Journal: Oncotarget

Article Title: Endoplasmic reticulum chaperone prolyl 4-hydroxylase, beta polypeptide (P4HB) promotes malignant phenotypes in glioma via MAPK signaling.

doi: 10.18632/oncotarget.18026

Figure Lengend Snippet: Figure 5: Transient over-expression of P4HB promoted glioma cell proliferation, migration, invasion and tube formation ability in vitro. (A) Western blot analysis showed upregulated expression of P4HB post-transfection. (B) MTT assay was performed on cells with transient P4HB over-expression (D54 P4HB, U87 P4HB and U251 P4HB) and empty vector controls (D54 Vec, U87 Vec and U251 Vec). After 5 days incubation, cells over-expressing P4HB showed higher proliferative rates than controls. (C) Migration assay showed greater motility of U87 P4HB and U251 P4HB than their respective vector controls (U87 Vec and U251 Vec). (D) Matrigel cell invasion assay similarly showed greater invasiveness in U87 and U251 cells with P4HB over-expression (** p <0.05). (E) Angiogenesis, as measured by tube formation ability, was again higher in P4HB overexpressing cells (U87 and U251). (F) Western blot analysis revealed that P4HB over-expression was associated with increased MAPK phosphorylation. (G) Suppression of MAPK activities by U0126 reduced VEGF expression in P4HB over-expressing cells, while P4HB expression level was unaffected. (H) Representative pictures from three independent assays showed decreased cell invasion abilities of U87 P4HB and U251 P4HB cells after treatment with U0126. (I) U0126 pretreatment also inhibited tube formation in these cells after 24 h (Magnification: ×200).

Article Snippet: Briefly, after blocking with 5% non-fat milk in TBS-T (20 mM Tris, 137 mM NaCl, 0.1% Tween-20, pH 7.6), the membrane was probed with one of the following primary antibodies (at 1:1000 dilution) at 4oC overnight: rabbit monoclonal antibodies against P4HB, total p44/42 MAP kinase (Erk 1/2) (total MAPK), phosphor-p44/42 MAP kinase (Erk1/2) (Thr202/Tyr204) (pMAPK), and VEGF (all from Cell Signaling Technology Inc.).

Techniques: Over Expression, Migration, In Vitro, Western Blot, Expressing, Transfection, MTT Assay, Plasmid Preparation, Incubation, Invasion Assay, Phospho-proteomics

Journal: Frontiers in pharmacology

Article Title: Curcumin Derivative Cur20 Attenuated Cerebral Ischemic Injury by Antioxidant Effect and HIF-1α/VEGF/TFEB-Activated Angiogenesis.

doi: 10.3389/fphar.2021.648107

Figure Lengend Snippet: FIGURE 6 | Expression of HIF-1 a (A), VEGF (B), TFEB (C), and CD34 (D) in cerebral cortex analyzed by immunohistochemistry. *p < 0.05 vs. sham group, **p < 0.01 vs. sham group; #p < 0.05 vs. Model group, ##p < 0.01 vs. Model group.

Article Snippet: Brain slices were incubated with antibodies (BosterBioengineering, Wuhan, China) against HIF-1α, CD34, NF-κB, VEGF, TFEB (rabbit polyclonal IgGantibody; 1:100) at 4°C and reacted with avidin-biotin-peroxidase complex and DAB (Boster Bio-engineering, Wuhan, China).

Techniques: Expressing, Immunohistochemistry

Journal: Frontiers in pharmacology

Article Title: Curcumin Derivative Cur20 Attenuated Cerebral Ischemic Injury by Antioxidant Effect and HIF-1α/VEGF/TFEB-Activated Angiogenesis.

doi: 10.3389/fphar.2021.648107

Figure Lengend Snippet: FIGURE 9 | Effect of Cur20 on the HIF-1α/VEGF/TFEB pathway. A–E, The protein levels of HIF-1α, NF-κB, VEGF, and TFEB analyzed by western blot. F, The content and translocation of TFEB in nuclei observed by confocal microscopy with immunofluorescence. rBMECs were pretreated with Cur20 for 2 h then treated with OGD for 4 h. L, M, or H represents the concentration of Cur20 as 0.1, 1, 10 μM, respectively. In the immunofluorescence experiment, the concentration of Cur20 was 10 μM. The experimental data was expressed by mean ± SD, n 3.*p < 0.05 vs. control group, #p < 0.05 vs. Model group.

