Journal: Oncology reports

Article Title: Radiosensitization of esophageal carcinoma cells by knockdown of HMGB1 expression.

doi: 10.3892/or.2018.6923

Figure Lengend Snippet: Figure 4. Suppression of HMGB1 increases the apoptosis rates of esophageal carcinoma cells after IR in vitro and affects the generation of apoptosis‑related proteins. (A) The apoptotic rate was calculated as the sum of B2 and B4. As the results revealed, compared with the NC group, silencing of HMGB1 sensitized ECA109 and TE13 cells to apoptosis both with 6 Gy radiation (A, lower images) and without (A, upper images). (B) After transfection with HMGB1‑shRNA, the expression of Bcl‑2 was attenuated, while the expression of Bax and caspase‑9 was increased in ECA109 and TE13 cells. (C) HMGB1‑shRNA and irradia- tion induced G0/G1 arrest in ESCC cells. (D) The results of flow cytometry revealed that the percentages of G0/G1‑phase cells in the HMGB1‑shRNA groups were significantly higher compared to those of the NC groups before IR, and the HMGB1‑shRNA group percentages significantly increased after irradiation. (E) In the HMGB1‑shRNA group, the expression of cyclin D1 and CDK4 was attenuated, while the expression of p16 was increased. Additionally, irradiation inhibited cyclin D1 and CDK4 expression in esophageal tumor cells. A comparison with the NC group is symbolized by an asterisk *P<0.05; and a comparison with the corresponding non‑irradiated group is symbolized by triangle ▲▲P<0.01. HMGB1, high mobility group box 1; IR, irradiation.

Article Snippet: The relevant antibodies were anti-HMGB1 (dilution 1:10,000; cat. no. ab79823), anti-γH2AX (di lut ion 1:1,000; cat. no. ab26350; Abcam), anti-MMP-2 (dilution 1:1,000; cat. no. 10373-2-AP), anti-MMP-9 (dilution 1:1,000; cat. no. 10375-2-AP), anti-p16 (cat. no. 10883-1-AP), anti-caspase-9 (dilution 1:1,000; cat. no. 10380-1-AP), anti-Bcl-2 (dilution 1:2,000; cat. no. 12789-1-AP), anti-CDK4 (dilution 1:2,000; cat. no. 11026-1-AP), anti-cyclin D1 (dilution 1:5,000; cat. no. 60186-1-lg), anti-Bax (dilution 1:5,000; cat. no. 50599-2-lg; ProteinTech Group, Inc., Chicago, IL, USA) and anti-β-actin (dilution 1:10,000; cat. no. AP0060; Bioworld Technology, Inc., St. Louis Park, MN, USA).

Techniques: In Vitro, Transfection, Expressing, Flow Cytometry, Irradiation, Comparison

Journal: Oncology reports

Article Title: Platycodin D, a metabolite of Platycodin grandiflorum, inhibits highly metastatic MDA-MB-231 breast cancer growth in vitro and in vivo by targeting the MDM2 oncogene.

doi: 10.3892/or.2016.4935

Figure Lengend Snippet: Figure 3. PD induced cell cycle arrest in the G0/G1 phase in MDA-MB-231 cells. (A) Cells were exposed to 2.5, 5, 10 or 20 µM of PD for 48 h and the cell cycle progression was assessed by flow cytometry. The 5, 10 and 20 µM concentrations of PD induced significant cell cycle arrest in the G0/G1 phase. (B and C) The expression levels of G0/G1 phase-related proteins was analyzed by western blotting after treatment with various concentrations of PD for 24 h. PD down regulated the expression of CDK2, CDK4, CDK6, and Cyclin E. (*p<0.05).

Article Snippet: The antibodies against human CDK2, CDK4, CDK6, and Cyclin E used in the western blot analyses were obtained from Boster Biotechnology (Wuhan, China).

Techniques: Flow Cytometry, Expressing, Western Blot

Journal: Oncology reports

Article Title: Platycodin D, a metabolite of Platycodin grandiflorum, inhibits highly metastatic MDA-MB-231 breast cancer growth in vitro and in vivo by targeting the MDM2 oncogene.

doi: 10.3892/or.2016.4935

Figure Lengend Snippet: Figure 9. The effects of PD on the expression levels of various proteins in MDA-MB-231 xenograft tumors. Western blotting was performed to assess the expression levels of proteins in the MDM2-MDMX-p53 pathway, down- stream proteins and G0/G1 phase-related proteins in mice after treatment with the three concentrations of PD for four weeks. (A) PD downregulated the expression of the MDM2, MDMX, and mutant p53 proteins. (B) PD upregulated the expression of p21 and p27. (C) PD decreased the expression of G0/G1 phase-related proteins (CDK2, CDK4, CDK6 and Cyclin E).

Article Snippet: The antibodies against human CDK2, CDK4, CDK6, and Cyclin E used in the western blot analyses were obtained from Boster Biotechnology (Wuhan, China).

