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  • 99
    Becton Dickinson matrigel
    Psoralidin inhibits CTPE xenografts tumour growth in nude mice. CTPE cells in a 50- μ l (final volume) of phosphate-buffered saline and <t>Matrigel</t> (1 : 1) were subcutaneously injected into separate flanks of mice. Mice received vehicle (sesame oil; circles) or Pso (20 mg kg −1 Bw; squares) orally for 5 days per week up to 5 weeks. Tumour volumes (mm 3 ) (upper panel) and mouse body weight (lower panel) were measured twice and once a week, respectively. Student’s t -test was used to calculate statistical significance between Pso-treated and -untreated animals at each time point. * P
    Matrigel, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 99/100, based on 96147 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/matrigel/product/Becton Dickinson
    Average 99 stars, based on 96147 article reviews
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    92
    Corning Life Sciences matrigel
    MBECs express endothelial-specific genes and display endothelial functional characteristics . (A) Primarily passaged MBECs were collected, and total RNA was isolated and prepared for reverse-transcriptase PCR analysis for expression of multiple endothelial cell genes. VE-cadherin, VWF, VEGFR-2, and PECAM (CD31) were all expressed by MBECs, confirming their endothelial cell lineage. (B) Primary MBECs (5 × 10 4 ) were plated in 24-well tissue culture plates precoated with <t>Matrigel</t> to analyze for evidence of capillary tube forming capability. As shown, MBECs readily formed tubelike structures, consistent with an endothelial cell lineage.
    Matrigel, supplied by Corning Life Sciences, used in various techniques. Bioz Stars score: 92/100, based on 12964 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson matrigel basement membrane matrix
    Effect of DMSA-Fe 2 O 3 on tube network formed by HAECs cultured on <t>Matrigel</t> within 14 h. ( a ) HAECs can form a capillary-like network on Matrigel-coated wells within 14 h. ( b ) An obvious failure to form networks by HAECs in the presence of 0.01 mg/ml DMSA-Fe 2 O 3 . ( c ) Few tube networks by HAECs in the presence of 0.02 mg/ml DMSA-Fe 2 O 3 . ( d ) The high urea solution (6M urea) was used as a positive control for the inhibition of tube formation.
    Matrigel Basement Membrane Matrix, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 4489 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/matrigel basement membrane matrix/product/Becton Dickinson
    Average 92 stars, based on 4489 article reviews
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    94
    Thermo Fisher matrigel
    Formation of tubular network by MDA-MB-231 cells was tested on <t>matrigel</t> in presence and absence of ephrinB2-Fc. Cells were treated with (a) PBS, (b) 3 μ g/mL ephrinB2-Fc and (c) 10 μ g/mL ephrinB2-Fc. Treatment with ephrinB2-Fc was expected to totally abrogate tubular formation and form aggregated colonies, yet it was surprisingly observed that tubular formation was not inhibited by ephrinB2-Fc treatment compared to PBS. Figures were inspected at 100x magnification and scale bars represent 20 μ m.