Article Snippet: Brain slices were incubated with antibodies (BosterBioengineering, Wuhan, China) against HIF-1α, CD34, NF-κB, VEGF, TFEB (rabbit polyclonal IgGantibody; 1:100) at 4°C and reacted with avidin-biotin-peroxidase complex and DAB (Boster Bio-engineering, Wuhan, China).

Techniques: Western Blot, Translocation Assay, Confocal Microscopy, Concentration Assay, Control

Journal: Cell death & disease

Article Title: B7-H3 promotes colorectal cancer angiogenesis through activating the NF-κB pathway to induce VEGFA expression.

doi: 10.1038/s41419-020-2252-3

Figure Lengend Snippet: Fig. 3 B7-H3 promoted the expression of VEGFA in CRC. a The expression of angiogenesis-related genes was detected by RT-qPCR in shB7-H3 HCT116 and RKO cells. b Western blot analysis of B7-H3 and VEGFA in the sh-NC and shB7-H3 CRC cell lines. β-actin served as a loading control. c Representative images of IHC for VEGFA in CRC tissues and matched normal tissues from the 125 clinical CRC patients. Scale bar, 100 μm. d VEGFA protein expression based on the staining index of CRC specimens and matched normal tissues. e VEGFA protein expression is shown for patients stratified into B7-H3 low (median value) groups. f Correlation analysis of the staining index of the protein expression levels of B7-H3 and CD31 in human CRC specimens (n = 125). The correlation coefficient (r) is shown. The data represent the means ± SEM. NS no significant difference; *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: After antigen retrieval with 10mM sodium citrate buffer (pH 6.0), the sections were incubated with goat anti-human 4IgB7-H3 antibody (R&D Systems, MN, USA, #AF1027, 1:100), mouse anti-human CD31 antibody (Abcam, Cambridge, MA, USA, #ab32457, 1:1500), or rabbit anti-human VEGFA antibody (Proteintech, Wuhan, China, #19003–1-AP, 1:500).

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

Journal: Cell death & disease

Article Title: B7-H3 promotes colorectal cancer angiogenesis through activating the NF-κB pathway to induce VEGFA expression.

doi: 10.1038/s41419-020-2252-3

Figure Lengend Snippet: Fig. 6 B7-H3 promoted angiogenesis via NF-κB/VEGFA pathway in Matrigel plugs in vivo. a, b CD31 (a) and VEGFA (b) protein expression based on their IHC staining index results in subcutaneous tumors formed by sh-NC-HCT116 and shB7-H3-HCT116 cells. N = 5. c, d CD31 (c) and VEGFA (d) protein expression based on their IHC staining index results in subcutaneous tumors formed by EV-HCT116 and B7-H3-HCT116 cells. N = 5. e, f CD31 (e) and VEGFA (f) protein expression based on their IHC staining index results in subcutaneous B7-H3-HCT116 tumors and B7-H3-HCT116 tumors treated with BAY11–7082 (6 mg/kg). g, h CD31 (g) and VEGFA (h) protein expression based on their IHC staining index results in subcutaneous B7-H3-HCT116 tumors and B7-H3-HCT116 tumors treated with bevacizumab (1 mg/kg). i, j CD31 (i) and VEGFA (j) protein expression based on their IHC staining index results in subcutaneous B7-H3-HCT116 tumors and B7-H3-HCT116 tumors co-treated with 3E8 (5 mg/kg) and BAY11–7082 (6 mg/kg) or bevacizumab (1 mg/kg). N = 5. The data represent the means ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: After antigen retrieval with 10mM sodium citrate buffer (pH 6.0), the sections were incubated with goat anti-human 4IgB7-H3 antibody (R&D Systems, MN, USA, #AF1027, 1:100), mouse anti-human CD31 antibody (Abcam, Cambridge, MA, USA, #ab32457, 1:1500), or rabbit anti-human VEGFA antibody (Proteintech, Wuhan, China, #19003–1-AP, 1:500).

Techniques: In Vivo, Expressing, Immunohistochemistry

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: linc00958/miR-185-5p/RSF-1 modulates cisplatin resistance and angiogenesis through AKT1/GSK3β/VEGFA pathway in cervical cancer

doi: 10.1186/s12958-022-00995-2

Figure Lengend Snippet: Antibodies used in western blot and IHC experiments

Article Snippet: The primary antibody against VEGFA (bs-4572R, Bioss, Beijing, China) was diluted with 1% donkey serum (1:200) and then added in each well.

Techniques: Western Blot