappa beta (NF-κB) (IκB-α), and phosphorylated IκB-α (p-IκB-α) at the protein level was quantified by western blot. The concentrations of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) were monitored by enzyme-linked immunosorbent assay (ELISA). The targeted relationship between miR-195-5p and MEG3 or FGFR2 was forecasted by the online software starBase v2.0 and verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. As a result, the expression of MEG3 and FGFR2 was decreased in caerulein-induced HPDE cells, while the expression of miR-195-5p was increased. MEG3 overexpression inhibited cell apoptosis and inflammatory responses that were induced by caerulein. Mechanically, miR-195-5p was targeted by MEG3 and abolished the effects of MEG3 overexpression. FGFR2 was a target of miR-195-5p, and MEG3 regulated the expression of FGFR2 by sponging miR-195-5p. FGFR2 overexpression abolished miR-195-5p enrichment-aggravated inflammatory injuries. Moreover, the NF-κB signaling pathway was involved in the MEG3/miR-195-5p/FGFR2 axis. Collectively, MEG3 participates in caerulein-induced inflammatory injuries by targeting the miR-195-5p/FGFR2 regulatory axis via mediating the NF-κB pathway in HPDE cells.

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  1. Department of Emergency and Critical Medicine, The Second Hospital of Jilin University, No. 218, Nanguan District, Ziqiang Street, Changchun, Jilin, 130041, China

    Xinghai Chen & Debiao Song

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Figure S1.

Cell viability was decreased in caerulein-treated HPDE cells in a dose/time-dependent manner. (A) Cell viability in HPDE cells exposed to different doses of caerulein (5, 10 and 15 nmol/L) was assessed by MTT assay. (B) Cell viability in HPDE cells exposed to 10 nmol/L caerulein for different time (4, 8 and 12 h) was assessed by MTT assay. *P < 0.05. (PNG 674 kb)

High resolution image (TIF 112 kb)

Figure S2.

LncRNA MEG3 was mainly distributed in the cytoplasm. The distribution of MEG3 in the cytoplasm and nucleus was examined using qRT-PCR, and GAPDH or U6 acted as the control in cytoplasm or nucleus, respectively. *P < 0.05. (PNG 365 kb)

High resolution image (TIF 48 kb)

Figure S3.

MEG3 repressed caerulein-induced apoptosis of HPDE cells by targeting miR-195-5p, and miR-195-5p promoted caerulein-induced apoptosis of HPDE cells by binding to FGFR2. (A) Cell apoptosis was monitored in caerulein-treated HPDE cells transfected with MEG3, pcDNA, MEG3+miR-195-5p or MEG3+miR-NC using flow cytometry assay. (B) Cell apoptosis was determined in caerulein-treated HPDE cells transfected with miR-195-5p, miR-NC, miR-195-5p+FGFR2 or miR-195-5p+pcDNA by flow cytometry analysis. *P < 0.05. (PNG 2892 kb)

High resolution image (TIF 941 kb)

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Chen, X., Song, D. LncRNA MEG3 Participates in Caerulein-Induced Inflammatory Injury in Human Pancreatic Cells via Regulating miR-195-5p/FGFR2 Axis and Inactivating NF-κB Pathway. Inflammation 44, 160–173 (2021). https://doi.org/10.1007/s10753-020-01318-6

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