Journal: Oncology Letters

Article Title: Anticancer effect of the oncolytic Newcastle disease virus harboring the PTEN gene on glioblastoma

doi: 10.3892/ol.2024.14752

Figure Lengend Snippet: Effect of rNDV-PTEN infection on apoptotic cell death through imbalance of Akt/mTOR pathway. U87-MG cells were infected with rNDV or rNDV-PTEN at an MOI of 1 for 12, 24 or 36 h. (A) Cell proliferation markers PCNA and MMP9, (B) mTOR signaling-related proteins and autophagy-related proteins and (C) pre-apoptotic cell death-related proteins were assessed using immunoblotting analysis in U87-MG cells. GAPDH was used as an internal control. *P<0.05 vs. CON. rNDV, recombinant Newcastle disease virus; PTEN, phosphatase and tensin homolog; PCNA, proliferating cell nuclear antigen; MMP9, matrix metallopeptidase 9; MOI, multiplicity of infection; CON, control; Bcl-2, B-cell lymphoma 2; Bax, Bcl-2-associated X protein.

Article Snippet: The membranes were incubated overnight at 4°C with the following primary antibodies: Anti-GAPDH (1:3,000; cat. no. sc-32233; Santa Cruz Biotechnology, Inc.); anti-LC3 (1:1,000; cat. no. NB100-2220; Novus Biologicals, LLC); anti-matrix metallopeptidase 9 (MMP9; 1:1,000; cat. no. MA5-15886; Thermo Fisher Scientific, Inc.); anti-proliferating cell nuclear antigen (PCNA; 1:500; cat. no. PC 10; Sigma-Aldrich; Merck KGaA); anti-P-mTOR (Ser2448; 1:1,000; cat. no. 2971S; Cell Signaling Technology, Inc.); anti-mTOR (1:1,000; cat. no. 2972S; Cell Signaling Technology, Inc.); anti-P-Akt (Ser473; 1:1,000; cat. no. 9271S; Cell Signaling Technology, Inc.); anti-Akt (1:1,000; cat. no. 9272S; Cell Signaling Technology, Inc.); anti-cleaved Caspase (Cas)9 (1:1,000; cat. no. 9509S; Cell Signaling Technology, Inc.); anti-cleaved Cas3 (1:1,000; cat. no. 9664S; Cell Signaling Technology, Inc.); anti-cleaved Cas8 (1:1,000; cat. no. 9496S; Cell Signaling Technology, Inc.); anti-B-cell lymphoma 2 (Bcl-2) associated X protein (Bax; 1:1,000; cat. no. 2772S; Cell Signaling Technology, Inc.); anti-p62 (1:1,000; cat. no. 5114S; Cell Signaling Technology, Inc.); anti-Occludin (1:1,000; cat. no. 5506S; Cell Signaling Technology, Inc.); anti-zonula occludens protein 1 (ZO-1; 1:1,000; cat. no. 5406S; Cell Signaling Technology, Inc.); anti-Clauddin-5 (E8F3D; 1:1,000; cat. no. 49564; Cell Signaling Technology, Inc.); and anti-PTEN (1:1,000; cat. no. 9552S; Cell Signaling Technology, Inc.).

Techniques: Infection, Western Blot, Control, Recombinant, Virus

Journal: Oncology Letters

Article Title: Anticancer effect of the oncolytic Newcastle disease virus harboring the PTEN gene on glioblastoma

doi: 10.3892/ol.2024.14752

Figure Lengend Snippet: PTEN restoration induces apoptotic cell death by impacting mTOR signaling and autophagy in the orthotopic GBM mouse model. (A) Immunohistochemical staining of Ki-67 (tumor cell proliferation marker) and MMP9 in the GBM orthotopic mouse tissue. Scale bar, 50 µm. (B) PCNA and MMP9 levels, (C) proteins related to the mTOR signaling pathway and autophagy, and (D) apoptosis markers were assessed using immunoblotting in GBM tissue. These data are expressed as the fold change in expression compared with PBS injected mice. GAPDH was used as an internal control. *P<0.05 vs. PBS-injected mice. PTEN, phosphatase and tensin homolog; MMP9, matrix metallopeptidase 9; PCNA, proliferating cell nuclear antigen; GBM, glioblastoma; CON, control.

