mouse anti human monoclonal ccnd1  (Santa Cruz Biotechnology)

Journal: Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics

Article Title: MicroRNA-623 Targets Cyclin D1 to Inhibit Cell Proliferation and Enhance the Chemosensitivity of Cells to 5-Fluorouracil in Gastric Cancer

doi: 10.3727/096504018x15193469240508

Figure Lengend Snippet: Figure 3. miR-623 directly targets cyclin D1 (CCND1) in GC. (A) Wild-type (WT) and mutated (MUT) miR-623 binding sequences in the 3¢-untranslated region (3¢-UTR) of CCND1. (B) RT-qPCR and (C) Western blot were performed to determine the mRNA and protein levels of CCND1 in SGC-7901 and BGC-823 cells transfected with miR-623 mimic or miR-NC. *p < 0.05 compared with miR-NC. miR-623 or miR-NC was transfected in (D) SGC-7901 and (E) BGC-823 cells with psiCHECK-WT-CCND1-3¢-UTR or psiCHECK-MUT-CCND1-3¢-UTR. Relative luciferase activity levels were measured at 48 h posttransfection. *p < 0.05 compared with miR-NC.

Article Snippet: In this study, mouse anti-human monoclonal CCND1 (1:1,000 dilution; Catalog No. sc-8396) and mouse antihuman monoclonal b-actin (1:1,000 dilution; Catalog No. sc-81178) primary antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). b-Actin was utilized as a loading control for protein level normalization.

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

Journal: Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics

Article Title: MicroRNA-623 Targets Cyclin D1 to Inhibit Cell Proliferation and Enhance the Chemosensitivity of Cells to 5-Fluorouracil in Gastric Cancer

doi: 10.3727/096504018x15193469240508

Figure Lengend Snippet: Figure 4. CCND1 overexpression in GC tissues is inversely correlated with miR-623 level. (A) RT-qPCR and (B) Western blot were applied to measure the mRNA and protein expression levels of CCND1 in GC tissues and adjacent normal tissues, respectively. *p < 0.05 compared with normal tissues. (C) The association between CCND1 mRNA and miR-623 levels in GC tissues was assessed through Spearman’s correlation analysis. r = −0.5849, p = 0.0005.

Article Snippet: In this study, mouse anti-human monoclonal CCND1 (1:1,000 dilution; Catalog No. sc-8396) and mouse antihuman monoclonal b-actin (1:1,000 dilution; Catalog No. sc-81178) primary antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). b-Actin was utilized as a loading control for protein level normalization.

Techniques: Over Expression, Quantitative RT-PCR, Western Blot, Expressing

Journal: Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics

Article Title: MicroRNA-623 Targets Cyclin D1 to Inhibit Cell Proliferation and Enhance the Chemosensitivity of Cells to 5-Fluorouracil in Gastric Cancer

doi: 10.3727/096504018x15193469240508

Figure Lengend Snippet: Figure 5. CCND1 overexpression reverses the effects of miR-623 on GC cells. (A) CCND1 protein expression was detected in SGC- 7901 and BGC-823 cells cotransfected with miR-623 mimic and pcDNA3.1 or pcDNA3.1-CCND1 through Western blot. *p < 0.05 compared with miR-NC. #p < 0.05 compared with miR-623 mimics + pDNA3.1-CCND1. CCK-8 assay (B), cell chemosensitivity assay (C), and flow cytometry analysis of cell apoptosis (D) were performed to determine cell proliferation, chemosensitivity to 5-FU, and apoptosis induced by 5-FU in differently treated SGC-7901 and BGC-823 cells, respectively. *p < 0.05 compared with miR-NC. #p < 0.05 compared with miR-623 mimics + pcDNA3.1-CCND1.

Article Snippet: In this study, mouse anti-human monoclonal CCND1 (1:1,000 dilution; Catalog No. sc-8396) and mouse antihuman monoclonal b-actin (1:1,000 dilution; Catalog No. sc-81178) primary antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). b-Actin was utilized as a loading control for protein level normalization.