Techniques: Expressing, Western Blot, Mutagenesis

Journal: Cells

Article Title: Artesunate Inhibits the Cell Growth in Colorectal Cancer by Promoting ROS-Dependent Cell Senescence and Autophagy

doi: 10.3390/cells11162472

Figure Lengend Snippet: Artesunate caused excessive mitochondrial ROS to induce cell senescence and inhibit cell proliferation. ( A ) Artesunate arrested cell cycle at G0/G1 phase in SW480 and HCT116. Cells were seeded in 6-well plates. Once attached, cells were maintained in FBS-free medium for about 16~24 h. Afterwards, cell were treated with artesunate (1, 2, and 4 μM in medium containing 10% FBS) for 72 h and then harvested for PI staining to analyze cell cycle by flow cytometry. ( B ) Artesunate induced cell senescence in SW480 and HCT116. Cells were seeded in 6-well plates and treated with artesunate (1, 2, and 4 μM) for 72 h. Cell senescence was represented via SA-β-gal activity, which was assayed using a SA-β-gal staining kit after artesunate treatment. ( C ) Artesunate treatment downregulated the protein levels of CDK 2/4/6 and upregulated the protein levels of CDKIs, p16, and p21 in SW480 and HCT116. Cells were seeded in 60 mm dishes and treated with artesunate (1, 2, and 4 μM) for 72 h. Then, cells were lysed with RIPA to extract total protein. The protein levels were measured by western blotting. The gray values of protein blots were evaluated by Image J. Relative protein expression was normalized to β-actin. ( D ) NAC attenuated the effect of artesunate on protein expression of p16 in SW480 and HCT116. NAC was used at a concentration of 2 mM and added alone or together with artesunate (4 μM) for 72 h. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. Ctrl. ### p < 0.001 vs. cells treated with artesunate alone. Data were shown as mean ± SD.

Article Snippet: CDK2, CDK4, CDK6, Cyclin D1, Cyclin E1, p21, Rb, phosphorylated-Rb, caspase 3, cleaved-caspase 3, PARP, cleaved-PARP, Bax, Bcl-2, BIP, PERK, IRE1α, CHOP, DR5, Beclin 1, LC3A/B, Atg3, Atg5, Atg7, and Atg12 primary antibodies for western blotting were purchased from Cell Signaling Technology Inc. (Danvers, MA, USA). p16 and phosphor-IRE1α primary antibodies for western blotting were purchased from Abcam (Cambridge, UK). p16, p21, and Cyclin D1 primary antibodies for immunohistochemistry and β-actin, XBP1S, and XBP1U primary antibodies for western blotting were purchased from Proteintech Group, Inc. (Manchester, UK).

Techniques: Staining, Flow Cytometry, Activity Assay, Western Blot, Expressing, Concentration Assay

Journal: Cells

Article Title: Artesunate Inhibits the Cell Growth in Colorectal Cancer by Promoting ROS-Dependent Cell Senescence and Autophagy

doi: 10.3390/cells11162472

Figure Lengend Snippet: Artesunate inhibited the growth of CT26-derived tumor in vivo. ( A ) Experimental timeline of CT26-derived tumor model in balb/c mice. Balb/c mice were injected CT-26 cells (1 × 10 5 cells for each mouse) subcutaneously to establish the CT26-derived tumor model. Tumor-loaded mice were gavaged with artesunate at 30 mg/kg or 60 mg/kg for 24 days. Body weights and tumor volumes were recorded every three days. ( B ) Curve of tumor volume. ( C ) Tumor weight. Tumor tissues were collected and weighted after treatment. ( D ) Immunohistochemical images of Ki67, Cyclin D1, p16, p21, LC3B, and p-IRE1α. Immunohistochemistry assay was performed using a SABC-POD staining kit. ( E ) Curve of body weight. ( F ) Organ indexes. Lung, heart, spleen, liver, and kidney were collected and weighted to calculate the organ indexes after treatment. * p < 0.05, *** p < 0.001 vs. Model group. Data were shown as mean ± SD.

Article Snippet: CDK2, CDK4, CDK6, Cyclin D1, Cyclin E1, p21, Rb, phosphorylated-Rb, caspase 3, cleaved-caspase 3, PARP, cleaved-PARP, Bax, Bcl-2, BIP, PERK, IRE1α, CHOP, DR5, Beclin 1, LC3A/B, Atg3, Atg5, Atg7, and Atg12 primary antibodies for western blotting were purchased from Cell Signaling Technology Inc. (Danvers, MA, USA). p16 and phosphor-IRE1α primary antibodies for western blotting were purchased from Abcam (Cambridge, UK). p16, p21, and Cyclin D1 primary antibodies for immunohistochemistry and β-actin, XBP1S, and XBP1U primary antibodies for western blotting were purchased from Proteintech Group, Inc. (Manchester, UK).

Techniques: Derivative Assay, In Vivo, Injection, Immunohistochemical staining, Immunohistochemistry, Staining

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: Screening of anti-phosphoT172 CDK4 mAbs using immobilized cyclin D3/CDK4 proteic fusions.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques:

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: Sera evaluation by CDK4 peptide ELISA.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques: Peptide ELISA

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: Evaluation of CDK4 T172-phosphospecificity of immune sera.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques:

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: CDK4 T172-phosphospecificity of cloned mAbs.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques: Clone Assay

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: T172-phosphospecific CDK4 sandwich ELISA.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques: Sandwich ELISA

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: S130-phosphorylated p21 is enriched in phospho-CDK4 immunoprecipitation.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques: Immunoprecipitation

Journal: Cell Cycle

Article Title: Monoclonal antibodies to activated CDK4: use to investigate normal and cancerous cell cycle regulation and involvement of phosphorylations of p21 and p27

doi: 10.1080/15384101.2021.1984663

Figure Lengend Snippet: S10-phosphorylated p27 is enriched in phospho-CDK4 immunoprecipitation.

Article Snippet: Equal amounts of whole cell extract protein were separated according to molecular mass and immunodetected using the following antibodies: monoclonal mouse antibodies against cyclin D3 (DCS-22) (Neomarkers/ThermoFisher); cyclin D1 (DCS-6), p16 (DCS-50), CDK4 (DCS-31) (Santa Cruz Biotechnology); p21, p27, CDK4 antibodies (respectively 12D1, D69C12, D9G3E, Cell Signaling Technology); or rabbit polyclonal (H22) CDK4 and anti-phospho-p27 (S10) antibodies (Santa Cruz Biotechnology).

Techniques: Immunoprecipitation