Journal: Vascular Cell

Article Title: EGCG, a major green tea catechin suppresses breast tumor angiogenesis and growth via inhibiting the activation of HIF-1α and NFκB, and VEGF expression

doi: 10.1186/2045-824X-5-9

Figure Lengend Snippet: Oral EGCG at 50–100 mg/kg/d in drinking water significantly reduced intratumoral microvessel density (Panel A: 109 ± 20 vs. 156 ± 12 microvessel #/mm^2; P < 0.01), plasma VEGF levels (Panel B; 26.48 ± 3.76 vs. 40.79 ± 3.5 pg/ml; P < 0.01), and tumor VEGF expression (Panel B; 45.72 ± 1.4 vs. 59.03 ± 3.8 pg/mg; P < 0.01) over the control, respectively in mice (n = 8). The digital images show CD31 immunohistochemistry staining in OCT-embedded cryosections of mouse breast cancer tumors obtained from a control (Figure A) or EGCG-treated (Figure A) mouse.

Article Snippet: Protein levels of VEGF in plasma, breast tumor, the heart, the limb muscle, and the medium cultured with E0771 cells were determined using mouse VEGF ELISA kits (R&D Systems, Minneapolis, MN), according to the manufacturer’s instructions.

Techniques: Clinical Proteomics, Expressing, Control, Immunohistochemistry, Staining

Journal: Vascular Cell

Article Title: EGCG, a major green tea catechin suppresses breast tumor angiogenesis and growth via inhibiting the activation of HIF-1α and NFκB, and VEGF expression

doi: 10.1186/2045-824X-5-9

Figure Lengend Snippet: EGCG at 50 μg/ml significantly inhibited VEGF expression (Panel A, 1752 ± 49 vs. 2254 ± 91 pg/mg, n = 6, P < 0.01), the activation of HIF-1α (Panel B, 0.11 ± 0.02 vs. 0.24 ± 0.02, n = 6, P < 0.01) and NFκB (Panel C, 1.15 ± 0.21 vs. 1.61 ± 0.32; n = 6, P < 0.01) in cultured E0771 cells, compared to the control, respectively.

Article Snippet: Protein levels of VEGF in plasma, breast tumor, the heart, the limb muscle, and the medium cultured with E0771 cells were determined using mouse VEGF ELISA kits (R&D Systems, Minneapolis, MN), according to the manufacturer’s instructions.

Techniques: Expressing, Activation Assay, Cell Culture, Control

Journal: Vascular Cell

Article Title: EGCG, a major green tea catechin suppresses breast tumor angiogenesis and growth via inhibiting the activation of HIF-1α and NFκB, and VEGF expression

doi: 10.1186/2045-824X-5-9

Figure Lengend Snippet: EGCG treatment did not affect the capillary density (3270 ± 162 vs. 3103 ± 226 #/mm^2; n = 8; P = 0.5215), and VEGF expression (261 ± 22 vs. 245 ± 19 pg/mg; n = 8; P = 0.4517) in the mouse heart, compared to the control group (Panel A), respectively. There was no significant difference in the capillary density (370 ± 55 vs. 381 ± 44 #/mm^2; n = 8; P = 0.5401), and VEGF expression (225 ± 16 vs. 214 ± 20 pg/mg; n = 8; P = 0.7825) in the limb skeletal muscles between the EGCG-treated mice and the control mice (Panel B ), respectively. The digital images show CD31 immunohistochemistry staining in OCT-embedded cryosections of the heart (Panel A ) and the limb muscle (Panel B ) of control mouse and EGCG-treated mouse, respectively.

Article Snippet: Protein levels of VEGF in plasma, breast tumor, the heart, the limb muscle, and the medium cultured with E0771 cells were determined using mouse VEGF ELISA kits (R&D Systems, Minneapolis, MN), according to the manufacturer’s instructions.

Techniques: Expressing, Control, Muscles, Immunohistochemistry, Staining

Journal: iScience

Article Title: Induced retinal pigment epithelial cells with anti-epithelial-to-mesenchymal transition ability delay retinal degeneration

doi: 10.1016/j.isci.2022.105050

Figure Lengend Snippet: iRPE cells have similar phenotype and functions as iPSC-RPE cells (A) Schematic for the transforming process. A cocktail of TF-expressing retroviruses was used to transfect De-iPSC-RPE cells. After seven days, iRPE clone was observed in the culture and picked out for subculturing. (B and C) RPE-specific and EMT-associated markers detected by (B) immunostaining and (C) western blotting after cells were cultured for 8 days. The expression pattern of these markers in iRPE cells is more similar to that in iPSC-RPE cells. Scale bar = 50 μm. (D) Electron micrographs of iPSC-RPE cells, De-iPSC-RPE cells, and iRPE cells demonstrated that quite a few microvilli were on the surface of iRPE and iPSC-RPE cells. Scale bar = 0.5 μm. (E and F) The bound and phagocyted POSs (pointed by arrows) in iPSC-RPE, De-iPSC-RPE, and iRPE cells (E) and quantification of phagocytosis (F) as determined by the number of bound and phagocyted POS per field. Scale bar = 50 μm. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n ≥ 5. (G and H) TER analysis (G) and HRP permeability assay (H) showed that iRPE cells maintained the same epithelial integrity as iPSC-RPE cells. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 6. (I) Expression levels of PEDF and VEGF from upper and lower chambers were determined by ELISA. iRPE cells and iPSC-RPE cells demonstrated similar secretion patterns. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 3.