Article Snippet: The membranes were incubated overnight at 4°C with the following primary antibodies: Anti-GAPDH (1:3,000; cat. no. sc-32233; Santa Cruz Biotechnology, Inc.); anti-LC3 (1:1,000; cat. no. NB100-2220; Novus Biologicals, LLC); anti-matrix metallopeptidase 9 (MMP9; 1:1,000; cat. no. MA5-15886; Thermo Fisher Scientific, Inc.); anti-proliferating cell nuclear antigen (PCNA; 1:500; cat. no. PC 10; Sigma-Aldrich; Merck KGaA); anti-P-mTOR (Ser2448; 1:1,000; cat. no. 2971S; Cell Signaling Technology, Inc.); anti-mTOR (1:1,000; cat. no. 2972S; Cell Signaling Technology, Inc.); anti-P-Akt (Ser473; 1:1,000; cat. no. 9271S; Cell Signaling Technology, Inc.); anti-Akt (1:1,000; cat. no. 9272S; Cell Signaling Technology, Inc.); anti-cleaved Caspase (Cas)9 (1:1,000; cat. no. 9509S; Cell Signaling Technology, Inc.); anti-cleaved Cas3 (1:1,000; cat. no. 9664S; Cell Signaling Technology, Inc.); anti-cleaved Cas8 (1:1,000; cat. no. 9496S; Cell Signaling Technology, Inc.); anti-B-cell lymphoma 2 (Bcl-2) associated X protein (Bax; 1:1,000; cat. no. 2772S; Cell Signaling Technology, Inc.); anti-p62 (1:1,000; cat. no. 5114S; Cell Signaling Technology, Inc.); anti-Occludin (1:1,000; cat. no. 5506S; Cell Signaling Technology, Inc.); anti-zonula occludens protein 1 (ZO-1; 1:1,000; cat. no. 5406S; Cell Signaling Technology, Inc.); anti-Clauddin-5 (E8F3D; 1:1,000; cat. no. 49564; Cell Signaling Technology, Inc.); and anti-PTEN (1:1,000; cat. no. 9552S; Cell Signaling Technology, Inc.).

Techniques: Immunohistochemical staining, Staining, Marker, Western Blot, Expressing, Injection, Control

Journal: Cureus

Article Title: Sterile Alpha Motif Domain-Containing 5 Suppresses Malignant Phenotypes and Tumor Growth in Breast Cancer: Regulation of Polo-Like Kinase 1 and c-Myc Signaling in a Xenograft Model

doi: 10.7759/cureus.73259

Figure Lengend Snippet: (A) and (C) Sterile Alpha Motif Domain-Containing 5 (SAMD5) overexpression was achieved in two triple-negative breast cancer cell lines (MDA-MB-231 and HCC1937) by introducing SAMD5 overexpression (OE), which was validated using Reverse Transcription Quantitative Polymerase Chain Reaction (RT-qPCR) and immunoblotting. (B) MDA-MB-231 and HCC1937 cells were transfected with SAMD5 OE and examined for cell viability using the Cell Counting Kit-8 (CCK-8) assay. (D) Colony formation was assessed in the transfected cells. (E) Cell invasion was evaluated using the Transwell assay. (F) The protein levels of the proliferation marker Ki67 and the invasion markers Matrix Metalloproteinase 2 (MMP2) and Matrix Metalloproteinase 9 (MMP9) were determined using immunoblotting. *p < 0.05; **p < 0.01

Article Snippet: The primary antibodies used were as follows: anti-SAMD5 (STJ194894; St John’s Laboratory, London, UK), anti-Ki67 (28074-1-AP; Proteintech), anti-Matrix Metallopeptidase 2 (MMP2) (10373-2-AP; Proteintech), anti-Matrix Metallopeptidase 9 (MMP9) (82854-1-RR; Proteintech), anti-PLK1 (ab189139; Abcam), anti-β-Catenin (51067-2-AP; Proteintech), anti-Cyclin-Dependent Kinase 4 (CDK4) (11026-1-AP; Proteintech), anti-Cyclin-Dependent Kinase 6 (CDK6) (14052-1-AP; Proteintech), anti-Cyclin D1 (26939-1-AP; Proteintech), anti-Cyclin E (11935-1-AP; Proteintech), and anti-GAPDH (60004-1-Ig; Proteintech).

