Journal: Cancer medicine

Article Title: Nuclear receptor coactivator 6 (NCoA6) promotes cell proliferation, migration, and invasion in pancreatic cancer.

doi: 10.1002/cam4.6427

Figure Lengend Snippet: FIGURE 3 NCoA6 knockdown decreases the proliferation, cell cycle, and apoptosis of pancreatic cancer cells. CCK-8 assays (A, B) and colony formation assays (C, D) were used to test the proliferation of PDAC cells transfected with NCoA6 shRNAs. The representative images and statistical results of the cell cycle (E, F) and apoptosis (G, H) using flow cytometry. (I, J) Western blotting analysis of p21 and p27 expression in NCoA6-silenced PANC-1 and SW1990 cells. (K, L) Western blotting analysis of CDK4, CDK2, Cyclin D1, Cyclin E1, and Cyclin A2 expression in NCoA6-silenced PANC-1 and SW1990 cells.

Article Snippet: Protein extraction and western blotting were performed as described in our previous research.16 Antibodies against NCoA6 (1:1000), E- cadherin (1:5000), N- cadherin (1:3000), FBW7 (1:1000), CDX2 (1:1000), cyclin- dependent kinase 4 (CDK4, 1:2000), cyclin- dependent kinase 2 (CDK2, 1:5000), Cyclin D1 (1:5000), Cyclin E1 (1:1000), and Cyclin A2 (1:2000) were obtained from Proteintech.

Techniques: Knockdown, CCK-8 Assay, Transfection, Flow Cytometry, Western Blot, Expressing

Journal: Journal of Biological Chemistry

Article Title: Selective Enhancing Effect of Early Mitotic Inhibitor 1 (Emi1) Depletion on the Sensitivity of Doxorubicin or X-ray Treatment in Human Cancer Cells

doi: 10.1074/jbc.m112.446351

Figure Lengend Snippet: FIGURE 7. Synergistic anticancer effect of Emi1 knockdown in combination with doxorubucin in normal cells. A, Emi1 or control siRNA was transfected into NHDF cells, and cells were collected after 48 h. The indicated proteins in siRNA-treated NHDF cells were examined by Western blotting. B, Emi1 siRNA and/or cyclin A siRNA were transfected into NHDF cells. The left panel shows the expression of Emi1 and cyclin A examined by Western blot analysis after 48 h of siRNA transfection. -Actin expression was used as a loading control. The right panel shows DAPI staining and percentage of cells with polyploidy after Emi1 siRNA and/or cyclin A siRNA transfection. Cells were stained with DAPI to visualize the nuclei, and percentage of cells with polyploidy was determined by DNA content analysis after PI staining using a flow cytometer. *, p 0.05. C, flow cytometric analysis of annexin V and PI staining in control and Emi1 siRNA treated NHDF or HFL III cells after treatment with doxorubucin (DOXY; 0.5 g/ml) for 12 h. We performed three independent experiments.

Article Snippet: Human cyclin A siRNA (sc-29282) and human E2F1 siRNA (sc-61861) were obtained from Santa Cruz Biotechnology.

Techniques: Knockdown, Control, Transfection, Western Blot, Expressing, Staining, Flow Cytometry

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Super enhancer-driven LINC01013 mediates hypoxia-induced mitochondrial dysfunction by HSPA9 to determine pulmonary arterial smooth muscle cell fate

doi: 10.1007/s00018-025-06071-3

Figure Lengend Snippet: Silencing of LINC01013 inhibited hypoxia-induced hPASMC proliferation and inflammation. A , B hPASMC proliferation was determined by CCK8 and EdU incorporation assays ( n = 6). EdU (red), DAPI (blue), scale bar, 50 μm. C Western blotting analysis of PCNA, cyclin A and cyclin D in hPASMCs ( n = 6). D RT‒qPCR analysis showed the mRNA levels of TNF-α and IL-6 after LINC01013 knockdown, β-actin was used as a internal reference gene ( n = 6). E Western blotting analysis of TNF-α and IL-6 in hPASMCs. ( n = 6). All values are presented as the mean ± SEM. Statistical analysis was performed with one-way ANOVA. * p < 0.05, ** p < 0.01, *** p < 0.001. Nor, normoxia; Hyp, hypoxia; NC, negative control; si, siRNA

Article Snippet: The antibody against CEBPB (PB9171, BA0670, 1:500, Boster, Wuhan, China), H3K27ac (A7253, 1:500, ABclonal, Wuhan, China), H3K4me1 (A2355, 1:500, Wuhan, China), PCNA (A00125, 1:500, Boster, Wuhan, China), Cyclin A (PB0515, 1:500, Boster, Wuhan, China), Cyclin D (BM4272, 1:500, Boster, Wuhan, China), IL-6 (AF7236, 1:500, Beyotime, Shanghai, China), TNF-α (AF8208, 1:500, Beyotime, Shanghai, China), PKM2 (4053, 1:1000, Cell Signaling, MA, US), HK II (66974-1-Ig, 1:1000, Proteintech, IL, USA), PDH (2784, 1:1000, Cell Signaling, MA, US), HSPA9 (14887-1-AP, 1:5000, Proteintech, IL, USA), VDAC1 (10866-1-AP, 1:5000, Proteintech, IL, USA), and β-actin (TA-09, 1:1000, ZSGB‐BIO, Beijing, China) was incubated at 4 °C overnight, followed by incubation with appropriate horseradish peroxidase-conjugated secondary antibodies at room temperature for 1 h, and proteins were visualized with enhanced chemiluminescence reagents.

