Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A) Quantitative real-time PCR of miR-145 expression in glioma samples (n = 19) compared with normal samples (n = 10). (B) quantitative real-time PCR of miR-145 expression in rat glioma tissues (n = 6) and U87, U251 glioma cells compared to normal tissues (n = 6). (C) Quantitative real-time PCR of BNIP3 mRNA expression in glioma samples (n = 19) compared with normal samples (n = 10). (D) quantitative real-time PCR of BNIP3 mRNA expression in rat glioma tissues and U87, U251 glioma cells compared to normal tissues. (E) Pathology observation of mice brain tissues sections stained with IHC (×100, ×200, ×400). (F) Immunofluorescence with BNIP3 (green) in rat normal tissues and glioma tissues (×200).*p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Real-time Polymerase Chain Reaction, Expressing, Staining, Immunofluorescence, Control

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A) Western analysis of BNIP3 and control β-actin in glioma samples compared with normal samples. (B) Western analysis of BNIP3 and control β-actin in rat glioma tissues compared to normal tissues. *p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Western Blot, Control

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A) Bioinformatics analysis shows the seed sequence of miR-145 binding to the 3′-UTR of BNIP3 mRNA. (B) , (C) Quantitative real-time PCR analysis of mRNA expression of BNIP3 in U87 cells treated with miR-145 mimics and inhibitor for 48 h. (D) Western analysis of protein expression of BNIP3 in U87 and U251 cells treated with miR-145 mimics and inhibitor for 48 h. (E) Immunofluorescence with BNIP3 (green) in U87 and U251 cells after miR-145 mimics or mimics NC treatment.*p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Sequencing, Binding Assay, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Immunofluorescence, Control

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A) Western analysis of BNIP3, which is localized in the nucleus or the cytoplasm in U87 and U251 cells treated with miR-145 mimics and inhibitor for 48 h. (B) Wild-type 3′-UTR of BNIP3 gene was cloned into the firefly and Renilla reporter plasmid. The BNIP3-3′UTR constructs or blank plasmid were transfected into U87 and U251 cells with control or miR-145 mimics, followed by dual luciferase assays. *p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Western Blot, Clone Assay, Plasmid Preparation, Construct, Transfection, Control, Luciferase

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A, B) Transfection effect of BNIP3-siRNA or BNIP3-vector was confirmed by quantitative real-time PCR. (C) U87 cells and U251 cells were stained with Hoechst 33342 dye after BNIP3-siRNA or control treatment. (D) U87 and U251 cells were stained with Tunel after BNIP3-siRNA or control treatment. (E) U87 cell apoptosis after BNIP3-siRNA or control treatment was determined by FACS. *p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Transfection, Plasmid Preparation, Real-time Polymerase Chain Reaction, Staining, Control, TUNEL Assay

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A, B) Protein expression of Notch1, p21 and Hes1 was determined by western blot analysis in U87 and U251 cells transfected with miR-145 mimics and mimics-NC, or miR-145 inhibitor and inhibitor-NC. (C, D) Western analysis of Notch1-related proteins in U87 and U251 cells transfected with Bnip3 siRNA and siRNA control, or BNIP3 expression vector and blank vector. *p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Expressing, Western Blot, Transfection, Control, Plasmid Preparation

Journal: Oncotarget

Article Title: MicroRNA-145 induces apoptosis of glioma cells by targeting BNIP3 and Notch signaling

doi: 10.18632/oncotarget.18604

Figure Lengend Snippet: (A) Protein expression of Notch1, p21, Hes1 was determined by western blot analysis in U87 and U251 cells co-transfected with miR-145 inhibitor and BNIP3-siRNA, or with miR-145 inhibitor. *p < 0.05, **p < 0.01 versus control group.

