Journal: iScience

Article Title: circGlis3 promotes β-cell dysfunction by binding to heterogeneous nuclear ribonucleoprotein F and encoding Glis3-348aa protein

doi: 10.1016/j.isci.2023.108680

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Article Snippet: Rabbit monoclonal anti-CCND2 , Cell Signaling Technology , Cat# 3741; RRID: AB_2070685.

Techniques: Virus, Recombinant, Modification, Lysis, Transfection, Enzyme-linked Immunosorbent Assay, Bicinchoninic Acid Protein Assay, CCK-8 Assay, Imaging, TUNEL Assay, Apoptosis Assay, In Situ Hybridization, Immunoprecipitation, Mass Spectrometry, Western Blot, shRNA, Plasmid Preparation, Software

Journal: Frontiers in Oncology

Article Title: Overexpression of miRNA-3613-3p Enhances the Sensitivity of Triple Negative Breast Cancer to CDK4/6 Inhibitor Palbociclib

doi: 10.3389/fonc.2020.590813

Figure Lengend Snippet: Overexpression of miR-3613-3p suppresses the proliferation, migration, and clonogenic ability induces G0/G1 cell-cycle arrest in TNBC cells. Effects of miR-3613-3p on TNBC cell growth and migration in vitro . After generation of miR-3613-3p stable overexpression in MDA-MB-231. (A) Cell vitality was evaluated by MTT assay, stably transfected with miR-3613-3p or NC lentivirus. (B, C) Colonigenic ability of different MDA-MB-231 cells were tested; generation of miR-3613-3p stable overexpression, the numbers of colony were counted and compared in the diagrams. (D, E) Cell migration of different MDA-MB-231 cells was tested using monolayer wound healing assay, generation of miR-3613-3p stable overexpression; the percent of wound closure was counted and compared in the diagrams. (F, G) Cell cycle of MDA-MB-231 cells stably transfected with miR-3613-3p or NC lentivirus was analyzed by flow cytometry assay; the percentage of cells in G0/G1, S, and G2/M phase is annotated in each column. (H, I) EdU assay of relative DAPI stained cells and EdU add-in cells. MDA-MB-231 cells were stably transfected with miR-3613-3p or NC lentivirus. At least 200 cells were counted per well. (J) Western blot analysis of positive cell cycle regulators CCND1, CCND2, CCND3, pRb, p-pRb, c-MYC, CDK4, and CDK6 protein in MDA-MB-231 cells transfected with transfected with miR-3613-3p lentivirus or vector control. P values were determined by two-tailed t-test, **P < 0.01; ****p < 0.0001.

Article Snippet: Primary antibodies were used as follows: anti-c-MYC (1:1,000, Proteintech), anti-EZH2, anti-SMAD2, anti-RB, anti-CDK4, anti-CDK6, anti-CCND1, anti-CCND2 and anti-CCND3 (1:1,000, Cell Signaling Technology) and anti-Actin (1:3,000, Sigma).

Techniques: Over Expression, Migration, In Vitro, MTT Assay, Stable Transfection, Transfection, Wound Healing Assay, Cytometry, EdU Assay, Staining, Western Blot, Plasmid Preparation, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: miR-198 Represses the Proliferation of HaCaT Cells by Targeting Cyclin D2

doi: 10.3390/ijms160817018

Figure Lengend Snippet: MiR-198 directly bound to the 3′-UTR of CCND2 mRNA. ( A ) Bioinformatics analyses showed that 10 potential target genes of miR-198 were predicted by three different databases; ( B ) The two distinct predicted binding sites of miR-198 in the 3′-UTR of CCND2 mRNA were allocated, and the fragments containing either mutated binding site were amplified according to the mature miR-198 sequence; ( C ) In pMIR-REPORT™ vector, CCND2 mRNA 3′-UTRfragment containing either the wild type or the mutated Site 1 was fused downstream the reporter gene. When the vectors were cotransfected with miR-198 mimic or mimic control, and the relative luciferase activity, normalised by β-gal, was significantly suppressed in vector with wild type Site 1 than that with mutated Site 1 (32.80% ± 6.89%); ( D ) In the presence of wild type Site 2, not the mutated one, miR-198 was able to significantly inhibit luciferase activity although to a less extent than wild type Site 1 (55.39% ± 8.48%). (NC, negative control; WT, wide-type; MT, mutated-type. * Compared with NC, p < 0.05).

Article Snippet: The antibodies used were as follows: mouse anti-GAPDH (AG019, Beyotime, China), mouse anti-CCND2 (BA2347-2, BOSTER, Wuhan, China), Horse Radish Peroxidase (HRP)-abeled Goat Anti-Mouse Immunoglobulin G (IgG) (H + L) (A0216, Beyotime, China).

Techniques: Binding Assay, Amplification, Sequencing, Plasmid Preparation, Luciferase, Activity Assay, Negative Control

Journal: International Journal of Molecular Sciences

Article Title: miR-198 Represses the Proliferation of HaCaT Cells by Targeting Cyclin D2

doi: 10.3390/ijms160817018

Figure Lengend Snippet: MiR-198 transfection repressed the mRNA and protein expression of CCND2. ( A ) MiR-198 mimic transfection reduced the expression of CCND2 mRNA at 24 h (68.09% ± 16.73%) and 48 h (45.68% ± 10.94%); ( B ) MiR-198 mimic or CCND2 siRNA transfection reduced the expression of CCND2 protein at 24 h (50.55% ± 24.04% or 34.45% ± 6.98%) and 48 h (17.38% ± 9.96% or 16.24% ± 9.23%). (NC, negative control. * Compared with NC, p < 0.05).

Article Snippet: The antibodies used were as follows: mouse anti-GAPDH (AG019, Beyotime, China), mouse anti-CCND2 (BA2347-2, BOSTER, Wuhan, China), Horse Radish Peroxidase (HRP)-abeled Goat Anti-Mouse Immunoglobulin G (IgG) (H + L) (A0216, Beyotime, China).

Techniques: Transfection, Expressing, Negative Control

Journal: International Journal of Molecular Sciences

Article Title: miR-198 Represses the Proliferation of HaCaT Cells by Targeting Cyclin D2

doi: 10.3390/ijms160817018

Figure Lengend Snippet: CCND2 siRNA transfection inhibited HaCaT cell proliferation by blocking cell cycle at G1 phase. ( A ) CCND2 siRNA transfection led to a significant reduction of mRNA expression of CCND2 in HaCaT cells both at 24 h (73.79% ± 11.45%) and 48 h (51.18% ± 8.85%); ( B ) Cell viability analysis showed that CCND2 siRNA transfection inhibited HaCaT cell proliferation at 24 h (67.98% ± 7.31%) and 48 h (67.45% ± 6.70%); ( C ) FCM analysis showed the effect of CCND2 siRNA transfection on cell cycle progression, and G1 phase arrest was obvious at 24 h (55.51% ± 6.18%) and 48 h (70.19% ± 4.10%) compared with negative control (42.98% ± 2.48%). (NC, negative control. * Compared with NC, p < 0.05).

Article Snippet: The antibodies used were as follows: mouse anti-GAPDH (AG019, Beyotime, China), mouse anti-CCND2 (BA2347-2, BOSTER, Wuhan, China), Horse Radish Peroxidase (HRP)-abeled Goat Anti-Mouse Immunoglobulin G (IgG) (H + L) (A0216, Beyotime, China).

Techniques: Transfection, Blocking Assay, Expressing, Negative Control