Techniques: Western Blot, Knockdown, Negative Control

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: MiR-29c-3p Suppresses the Migration, Invasion and Cell Cycle in Esophageal Carcinoma via CCNA2/p53 Axis

doi: 10.3389/fbioe.2020.00075

Figure Lengend Snippet: Primer sequence.

Article Snippet: In order to investigate the underlying mechanism of CCNA2 on EC cells, cells were classified into four groups: si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβand si-CCNA2 + PFTβ groups [PFTβ, p53 inhibitor, HY-16702, MedChemExpress, 10 μM ( )]. qRT-PCR was firstly performed to test CCNA2 level in each group, finding that CCNA2 was markedly decreased in cells transfected with si-CCNA2 + DMSO ( ).

Techniques: Sequencing

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: MiR-29c-3p Suppresses the Migration, Invasion and Cell Cycle in Esophageal Carcinoma via CCNA2/p53 Axis

doi: 10.3389/fbioe.2020.00075

Figure Lengend Snippet: MiR-29c-3p is decreased in EC tissues accompanied by low survival rate and associated with the increase of CCNA2. TCGA database was utilized to access expression data of miRNAs and mRNAs of ESCA, and (A) the results of differential analysis were plotted in Volcano plots, with red representing high expression and green representing low expression. In panel (B) , miR-29c-3p level in EC tissues were determined as shown in a box plot. (C) Survival analysis of miR-29c-3p in TCGA-ESCA dataset was performed, with the red line as high expression and blue line as low expression. In panel (D) , Venn diagram was made to find the candidate targets of miR-29c-3p, acquiring 10 DEmRNAs. In panel (E) , correlation analysis was conducted between miR-29c-3p and CCNA2 (–0.57) as plotted in a heat map. In panel (F) , CCNA2 expression in EC cells was examined. Clinical tissue samples were used to further explore the (G) expression of miR-29c-3p and CCNA2 mRNA in EC tissues by qRT-PCR, (H) the protein level of CCNA2 (P1, P2, P3 referred to three EC samples) via Western blot and (I) the correlation between miR-29c-3p and CCNA2. * P < 0.05.

Article Snippet: In order to investigate the underlying mechanism of CCNA2 on EC cells, cells were classified into four groups: si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβand si-CCNA2 + PFTβ groups [PFTβ, p53 inhibitor, HY-16702, MedChemExpress, 10 μM ( )]. qRT-PCR was firstly performed to test CCNA2 level in each group, finding that CCNA2 was markedly decreased in cells transfected with si-CCNA2 + DMSO ( ).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: MiR-29c-3p Suppresses the Migration, Invasion and Cell Cycle in Esophageal Carcinoma via CCNA2/p53 Axis

doi: 10.3389/fbioe.2020.00075

Figure Lengend Snippet: MiR-29c-3p targets CCNA2 and inhibits its expression. Targeted binding sites of miR-29c-3p and CCNA2 were predicted before as shown in panel (A) . To investigate their targeted relationship, (B) dual-luciferase assay was performed to confirm their targeted binding, and (C) RIP was conducted to describe the effect of miR-29c-3p on CCNA2. Moreover, (D,E) qRT-PCR and Western blot were carried out to determine CCNA2 expression in mRNA and protein levels in miR-29c-3p mimic transfected cells, so as to further verify such relationship. * P < 0.05.

Article Snippet: In order to investigate the underlying mechanism of CCNA2 on EC cells, cells were classified into four groups: si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβand si-CCNA2 + PFTβ groups [PFTβ, p53 inhibitor, HY-16702, MedChemExpress, 10 μM ( )]. qRT-PCR was firstly performed to test CCNA2 level in each group, finding that CCNA2 was markedly decreased in cells transfected with si-CCNA2 + DMSO ( ).

Techniques: Expressing, Binding Assay, Luciferase, Quantitative RT-PCR, Western Blot, Transfection

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: MiR-29c-3p Suppresses the Migration, Invasion and Cell Cycle in Esophageal Carcinoma via CCNA2/p53 Axis

doi: 10.3389/fbioe.2020.00075

Figure Lengend Snippet: CCNA2 silencing regulates the migration, invasion and cell cycle in EC by promoting p53 signaling pathway. si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβ and si-CCNA2 + PFTβ were transfected into cells. qRT-PCR and Western blot were conducted to determine (A) the CCNA2 mRNA and (B) protein levels of CCNA2 as well as p53. MTT, Transwell, and flow cytometry were performed to investigate the effects of silencing CCNA2 on EC cell activities, including (C) cell viability, (D) migration and invasion, (E) cell cycle. * P < 0.05.