Article Snippet: Rabbit monoclonal antibodies against Bax, Bcl-2, Caspase-3, and p21 (Abcam, USA) were used at 1:600 dilution, rabbit monoclonal antibody against Notch 1 Hes1 (Cell Signaling Technology, USA) was used at 1:600, anti-BNIP3 antibodies (Boster, China), anti-β-actin and anti-PCNA (ZSGB-BIO, China) were diluted 1:300

Techniques: Expressing, Western Blot, Transfection, Control

Journal: Frontiers in Microbiology

Article Title: Autophagy Induced by the N-Terminus of the Classic Swine Fever Virus Nonstructural Protein 5A Protein Promotes Viral Replication

doi: 10.3389/fmicb.2021.733385

Figure Lengend Snippet: Nonstructural protein 5A expression induces cellular autophagy. (A) PK-15 cells were transfected for NS5A expression, infected with CSFV (MOI=1) for 36h or treated with rapamycin (Rapa, 100nM) for 48h. (B-D) The expression of light chain 3 (LC3), ATG5, BECN1, and GAPDH (loading control) was analyzed by immunoblotting with specific antibodies described in the Materials and Methods. (E) PK-15 cells were transfected with NS5A or treated with Rapa (100nM) for 48h. The cells were then fixed and processed for indirect immunofluorescence using antibodies against LC3B, followed by the corresponding secondary antibodies conjugated to FITC as described in the Materials and Methods. The cell nuclei were counterstained with DAPI. The fluorescence signals were visualized by confocal immunofluorescence microscopy. In the images, nuclear staining is shown in blue, and LC3B staining is shown in green. (F) The average number of LC3B puncta in each cell was determined from at least 100 cells in each group. The data represent the mean±SD of three independent experiments. Statistical comparisons were evaluated using ANOVA followed by Tukey’s HSD post hoc analysis. *** p <0.001, ** p <0.01, and compared with the mock group.

Article Snippet: The primary antibodies were as follows: anti-LC3B (Abcam, ab-229327), anti-ATG5 (Bioss, bs-4005R), anti-Beclin 1 (Bioss, bs-1353R), and anti-GAPDH (Bioss, bs-0755R).

Techniques: Expressing, Transfection, Infection, Western Blot, Immunofluorescence, Fluorescence, Microscopy, Staining

Journal: Frontiers in Microbiology

Article Title: Autophagy Induced by the N-Terminus of the Classic Swine Fever Virus Nonstructural Protein 5A Protein Promotes Viral Replication

doi: 10.3389/fmicb.2021.733385

Figure Lengend Snippet: Expression of autophagy marker proteins in cells expressing different portions of the NS5A gene. (A) PK-15 cells were transfected with sequences expressing different portions of the NS5A gene or a mock plasmid. The expression of LC3, ATG5, BECN1, and GAPDH (loading control) was analyzed by immunoblotting with specific antibodies described in the Materials and Methods. The data represent the mean±SD of three independent experiments. Statistical comparisons were evaluated using ANOVA followed by Tukey’s HSD post hoc analysis. *** p <0.001, compared with the mock group.

Article Snippet: The primary antibodies were as follows: anti-LC3B (Abcam, ab-229327), anti-ATG5 (Bioss, bs-4005R), anti-Beclin 1 (Bioss, bs-1353R), and anti-GAPDH (Bioss, bs-0755R).

Techniques: Expressing, Marker, Transfection, Plasmid Preparation, Western Blot

Journal: Frontiers in Microbiology

Article Title: Autophagy Induced by the N-Terminus of the Classic Swine Fever Virus Nonstructural Protein 5A Protein Promotes Viral Replication

doi: 10.3389/fmicb.2021.733385

Figure Lengend Snippet: The key amino acids of NS5A affect the expression of autophagy marker proteins. (A) PK-15 cells were transfected with constructs expressing NS5A with specific amino acid substitutions. The expression of LC3, ATG5, BECN1, and GAPDH (loading control) was analyzed by immunoblot with specific antibodies described in the Materials and Methods. The data represent the mean±SD of three independent experiments. Statistical comparisons were evaluated using ANOVA followed by Tukey’s HSD post hoc analysis. *** p <0.001, ** p <0.01, compared with the mock group.

Article Snippet: The primary antibodies were as follows: anti-LC3B (Abcam, ab-229327), anti-ATG5 (Bioss, bs-4005R), anti-Beclin 1 (Bioss, bs-1353R), and anti-GAPDH (Bioss, bs-0755R).

Techniques: Expressing, Marker, Transfection, Construct, Western Blot