bcl2 d17c4 antibody  (Cell Signaling Technology Inc)

Journal: Pharmaceuticals

Article Title: IL-13Rα2 Is Involved in Resistance to Doxorubicin and Survival of Osteosarcoma Patients

doi: 10.3390/ph17111526

Figure Lengend Snippet: IL-13Rα2 is involved in the regulation of signaling molecules related to osteosarcoma cell proliferation and apoptosis: ( A ) Two osteosarcoma cells, U2OS and KHOS/NP, were transfected with empty vector, shRNA for IL-13Rα2, or wild-type IL-13Rα2 plasmids, and Western blotting performed for IL-13Rα2, TGF-β, snail, cyclin D1, BAX, BCL2, and actin. ( B ) Quantitative reverse-transcription polymerase chain reaction was performed for IL-13Rα2, TGF-β1, snail, cyclin D1, BAX, BCL2, and GAPDH in U2OS and KHOS/NP cells after transfection of vectors. The data were normalized with GAPDH expression. *** p < 0.001; EV, empty vector for wild-type IL-13Rα2; IL-13Rα2-OE, vector for wild-type IL-13Rα2; shControl, control vector for shRNA; shIL-13Rα2, vector for shRNA for IL-13Rα2.

Article Snippet: The following antibodies were used in Western blotting: IL-13RA2/CD213a2 (E7U7B) (#85677, Cell Signaling Technology, Beverly, MA, USA), transforming growth factor β (TGF-β) (#3709, Cell Signaling Technology, Beverly, MA, USA), snail (#ab180714, Abcam, Cambridge, UK), cyclin D1 (#2922, Cell Signaling Technology, Beverly, MA, USA), BAX (#2774, Cell Signaling Technology, Beverly, MA, USA), BCL2 (D17C4) (#3498, Cell Signaling Technology, Beverly, MA, USA), forkhead box O3 (FOXO3a) (75D8) (#2497, Cell Signaling Technology, Beverly, MA, USA), cleaved caspase-3 (Asp175) (5A1E) (#9664, Cell Signaling Technology, Beverly, MA, USA), and actin (#sc-47778, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Transfection, Plasmid Preparation, shRNA, Western Blot, Reverse Transcription, Polymerase Chain Reaction, Expressing, Control

Journal: Pharmaceuticals

Article Title: IL-13Rα2 Is Involved in Resistance to Doxorubicin and Survival of Osteosarcoma Patients

doi: 10.3390/ph17111526

Figure Lengend Snippet: Effects of IL-13Rα2 on apoptosis of osteosarcoma cells under treatment with doxorubicin: ( A ) U2OS and KHOS/NP cells were transfected with vectors, followed by 0.2 μM doxorubicin treatment and Western blot for IL-13Rα2, FOXO3a, BAX, BCL2, cleaved caspase 3, and actin was performed. ( B ) Flow cytometric analysis for apoptosis of osteosarcoma cells under treatment with doxorubicin. U2OS and KHOS/NP cells transfected with vectors were treated with 0.2 μM doxorubicin for 24 h, stained with Annexin V and propidium iodide, and examined using flow cytometry. The right upper and right lower quadrants were considered apoptotic populations. * p < 0.05; *** p < 0.001; ns , not significant; EV, empty vector for wild-type IL-13Rα2; IL-13Rα2-OE, vector for wild-type IL-13Rα2; shControl, control vector for shRNA; shIL-13Rα2, vector for shRNA for IL-13Rα2; DOX, doxorubicin.

Article Snippet: The following antibodies were used in Western blotting: IL-13RA2/CD213a2 (E7U7B) (#85677, Cell Signaling Technology, Beverly, MA, USA), transforming growth factor β (TGF-β) (#3709, Cell Signaling Technology, Beverly, MA, USA), snail (#ab180714, Abcam, Cambridge, UK), cyclin D1 (#2922, Cell Signaling Technology, Beverly, MA, USA), BAX (#2774, Cell Signaling Technology, Beverly, MA, USA), BCL2 (D17C4) (#3498, Cell Signaling Technology, Beverly, MA, USA), forkhead box O3 (FOXO3a) (75D8) (#2497, Cell Signaling Technology, Beverly, MA, USA), cleaved caspase-3 (Asp175) (5A1E) (#9664, Cell Signaling Technology, Beverly, MA, USA), and actin (#sc-47778, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Transfection, Western Blot, Staining, Flow Cytometry, Plasmid Preparation, Control, shRNA

Journal: Pharmaceuticals

Article Title: IL-13Rα2 Is Involved in Resistance to Doxorubicin and Survival of Osteosarcoma Patients

doi: 10.3390/ph17111526

Figure Lengend Snippet: Primer sequences used for a quantitative reverse-transcription polymerase chain reaction.