Article Snippet: In order to investigate the underlying mechanism of CCNA2 on EC cells, cells were classified into four groups: si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβand si-CCNA2 + PFTβ groups [PFTβ, p53 inhibitor, HY-16702, MedChemExpress, 10 μM ( )]. qRT-PCR was firstly performed to test CCNA2 level in each group, finding that CCNA2 was markedly decreased in cells transfected with si-CCNA2 + DMSO ( ).

Techniques: Migration, Transfection, Quantitative RT-PCR, Western Blot, Flow Cytometry

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: MiR-29c-3p Suppresses the Migration, Invasion and Cell Cycle in Esophageal Carcinoma via CCNA2/p53 Axis

doi: 10.3389/fbioe.2020.00075

Figure Lengend Snippet: MiR-29c-3p mediates the migration, invasion and cell cycle in EC via CCNA2/p53 axis. Cells were treated with inhibitor NC + si-NC, inhibitor NC + si-CCNA2, miR-29c-3p inhibitor + si-NC and miR-29c-3p inhibitor + si-CCNA2, and then harvested for (A) Western blot to detect the protein levels of CCNA2 and p53. (B) MTT was performed to test cell viability, (C) Transwell was conducted to assay the ability of cell migration and invasion, and (D) flow cytometry was carried out to determine the effect of miR-29c-3p on cell cycle. * P < 0.05.

Article Snippet: In order to investigate the underlying mechanism of CCNA2 on EC cells, cells were classified into four groups: si-NC + DMSO, si-CCNA2 + DMSO, si-NC + PFTβand si-CCNA2 + PFTβ groups [PFTβ, p53 inhibitor, HY-16702, MedChemExpress, 10 μM ( )]. qRT-PCR was firstly performed to test CCNA2 level in each group, finding that CCNA2 was markedly decreased in cells transfected with si-CCNA2 + DMSO ( ).

Techniques: Migration, Western Blot, Flow Cytometry

Journal: International immunopharmacology

Article Title: Immunosensitivity mediated by downregulated AKT1-SKP2 induces anti-PD-1-associated thyroid immune injury.

doi: 10.1016/j.intimp.2023.110452

Figure Lengend Snippet: Fig. 5. The downstream regulation of AKT1-SKP2 in NTHY cells.

Article Snippet: SKP2 inhibitor (SKP2i, MCE, 5uM) is solubilized by DMSO.

Techniques:

Journal: International immunopharmacology

Article Title: Immunosensitivity mediated by downregulated AKT1-SKP2 induces anti-PD-1-associated thyroid immune injury.

doi: 10.1016/j.intimp.2023.110452

Figure Lengend Snippet: Fig. 6. Interaction between SKP2 and PD-L1. (A) DAPI is in blue, SKP2 in red, and PD-L1 in green. In NTHY cells, SKP2 is mainly expressed in the nucleus and PD- L1 in non-nuclear area. (B) NTHY cells are co-transfected with pcDNA3.1-SKP2 plasmid and pcDNA3.1-PDL1 plasmid. The interaction between SKP2 and PD-L1 protein is proved by immunoprecipitation.

Article Snippet: SKP2 inhibitor (SKP2i, MCE, 5uM) is solubilized by DMSO.

Techniques: Transfection, Plasmid Preparation, Immunoprecipitation

Journal: International immunopharmacology

Article Title: Immunosensitivity mediated by downregulated AKT1-SKP2 induces anti-PD-1-associated thyroid immune injury.

doi: 10.1016/j.intimp.2023.110452

Figure Lengend Snippet: Fig. 8. Hypothesis: NIVO increases the immunosensitivity of NTHY cells by downregulating AKT1-SKP2.

Article Snippet: SKP2 inhibitor (SKP2i, MCE, 5uM) is solubilized by DMSO.

Techniques:

Journal: International immunopharmacology

Article Title: Immunosensitivity mediated by downregulated AKT1-SKP2 induces anti-PD-1-associated thyroid immune injury.

doi: 10.1016/j.intimp.2023.110452

Figure Lengend Snippet: Fig. 7. AKT1, SKP2, PD-L1 and IFN-γ in thyroid tissue. (A) The thyroid gland of mice is used for detection by immunohistochemistry. Fewer AKT1, SKP2, PD-L1 and more IFN-γ are observed in the NIVO group than in the CTRL group (10X). (B) Further, levels of various proteins in mouse thyroid are quantified by WB.(C) Human thyroid tissue is used for detection by WB. results revealed that PD-L1 is down-regulated and IFNG is up-regulated in the HT group compared to the non-HT group, however, there is no difference in AKT1 and SKP2.

Article Snippet: SKP2 inhibitor (SKP2i, MCE, 5uM) is solubilized by DMSO.

Techniques: Immunohistochemistry