Article Snippet: PEDF and VEGF were quantified by PEDF ELISA kit (Elabscience, Wuhan, China) and VEGF ELISA kit (Proteintech).

Techniques: Expressing, Subculturing Assay, Immunostaining, Western Blot, Cell Culture, Permeability, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Induced retinal pigment epithelial cells with anti-epithelial-to-mesenchymal transition ability delay retinal degeneration

doi: 10.1016/j.isci.2022.105050

Figure Lengend Snippet: BMP7 and FOXF2 are critical regulators of EMT in iRPE cells (A) The higher level of BMP7 secreted by iRPE cells compared with De-iPSC-RPE cells was determined by ELISA. Data are mean ± SD, unpaired two-sided t-tests, n = 4. (B and C) The reduced expression level of FOXF2 in iRPE compared with that in De-iPSC-RPE cells was determined by (B) immunostaining and (C) western blotting. Scale bar = 50 μm. (D) The efficiency of bmp7 knockdown was determined by qRT-PCR; shBmp7-1 was slightly more efficient at reducing the mRNA level of bmp7 than shBmp7-2; therefore, it was selected for subsequent experiments. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 4. (E) The shBmp7 construct contained the ZsGreen expression element to indicate successful transfection. Scale bar = 50 μm. (F and G) FLAG-FOXF2 overexpression (ov-FOXF2) in (F) iRPE cells (G) shBmp7-iRPE cells. Scale bar = 50 μm. (H and I) The expression levels of RPE-specific markers and EMT markers were determined by (H) western blotting and (I) quantitative analysis. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 3. (J) BMP7 expression levels in shCont-iRPE, shBmp7-iRPE, ov-FOXF2- iRPE, and shBmp7 + ov-FOXF2-iRPE. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 4. (K) The immunostaining of RPE-specific markers and EMT markers. Overexpression of FOXF2 exhibited similar effects to knockdown of bmp7 in iRPE cells by downregulating RPE markers and upregulating EMT marker. Scale bar = 50 μm.

Article Snippet: PEDF and VEGF were quantified by PEDF ELISA kit (Elabscience, Wuhan, China) and VEGF ELISA kit (Proteintech).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Immunostaining, Western Blot, Knockdown, Quantitative RT-PCR, Construct, Transfection, Over Expression, Marker

Journal: iScience

Article Title: Induced retinal pigment epithelial cells with anti-epithelial-to-mesenchymal transition ability delay retinal degeneration

doi: 10.1016/j.isci.2022.105050

Figure Lengend Snippet: Four TFs transcriptionally regulate bmp7 , foxf2 , lin7a , pard6b , and ppm1a in a direct or indirect manner (A) ELISA analysis demonstrated that BMP7 levels was reduced in 4TFs−nr2e1-RPE, 4TFs−mitf-a-RPE, 4TFs−c-myc-RPE, and 4TFs−crx-RPE cells compared with that in iRPE cells. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 4. (B and C) The levels of FOXF2, LIN7A, PARD6B, and PPM1A were determined by (B) western blotting and (C) quantitative analysis. Data are mean ± SD, one-way ANOVA and post hoc Bonferroni test, n = 3. (D) Generation of FLAG-MITF-A-iRPE, FLAG-CRX-iRPE, FLAG-NR2E1-iRPE, and FLAG-C-MYC-iRPE cells. Scale bar = 50 μm. (E–I) Enriched peaks of CRX binding to the target genes and fold enrichment of CRX immunoprecipitation compared with IgG control, as determined by qRT-PCR. Data are mean ± SD, unpaired two-sided t-tests, n = 3. (J–M) Enriched peaks of MITF-A and NR2E1 binding to the target gene lin7a and fold enrichment of MITF-A and NR2E1 immunoprecipitation compared with IgG control, as determined by qRT-PCR. Data are mean ± SD, unpaired two-sided t-tests, n = 3. (N) Schematic model for the regulation of EMT and MET processes by the four TFs. CRX, MITF-A, NR2E1, and C-MYC directly or indirectly regulated the expression of bmp7 , lin7a , pard6b, ppm1a , and foxf2 to inhibit EMT and promote MET.

Article Snippet: PEDF and VEGF were quantified by PEDF ELISA kit (Elabscience, Wuhan, China) and VEGF ELISA kit (Proteintech).