Techniques: Over Expression, Expressing, Western Blot, CCK-8 Assay, Flow Cytometry

Journal: Molecular medicine reports

Article Title: MicroRNA‑720 inhibits pancreatic cancer cell proliferation and invasion by directly targeting cyclin D1.

doi: 10.3892/mmr.2017.7732

Figure Lengend Snippet: Figure 3. CCND1 is a direct target of miR‑720 in pancreatic cancer. (A) Wild type and mutant of putative miR‑720 binding sites in the 3'‑untranslated region (3'‑UTR) of CCND1. (B) Relative luciferase activities in Panc‑1 and Sw1990 cells transfected with miR‑720 mimics or NC, together with pMIR‑CCND1‑3'‑UTR WT or pMIR‑CCND1‑3'‑UTR MUT. *P<0.05 compared with NC. (C and D) RT‑qPCR and Western blot analyses showed that miR‑720 overexpression decreased CCND1 mRNA and protein expression levels in Panc‑1 and Sw1990 cells. *P<0.05 compared with NC.

Article Snippet: Subsequently, the membranes were blocked by 5% non-fat milk in Tris-based saline-Tween 20 (TBST) for 1 h at room temperature and blotted with primary antibodies: mouse anti-human CCND1 monoclonal antibody (sc-450; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA) or mouse anti-human GAPDH monoclonal antibody (sc-47724; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA).

Techniques: Mutagenesis, Binding Assay, Luciferase, Transfection, Western Blot, Over Expression, Expressing

Journal: Molecular medicine reports

Article Title: MicroRNA‑720 inhibits pancreatic cancer cell proliferation and invasion by directly targeting cyclin D1.

doi: 10.3892/mmr.2017.7732

Figure Lengend Snippet: Figure 4. Inverse correlation between miR‑720 and CCND1 in pancreatic cancer tissues. (A and B) MRNA and protein expression levels of CCND1 in pancreatic cancer tissues and matched adjacent normal pancreatic tissues were determined using RT‑qPCR and Western blot analyses. *P<0.05 compared with adjacent normal pancreatic tissues. T, pancreatic cancer tissues; N, adjacent normal pancreatic tissues. (C) Evaluation of the inverse correlation between miR‑720 and CCND1 in pancreatic cancer tissues by Spearman's correlation analysis (r=‑0.6105, P=0.0020).

Article Snippet: Subsequently, the membranes were blocked by 5% non-fat milk in Tris-based saline-Tween 20 (TBST) for 1 h at room temperature and blotted with primary antibodies: mouse anti-human CCND1 monoclonal antibody (sc-450; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA) or mouse anti-human GAPDH monoclonal antibody (sc-47724; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA).

Techniques: Expressing, Western Blot

Journal: Molecular medicine reports

Article Title: MicroRNA‑720 inhibits pancreatic cancer cell proliferation and invasion by directly targeting cyclin D1.

doi: 10.3892/mmr.2017.7732

Figure Lengend Snippet: Figure 5. Upregulation of CCND1 prevents the inhibitory effects of miR‑720 on pancreatic cancer cells. (A) Protein expression of CCND1 in Panc‑1 and Sw1990 cells after transfection with miR‑720 mimics, NC or miR‑720 mimics, along with pcDNA 3.1‑CCND1. *P<0.05 compared with NC and miR‑720 mimics + pcDNA 3.1‑CCND1. (B and C) CCK8 assay and Matrigel invasion assay showed that CCND1 upregulation partly reversed the suppressive effects of miR‑720 on the proliferation and invasion of Panc‑1 and Sw1990 cells. *P<0.05 compared with NC and miR‑720 mimics + pcDNA 3.1‑CCND1.

Article Snippet: Subsequently, the membranes were blocked by 5% non-fat milk in Tris-based saline-Tween 20 (TBST) for 1 h at room temperature and blotted with primary antibodies: mouse anti-human CCND1 monoclonal antibody (sc-450; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA) or mouse anti-human GAPDH monoclonal antibody (sc-47724; 1:1,000 dilution; Santa Cruz Biotechnology, CA, USA).

Techniques: Expressing, Transfection, CCK-8 Assay, Invasion Assay

Journal: Oncology reports

Article Title: Downregulation of miR-34a contributes to the proliferation and migration of laryngeal carcinoma cells by targeting cyclin D1.

doi: 10.3892/or.2016.4823

Figure Lengend Snippet: Figure 3. CCND1 is a potential target of miR-34a. (A) Schematic of the putative miR-34a binding site in the 3'-UTR region of CCND1 and interspecies conservation of seed matching sequences (gray box). (B) Diagram of CCND1 3'-UTR containing the reporter constructs. (C) Luciferase reporter assays in HEp-2 cells co-transfected with wt/mut 3'-UTR and miR-34a/miR-Ctrl as indicated. (D) Expression levels of CCND1 were tested after miR-34a transfection at 50 nM in HEp-2 cells by western blotting assay. *P<0.05 compared with the control. 3'-UTR, 3'-untranslated region.

