Journal: Journal of Immunology Research

Article Title: The Long Noncoding RNA MEG3 Retains Epithelial-Mesenchymal Transition by Sponging miR-146b-5p to Regulate SLFN5 Expression in Breast Cancer Cells

doi: 10.1155/2022/1824166

Figure Lengend Snippet: Intervention with MEG3 affects SLFN5 expression in BRCA cells. (a) Real-time PCR analysis of MEG3 RNA level in BRCA cell lines with different invasive capabilities, high-invasive capability cell lines MDA-MB-231 and BT549, and low-invasive capability cell lines MCF7 and T-47D. (b) Expression change of MEG3 RNA and SLFN5 mRNA in T-47D cells and MCF7 cells interfered with si-MEG3 or negative control siRNA (si-NC) analyzed by real-time PCR. (c) SLFN5 protein change in T-47D cells and MCF7 cells interfered with si-MEG3 or si-NC analyzed by Western blotting. (d) Expression change of MEG3 RNA and SLFN5 mRNA in BT-549 cells and MDA-MB-231 cells treated with MEG3 plasmids or control plasmids (NC) analyzed by real-time PCR. (e) SLFN5 protein change in BT-549 cells and MDA-MB-231 cells treated with MEG3 or NC plasmids analyzed by Western blotting. ∗∗ P < 0.01.

Article Snippet: Four types of human BRCA cells (MDA-MB-231, BT-549, MCF-7, and T-47D) and 293T cells were all from ATCC and cultured in Dulbecco's modified Eagle's medium (Gibco: C11885500BT, Australia) supplemented with 10% fetal bovine serum (Gibco: 10099-141, Australia).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Negative Control, Western Blot, Control

Journal: Journal of Immunology Research

Article Title: The Long Noncoding RNA MEG3 Retains Epithelial-Mesenchymal Transition by Sponging miR-146b-5p to Regulate SLFN5 Expression in Breast Cancer Cells

doi: 10.1155/2022/1824166

Figure Lengend Snippet: MEG3 represses BRCA cell migration, invasion, and EMT. (a, b) Morphological changes responding to si-MEG3 or si-NC interference (a) and MEG3 plasmid transfection (b) in indicated BRCA cells. (c, d) Relative mRNA level change of EMT molecular markers, vimentin, E-cadherin, and ZEB1, responding to si-MEG3 interference (c) or MEG3 plasmid transfection (d) in indicated BRCA cells. (e, f) Western blotting analyses of protein change of EMT markers responding to si-MEG3 transfection (e) or MEG3 plasmid transfection (f). (g–j) Cell migration and invasion capabilities evaluated by transwell assays responding to si-MEG3 (g, h) or MEG3 plasmid transfection (i, j). (k) Morphological changes of MCF-7 and T-47D cells responding to si-SLFN5 or si-NC interference. (l, m) Real-time and Western blotting analyses of vimentin, E-cadherin, and ZEB1, responding to si-SLFN5 or si-NC interference. ∗ P < 0.05, ∗∗ P < 0.01.

Article Snippet: Four types of human BRCA cells (MDA-MB-231, BT-549, MCF-7, and T-47D) and 293T cells were all from ATCC and cultured in Dulbecco's modified Eagle's medium (Gibco: C11885500BT, Australia) supplemented with 10% fetal bovine serum (Gibco: 10099-141, Australia).

Techniques: Migration, Plasmid Preparation, Transfection, Western Blot

Journal: Journal of Immunology Research

Article Title: The Long Noncoding RNA MEG3 Retains Epithelial-Mesenchymal Transition by Sponging miR-146b-5p to Regulate SLFN5 Expression in Breast Cancer Cells

doi: 10.1155/2022/1824166

Figure Lengend Snippet: MiR-146b-5p could bind to MEG3 and SLFN5 3′UTR directly by competing endogenous RNAs mechanism. (a, b) Twelve miRNAs predicted to regulate both SLFN5 and MEG3 by bioinformatics. (c, d) Comparison of 12 miRNA expression between control group and si-MEG3/MEG3 group in BRCA cell lines. (e) Schematic representation of binding sites between MEG3 and miR-146b-5p predicted by RNAhybrid. WT: wild type. MUT: mutant. (f) miR-146b-5p expression change in MCF7 cells after MEG3 plasmid transfection. (g) MEG3 expression change after transfection of either mimic (Mi) or inhibitor (AMO) of miR-146b-5p. (h) Luciferase reporter assay in 293T cells after transfection with MEG3 plasmids, either wild type (WT) or mutant (MUT), and Mi of miR-146b-5p. (i) RNA pull down assay of MEG3 binding to miR-146b-5p in MCF7 cells. (j) Schematic representation of binding sites between miR-146b-5p and SLFN5's 3′-UTR predicted by RNAhybrid. (k) Relative mRNA levels of SLFN5 responding to transfection of either mimic (Mi) or inhibitor (AMO) of miR-146b-5p. (l, m) Relative change of SLFN5 protein and mRNA responding to transfection of si-MEG3 or miR-146b-5p Mi or si-MEG3 plus miR-146b-5p inhibitor. (n) Luciferase reporter assay of transfection with SLFN5 plasmid, either WT or MUT, and miR-146b-5p Mi, or si-MEG3, or si-MEG3 plus miR-146b-5p AMO together.

Article Snippet: Four types of human BRCA cells (MDA-MB-231, BT-549, MCF-7, and T-47D) and 293T cells were all from ATCC and cultured in Dulbecco's modified Eagle's medium (Gibco: C11885500BT, Australia) supplemented with 10% fetal bovine serum (Gibco: 10099-141, Australia).

Techniques: Comparison, Expressing, Control, Binding Assay, Mutagenesis, Plasmid Preparation, Transfection, Luciferase, Reporter Assay, Pull Down Assay