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Binding Assay, Immunoprecipitation, Control, Quantitative RT-PCR, Expressing

Journal: Advanced Functional Materials

Article Title: 3D Hybrid Nanofiber Aerogels Combining with Nanoparticles Made of a Biocleavable and Targeting Polycation and MiR‐26a for Bone Repair

doi: 10.1002/adfm.202005531

Figure Lengend Snippet: Figure 7. a,b) Relative miR-26a expression in rBMSCs mediated by Lipo2000-, PEI-, PGEA-, and HA–SS–PGEA-containing NPs, relative to the internal reference U6. c) VEGF expression in rBMSCs mediated by HA–SS–PGEA/miR-NC and HA–SS–PGEA/miR-26a at the N/P ratio of 20 after transfection for 2, 3, 4, and 5 days. d) Relative mRNA expression of GSK-3β in rBMSCs mediated by HA–SS–PGEA/miR-NC and HA–SS–PGEA/miR-26a at the N/P ratio of 20 after transfection for 2, 3, 4, and 5 days. The internal reference is GAPDH. e,f) Relative mRNA expressions of Runx2, Alp, OCN, and BSP in rBMSCs mediated by HA–SS–PGEA/miR-NC and HA–SS–PGEA/miR-26a at the N/P ratio of 20 after 7 and 21 d of culture in osteo-differentiation medium (*p < 0.05; n = 3 per group). The internal reference is GAPDH.

Article Snippet: The supernatant was collected at 2, 3, 4, and 5 d. VEGF levels in the supernatant were detected with a VEGF ELISA kit (AVIVA System Biology, San Diego, US) according to the manual.

Techniques: Expressing, Transfection

Journal: iScience

Article Title: Phosphate-induced activation of VEGFR2 leads to caspase-9-mediated apoptosis of hypertrophic chondrocytes

doi: 10.1016/j.isci.2023.107548

Figure Lengend Snippet: Pi increases VEGFA bioavailability (A) HCs were pretreated for 2 h with the pan-MMP inhibitor, MMP-V (10 μM) followed by treatment with Si (7 mM) or Pi (7 mM) for 10 min. ELISA was performed to quantitate VEGFA in the conditioned media. Data represent the mean ± SD; of that obtained from three to four independent chondrocyte preparations; p values are indicated above the line. (B) Western analyses of the treated HCs using anti-pERK1/2 & tERK1/2 antibodies. (C) Densitometric quantitation of western data in (B). Densitometric data represent mean ± SD; of that obtained from three to four independent chondrocyte preparations; p values are indicated above the line.

Article Snippet: Mouse VEGFA was quantitated in the conditioned media of hypertrophic chondrocytes using a mouse VEGFA ELISA kit (Bioss, Woburn, Massachusetts, USA Cat# BSKM1018) following the manufacturer’s protocol.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Quantitation Assay

Journal: iScience

Article Title: Phosphate-induced activation of VEGFR2 leads to caspase-9-mediated apoptosis of hypertrophic chondrocytes

doi: 10.1016/j.isci.2023.107548

Figure Lengend Snippet:

Article Snippet: Mouse VEGFA was quantitated in the conditioned media of hypertrophic chondrocytes using a mouse VEGFA ELISA kit (Bioss, Woburn, Massachusetts, USA Cat# BSKM1018) following the manufacturer’s protocol.

Techniques: Recombinant, Modification, Bicinchoninic Acid Protein Assay, Reverse Transcription, Enzyme-linked Immunosorbent Assay, In Situ, Software

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: STAT3/miR-130b-3p/MBNL1 feedback loop regulated by mTORC1 signaling promotes angiogenesis and tumor growth

doi: 10.1186/s13046-022-02513-z

Figure Lengend Snippet: miR-130b-3p promotes angiogenesis via targeting MBNL1. A-D Tsc2 − / − MEFs were infected with lentiviruses harboring control vector (Lv) or MBNL1 overexpression constructs (Lv-MBNL1). A The expression of MBNL1 was determined by western blot. B Cell culture supernatants from the cells were analyzed for VEGF-C by ELISA. (RU, relative unit). C and D The effect on angiogenesis of was detected by tube formation assay ( C , scale bar, 50 μm) and CAM assay ( D ). E–G anti-miR-130b-3p-expressing NTC/T2 null cells were infected with lentivirus harboring shRNAs targeting MBNL1 (shMBNL1-1 and shMBNL1-2) or a control shRNA (shSc). E Cell lysates were subjected to immunoblotting with the indicated antibodies. F and G The angiogenic abilities of the indicated cells were detected by tube formation assay ( F , scale bar, 50 μm) and CAM assay ( G ). Representative images (left panels) and quantifications (right panels). H–K The indicated cells were inoculated subcutaneously into nude mice ( n = 5), and tumor growth was monitored. H Tumor pictures. Scale bar, 1 cm. I Tumor volumes, J Tumor weight. K Tumor tissues were subjected to HE and IHC staining. Scale bar, 50 μm. Data indicate mean ± SD of 3–5 replicates. * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001

Article Snippet: The levels of secreted VEGF-C in cell-free supernatant of MBNL1-overexpressing cells and the control cells were quantified using a mouse VEGF-C ELISA Kit (Novus Biologicals, CO, USA) according to the manufacturer’s instructions.

Techniques: Infection, Control, Plasmid Preparation, Over Expression, Construct, Expressing, Western Blot, Cell Culture, Enzyme-linked Immunosorbent Assay, Tube Formation Assay, Chick Chorioallantoic Membrane Assay, shRNA, Immunohistochemistry