Article Snippet: After blocking for 1 h at room temperature and incubation overnight at 4̊C in Tris-buffered saline containing 0.05% Tween (TbsT) with 5% milk, the membranes were incubated with mouse monoclonal antibody against human CCND1 (Cell Signaling Technology, Danvers, MA, USA) or tubulin (Sigma-Aldrich) as a protein loading control, followed by horseradish peroxidase (HRp)-conjugated goat-anti-mouse igg (Abcam), and the bands were detected using the Supersignal West Pico eCL chemiluminescence kit (pierce) and Kodak X-ray film (Eastman Kodak Co, Rochester, NY, USA).

Techniques: Binding Assay, Construct, Luciferase, Transfection, Expressing, Western Blot, Control

Journal: Oncology reports

Article Title: Downregulation of miR-34a contributes to the proliferation and migration of laryngeal carcinoma cells by targeting cyclin D1.

doi: 10.3892/or.2016.4823

Figure Lengend Snippet: Figure 4. CCND1 is essential for LSCC proliferation and migration, and is involved in the miR-34a-induced effect. (A) Analysis of CCND1 protein expres- sion by western blot analysis. CCND1 protein was reduced by small interfering RNA (CCND1-siRNA) in the HEp-2 cells. (B) MTT assay showed that cell proliferation was inhibited in the HEp-2 cells after transfection with CCND1-siRNA, compared with the scrambled sequence (Ctrl-siRNA). (C) Apoptosis analysis of HEp-2 cells in response to CCND1-siRNA at 48 h, by FACS assay. (D) Analysis of the effect of CCND1-siRNA on the migration of HEp-2 cells by Transwell migration assay. (E) MTT assay shows the cell proliferation in HEp-2 cells co-transfected with miR-34a and wt/mut 3'-UTR-CCND1 compared with miR-Ctrl. *P<0.05 compared with the control. LSCC, laryngeal squamous cell carcinomas.

Article Snippet: After blocking for 1 h at room temperature and incubation overnight at 4̊C in Tris-buffered saline containing 0.05% Tween (TbsT) with 5% milk, the membranes were incubated with mouse monoclonal antibody against human CCND1 (Cell Signaling Technology, Danvers, MA, USA) or tubulin (Sigma-Aldrich) as a protein loading control, followed by horseradish peroxidase (HRp)-conjugated goat-anti-mouse igg (Abcam), and the bands were detected using the Supersignal West Pico eCL chemiluminescence kit (pierce) and Kodak X-ray film (Eastman Kodak Co, Rochester, NY, USA).

Techniques: Migration, Western Blot, Small Interfering RNA, MTT Assay, Transfection, Sequencing, Transwell Migration Assay, Control

Journal: Oncology reports

Article Title: Downregulation of miR-34a contributes to the proliferation and migration of laryngeal carcinoma cells by targeting cyclin D1.

doi: 10.3892/or.2016.4823

Figure Lengend Snippet: Figure 5. miR-34a and CCND1 are inversely correlated in LSCC tissues. (A) Statistical quantification of the mRNA expression levels of CCND1 (analyzed with q-PCR) between LSCC and paired NAT specimen. The CCND1 level was normalized to GAPDH (P<0.01). (B) A scatter diagram shows an inverse correlation between miR-34a and CCND1 expression in the same set of LSCC tissues (Spearman's correlation analysis, r =-7604; P<0.0001). NATs, normal adjacent tissues; LSCC, laryngeal squamous cell carcinomas.

Article Snippet: After blocking for 1 h at room temperature and incubation overnight at 4̊C in Tris-buffered saline containing 0.05% Tween (TbsT) with 5% milk, the membranes were incubated with mouse monoclonal antibody against human CCND1 (Cell Signaling Technology, Danvers, MA, USA) or tubulin (Sigma-Aldrich) as a protein loading control, followed by horseradish peroxidase (HRp)-conjugated goat-anti-mouse igg (Abcam), and the bands were detected using the Supersignal West Pico eCL chemiluminescence kit (pierce) and Kodak X-ray film (Eastman Kodak Co, Rochester, NY, USA).

Techniques: Expressing