Article Snippet: The following antibodies were used in Western blotting: IL-13RA2/CD213a2 (E7U7B) (#85677, Cell Signaling Technology, Beverly, MA, USA), transforming growth factor β (TGF-β) (#3709, Cell Signaling Technology, Beverly, MA, USA), snail (#ab180714, Abcam, Cambridge, UK), cyclin D1 (#2922, Cell Signaling Technology, Beverly, MA, USA), BAX (#2774, Cell Signaling Technology, Beverly, MA, USA), BCL2 (D17C4) (#3498, Cell Signaling Technology, Beverly, MA, USA), forkhead box O3 (FOXO3a) (75D8) (#2497, Cell Signaling Technology, Beverly, MA, USA), cleaved caspase-3 (Asp175) (5A1E) (#9664, Cell Signaling Technology, Beverly, MA, USA), and actin (#sc-47778, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Reverse Transcription, Sequencing

Journal: Scientific Reports

Article Title: miR-124-3p downregulates EGR1 to suppress ischemia-hypoxia reperfusion injury in human iPS cell-derived cardiomyocytes

doi: 10.1038/s41598-024-65373-x

Figure Lengend Snippet: IH/R increases the expression of EGR1 and induces apoptosis in hiPSC-CMs. ( A ) A chart of the experiments of the induction of IH/R in hiPSC-CMs. ( B ) Representative immunofluorescence staining for cardiac isoform of troponin-T (cTnT) for hiPSC-CMs. ( C ) qRT-PCR of human EGR1 and RPLP1 (endogenous control) for hiPSC-CMs (n = 6). **P < 0.01. ( D , E ). Western blotting of human EGR1 and GAPDH (endogenous control) (n = 9). ****P < 0.0001. ( F , G ). Immunofluorescence staining for cleaved Caspase-3 (red), cTnT (green) and DAPI (blue) (n = 42). Scale bars 100 μm. ***P < 0.001. ( H ). Western blotting of human BCL2, BAX and GAPDH (endogenous control) (n = 6). ( I ). The ratio of BCL2/BAX in Control group and IH/R group, **P < 0.01.

Article Snippet: Subsequently, the membrane was blocked using 5% skim milk TBST buffer for 1 h, followed by incubation with the primary antibodies: EGR1 (44D5) Rabbit mAb (#4154, 75 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA), BCL2 (D17C4) Rabbit mAb (#3498, 26 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA), BAX (E4U1V) Rabbit mAb (#41,162, 20 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA) and GAPDH (100,118, 36KD, 1:5000, GeneTex, Irvine, CA, USA) at 4 °C overnight.

Techniques: Expressing, Immunofluorescence, Staining, Quantitative RT-PCR, Control, Western Blot

Journal: Scientific Reports

Article Title: miR-124-3p downregulates EGR1 to suppress ischemia-hypoxia reperfusion injury in human iPS cell-derived cardiomyocytes

doi: 10.1038/s41598-024-65373-x

Figure Lengend Snippet: miR-124-3p inhibits the pro-apoptotic effect of IH/R on hiPSC-CMs by downregulating EGR1. ( A ) qRT-PCR of human miR-124-3p and U6 (endogenous control) (n = 6). *P < 0.05. ( B ) qRT-PCR of EGR1 and RPLP1 (endogenous control) (n = 6). ****P < 0.0001, *P < 0.05. ( C ) qRT-PCR of GAPDH and RPLP1 (endogenous control) (n = 6). This experiment was conducted to demonstrate the technical efficiency of miRNA transfection. miRNA positive control was designed to antagonize GAPDH mRNA. ***P < 0.001, ****P < 0.0001, ( D , E ) Western blotting of EGR1 and GAPDH (endogenous control) (n = 7). **P < 0.01, ****P < 0.0001. ( F ). Western blotting of BCL2, BAX and GAPDH (endogenous control) (n = 6). ( G ). The ratio of BCL2/BAX in Control and IH/R groups. ***P < 0.001, ***P < 0.001 ( H , I ) Immunofluorescence staining for EGR1 (red), cTnT (green) and DAPI (blue) (n = 14). Scale bars 100 μm, ****P < 0.0001. ( J , K ) Immunofluorescence staining for cleaved Caspase-3 (red), cTnT (green) and DAPI (blue) (n = 14). Scale bars 100 μm, ****P < 0.0001.

Article Snippet: Subsequently, the membrane was blocked using 5% skim milk TBST buffer for 1 h, followed by incubation with the primary antibodies: EGR1 (44D5) Rabbit mAb (#4154, 75 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA), BCL2 (D17C4) Rabbit mAb (#3498, 26 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA), BAX (E4U1V) Rabbit mAb (#41,162, 20 kDa, 1:1000, Cell Signaling Technology, Danvers, MA, USA) and GAPDH (100,118, 36KD, 1:5000, GeneTex, Irvine, CA, USA) at 4 °C overnight.

Techniques: Quantitative RT-PCR, Control, Transfection, Positive Control, Western Blot, Immunofluorescence, Staining