Techniques: Sterility, Over Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Transfection, Cell Counting, CCK-8 Assay, Transwell Assay, Marker

Journal: Cureus

Article Title: Sterile Alpha Motif Domain-Containing 5 Suppresses Malignant Phenotypes and Tumor Growth in Breast Cancer: Regulation of Polo-Like Kinase 1 and c-Myc Signaling in a Xenograft Model

doi: 10.7759/cureus.73259

Figure Lengend Snippet: (A) Polo-like Kinase 1 (PLK1) knockdown was achieved in two triple-negative breast cancer cell lines (MDA-MB-231 and HCC1937) by introducing small interfering RNA against PLK1 (siRNA-PLK1-1/2/3), which was validated using immunoblotting. si-PLK1-2 was selected for further investigations due to its higher efficiency. (B) MDA-MB-231 and HCC1937 cells were transfected with si-PLK1 and examined for cell viability using the Cell Counting Kit-8 (CCK-8) assay. (C) Colony formation was assessed in the transfected cells. (D) Cell invasion was evaluated using the Transwell assay. (E) The protein levels of proliferation marker Ki67 and invasion markers Matrix Metalloproteinase 2 (MMP2) and Matrix Metalloproteinase 9 (MMP9) were determined using immunoblotting. *p < 0.05; **p < 0.01

Article Snippet: The primary antibodies used were as follows: anti-SAMD5 (STJ194894; St John’s Laboratory, London, UK), anti-Ki67 (28074-1-AP; Proteintech), anti-Matrix Metallopeptidase 2 (MMP2) (10373-2-AP; Proteintech), anti-Matrix Metallopeptidase 9 (MMP9) (82854-1-RR; Proteintech), anti-PLK1 (ab189139; Abcam), anti-β-Catenin (51067-2-AP; Proteintech), anti-Cyclin-Dependent Kinase 4 (CDK4) (11026-1-AP; Proteintech), anti-Cyclin-Dependent Kinase 6 (CDK6) (14052-1-AP; Proteintech), anti-Cyclin D1 (26939-1-AP; Proteintech), anti-Cyclin E (11935-1-AP; Proteintech), and anti-GAPDH (60004-1-Ig; Proteintech).

Techniques: Knockdown, Small Interfering RNA, Western Blot, Transfection, Cell Counting, CCK-8 Assay, Transwell Assay, Marker

Journal: Cureus

Article Title: Sterile Alpha Motif Domain-Containing 5 Suppresses Malignant Phenotypes and Tumor Growth in Breast Cancer: Regulation of Polo-Like Kinase 1 and c-Myc Signaling in a Xenograft Model

doi: 10.7759/cureus.73259

Figure Lengend Snippet: (A) Polo-like Kinase 1 (PLK1) overexpression was achieved in two triple-negative breast cancer cell lines (MDA-MB-231 and HCC1937) by introducing PLK1 overexpression (OE), which was validated using immunoblotting. (B) MDA-MB-231 and HCC1937 cells were then co-transfected with Sterile Alpha Motif Domain-Containing 5 (SAMD5) overexpression (OE) and PLK1 OE, and the protein levels of SAMD5 were analyzed using immunoblotting. (C) Cell viability was assessed using the Cell Counting Kit-8 (CCK-8) assay. (D) Colony formation was evaluated in the transfected cells. (E) Cell invasion was measured using the Transwell assay. (F)-(G) The protein levels of proliferation marker Ki67 and invasion markers Matrix Metalloproteinase 2 (MMP2) and Matrix Metalloproteinase 9 (MMP9) were determined using immunoblotting. **p < 0.01 vs. vector group; #p < 0.05; ##p < 0.01 vs. SAMD5 group

Article Snippet: The primary antibodies used were as follows: anti-SAMD5 (STJ194894; St John’s Laboratory, London, UK), anti-Ki67 (28074-1-AP; Proteintech), anti-Matrix Metallopeptidase 2 (MMP2) (10373-2-AP; Proteintech), anti-Matrix Metallopeptidase 9 (MMP9) (82854-1-RR; Proteintech), anti-PLK1 (ab189139; Abcam), anti-β-Catenin (51067-2-AP; Proteintech), anti-Cyclin-Dependent Kinase 4 (CDK4) (11026-1-AP; Proteintech), anti-Cyclin-Dependent Kinase 6 (CDK6) (14052-1-AP; Proteintech), anti-Cyclin D1 (26939-1-AP; Proteintech), anti-Cyclin E (11935-1-AP; Proteintech), and anti-GAPDH (60004-1-Ig; Proteintech).

Techniques: Over Expression, Western Blot, Transfection, Sterility, Cell Counting, CCK-8 Assay, Transwell Assay, Marker, Plasmid Preparation