Journal: Molecular Medicine Reports

Article Title: Silencing the lncRNA NORAD inhibits EMT of head and neck squamous cell carcinoma stem cells via miR-26a-5p

doi: 10.3892/mmr.2021.12383

Figure Lengend Snippet: Isolation and identification of tumor stem cells from HNSCC cell lines. (A) Flow cytometry was used to sort ALDH − and ALDH + cells from HNSCC cell lines H357 and HSC-3. (B) ALDH + cells suspended in serum-free medium formed cell microspheres (magnification, ×100). (C) Protein expression levels of ALDH in control, ALDH − and ALDH + cells were detected using western blotting, (D) and the results were semiquantified. (E) Western blotting results of the (F) expression levels of tumor stem cell marker genes CD44, Oct-4 and Nanog in ALDH − and ALDH + H357 cells. (G) Western blotting results of the (H) expression levels of tumor stem cell marker genes CD44, Oct-4 and Nanog in ALDH − and ALDH + HSC-3 cells. *P<0.05, **P<0.01, ***P<0.001 vs. Control; ^^ P<0.01, ^^^ P<0.001 vs. ALDH − . ALDH, aldehyde dehydrogenase; HNSCC, head and neck squamous cell carcinoma.

Article Snippet: As recommended by the manufacturer, H357 cell lines (cat. no. 06092004) purchased from The European Collection of Authenticated Cell Cultures were cultured in a 37°C, 5% CO 2 environment in DMEM:F12 medium (cat. no. 30-2006) containing 2 mM glutamine (cat. no. 30-2214), 10% FBS (cat. no. 30-2020; all from the American Type Culture Collection), and 0.5 μg/ml sodium hydrocortisone succinate (cat. no. 125-04-2; Pure Chemistry Scientific, Inc.).

Techniques: Isolation, Flow Cytometry, Expressing, Control, Western Blot, Marker

Journal: Molecular Medicine Reports

Article Title: Silencing the lncRNA NORAD inhibits EMT of head and neck squamous cell carcinoma stem cells via miR-26a-5p

doi: 10.3892/mmr.2021.12383

Figure Lengend Snippet: Overexpression and knockdown of NORAD affects the expression levels of stem cell markers in H357 and HSC-3 cell lines. Transfection efficiencies of (A) overexpression and (B) siRNA vector were measured using reverse transcription-quantitative PCR. (C) Western blotting was used to detect the effect of NORAD overexpression on the expression levels of CD44, Oct-4 and Nanog in from H357 ALDH − cells. (D) Relative protein expression levels in H357 ALDH − cells. (E) Western blotting was used to detect the effect of NORAD overexpression on the expression levels of CD44, Oct-4 and Nanog in HSC-3 ALDH − cells. (F) Relative protein expression levels in HSC-3 ALDH − cells. (G) Western blotting was used to detect the effect of NORAD knockdown on the expression levels of CD44, Oct-4 and Nanog in H357 ALDH + cells. (H) Relative protein expression levels in H357 ALDH + cells. (I) Western blotting was used to detect the effect of NORAD knockdown on the expression levels of CD44, Oct-4 and Nanog in HSC-3 ALDH + cells. (J) Relative protein expression levels in HSC-3 ALDH + cells. GAPDH was used as a control. ^^^ P<0.001 vs. NORAD-NC; *P<0.05, **P<0.01, ***P<0.001 vs. siNORAD-siNC. NC, negative control; siRNA, small interfering RNA; NORAD, non-coding RNA activated by DNA damage; ALDH, aldehyde dehydrogenase; lncRNA, long non-coding RNA.

Article Snippet: As recommended by the manufacturer, H357 cell lines (cat. no. 06092004) purchased from The European Collection of Authenticated Cell Cultures were cultured in a 37°C, 5% CO 2 environment in DMEM:F12 medium (cat. no. 30-2006) containing 2 mM glutamine (cat. no. 30-2214), 10% FBS (cat. no. 30-2020; all from the American Type Culture Collection), and 0.5 μg/ml sodium hydrocortisone succinate (cat. no. 125-04-2; Pure Chemistry Scientific, Inc.).

Techniques: Over Expression, Knockdown, Expressing, Transfection, Plasmid Preparation, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot, Control, Negative Control, Small Interfering RNA

Journal: Molecular Medicine Reports

Article Title: Silencing the lncRNA NORAD inhibits EMT of head and neck squamous cell carcinoma stem cells via miR-26a-5p

doi: 10.3892/mmr.2021.12383

Figure Lengend Snippet: Long non-coding RNA NORAD knockdown decreases the migration and invasion of tumor stem cells via miR-26a-5p in HNSCC. Wound healing assay results from (A) H357 and (B) HSC-3 cells indicating the migration distances of the Blank, siNORAD-siNC, siNORAD, IC, I and siNORAD+I groups; magnification, ×100. (C) Relative migration rates are presented as bar diagrams. (D) Number of invasive tumor stem cells in each group was determined using Transwell assays in (E) H357 and (F) HSC-3 cells; magnification, ×250). *P<0.05, ***P<0.001 vs. siNORAD-siNC; ^ P<0.05, ^^ P<0.01, ^^^ P<0.001 vs. IC; # P<0.05, ### P<0.001 vs. I; Δ P<0.05, ΔΔΔ P<0.001 vs. siNORAD. I, microRNA-26a-5p inhibitor; IC, inhibitor control; NC, negative control; siRNA, small interfering RNA; NORAD, non-coding RNA activated by DNA damage.

Article Snippet: As recommended by the manufacturer, H357 cell lines (cat. no. 06092004) purchased from The European Collection of Authenticated Cell Cultures were cultured in a 37°C, 5% CO 2 environment in DMEM:F12 medium (cat. no. 30-2006) containing 2 mM glutamine (cat. no. 30-2214), 10% FBS (cat. no. 30-2020; all from the American Type Culture Collection), and 0.5 μg/ml sodium hydrocortisone succinate (cat. no. 125-04-2; Pure Chemistry Scientific, Inc.).

Techniques: Knockdown, Migration, Wound Healing Assay, Control, Negative Control, Small Interfering RNA

Journal: Molecular Medicine Reports

Article Title: Silencing the lncRNA NORAD inhibits EMT of head and neck squamous cell carcinoma stem cells via miR-26a-5p

doi: 10.3892/mmr.2021.12383

Figure Lengend Snippet: lncRNA NORAD knockdown attenuates the expression of EMT-related proteins in HNSCC tumor stem cells via miR-26a-5p. (A) Western blot analysis was used to detect the expression of MMP-2, MMP-9, E-cadherin, N-cadherin and Vimentin, and (B) relative protein expression is presented as bar diagrams in H357 cells. Reverse transcription-quantitative PCR was used to determine the mRNA expression of MMP-2, MMP-9, E-cadherin, N-cadherin and Vimentin in (C) H357 and (D) HSC-3 cells. (E) Western blot analysis was used to detect the expression of MMP-2, MMP-9, E-cadherin, N-cadherin and Vimentin, and (F) relative protein expression is presented as bar diagrams in HSC-3 cells. *P<0.05, **P<0.01, ***P<0.001 vs. siNORAD-siNC; ^ P<0.05, ^^ P<0.01, ^^^ P<0.001 vs. IC; # P<0.05, ## P<0.01, ### P<0.001 vs. I; Δ P<0.05, ΔΔ P<0.01, ΔΔΔ P<0.001 vs. siNORAD. I, microRNA-26a-5p inhibitor; IC, inhibitor control; NC, negative control; siRNA, small interfering RNA; NORAD, non-coding RNA activated by DNA damage.

Article Snippet: As recommended by the manufacturer, H357 cell lines (cat. no. 06092004) purchased from The European Collection of Authenticated Cell Cultures were cultured in a 37°C, 5% CO 2 environment in DMEM:F12 medium (cat. no. 30-2006) containing 2 mM glutamine (cat. no. 30-2214), 10% FBS (cat. no. 30-2020; all from the American Type Culture Collection), and 0.5 μg/ml sodium hydrocortisone succinate (cat. no. 125-04-2; Pure Chemistry Scientific, Inc.).

Techniques: Knockdown, Expressing, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction, Control, Negative Control, Small Interfering RNA