    Matrigel, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 4126 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Biocoat bd biocoat matrigel invasion chambers
    c-MET protein and mRNA levels are highly upregulated in invasive CRC daughter cell lines BD <t>BioCoat</t> <t>Matrigel</t> invasion chambers were used to isolate invasive subpopulations from a panel of CRC cells. ( A ) Migration and invasion assays of invasive subpopulations of HCT116, HKH-2 and LoVo sublines were compared with the parental cells using the quantitative xCELLigence system. ( B ) qRT-PCR assessment of MET gene expression levels in parental and invasive cell models. ( C ) Western blot analysis of pc-MET 1234/1235 and c-MET protein expression levels in parental (Par) and invasive (Inv) cell models. ( D ) HGF protein levels in the culture media of parental (Par) and invasive (Inv) subpopulations were measured by ELISA [Positive control: CCD-18Co (18Co)]. *= p
    Bd Biocoat Matrigel Invasion Chambers, supplied by Biocoat, used in various techniques. Bioz Stars score: 92/100, based on 1622 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Biocoat biocoat matrigel invasion chamber
    Role of miR-200b subfamily and miR-205 on prostate cancer proliferation and invasion ( A ) Expression levels of miR-200a, miR-200b, miR-429, miR-205, and ERG in commonly used prostate cancer cell lines, including LNCaP, VCaP, 22RV1, PC3, DU145. Mature miRNAs expression levels were measured by qRT-PCR, normalized against RNU48 internal control. ( B ) Expression of miRNAs in PC3 stable cell lines, including PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( C ) The effect of miRNAs on PC3 cell growth as determined by MTT assay. ( D ) PC3 stable cell <t>Matrigel</t> invasion assay using BD <t>Biocoat</t> Matrigel Invasion Chambers. Representative pictures of invaded cells stained with crystal violet. Four stable cell lines, PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( E ) Quantification of invasion. Invaded cells were stained with crystal violet, which was then solubilized in 1% SDS. The absorbance was measured at 595 nm. Error bars, mean ± SEM. * P
    Biocoat Matrigel Invasion Chamber, supplied by Biocoat, used in various techniques. Bioz Stars score: 93/100, based on 1550 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson matrigel coated membrane
    Invasion, attachment, and proliferation assay of glioblastoma cells . All assays were performed with four different glioblastoma cell lines: U87, U251, U373, and C6 (Top to bottom). Values in (A) and (B) are expressed as a percentage of corresponding parental line data. (A) Invasion: IM-3 subpopulations were compared to their parental lines. (B) Attachment: Cells adherent to <t>Matrigel</t> ® after 4 h were stained with hematoxylin QS and counted. (C) Proliferation: The rate of cell proliferation was evaluated with CellTiter 96 Aqueous One Solution (Promega) at five different time points, 4 h, 24 h, 48 h, 72 h and 96 h after seeding cells. Triangles - IM-3 lines, squares - parental. Data are represented as means ± SD (n = 3). P, parental cells; IM3, IM3 cells; IM3, IM3 cell line; **, p
    Matrigel Coated Membrane, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 1031 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Becton Dickinson phosphate buffered saline matrigel
    Establishment of mouse prostate organoid cultures A) Overview of the isolation of the prostate from the mouse urogenital system (for a detailed isolation protocol see 25 ). The procedure refers to pictures I – IX. B) Schematic representation of the anatomy of the mouse prostate. AP, anterior prostate; DLP, dorsolateral prostate; VP, ventral prostate. C) Example of how to plate the <t>matrigel</t> disc in a well of a tissue culture plate. D) Representative pictures of organoids growing from mouse prostate tissue 1 and 7 days after plating. Scale bars, 100 micron.
    Phosphate Buffered Saline Matrigel, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 88/100, based on 44 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Corning Life Sciences matrigel basement membrane matrix
    Generation of induced pluripotent stem cells (iPSCs) and retinal pigment epithelium cells (RPE) from RP2 R120X fibroblasts. ( A ) Schematic representation of iPSC and RPE cell generation from fibroblasts. A skin biopsy was taken and cut into small pieces using scalpel blades. The cells were then cultured in DMEM and outgrowing fibroblasts from the skin biopsy were isolated manually. To reprogram fibroblasts into iPSCs, cells were electroporated with plasmids containing Oct4, Sox2, Myc, Lin28 and Klf4, plated onto <t>matrigel-coated</t> dishes and cultured in mTESR medium. Forming iPSC colonies were isolated manually. iPSC were differentiated into RPE cells by culturing in HESC—bFGF medium. Pigmented RPE colonies were excised manually using crescent blades. ( B ) iPSC derived RPE from controls and the RP2 R120X patient showed nuclear (DAPI, blue) localization of the iPSC markers Nanog (red) and Oct4 (red). Scale bar 10 μm. ( C ) Differentiated control and R120X iPSC-derived RPE cells display the classic cobblestone pigmented morphology. Scale bar 10 μm. ( D ) Sectioned RPE cells were stained for RPE and differentiation markers: ATP1B1 (green), Col4 (red), Bestrophin (green), MerTK (red), Pmel17 (green), ZO1 (red), mitf (green) or ( E ) RPE65 (green). Scale bar 10 μm ( F ) RP2 protein is undetectable in iPSC-derived R120X RPE cells at the plasma membrane and the basal body by ICC. Scale bar 10 μm. ( G ) RP2 protein is undetectable in the iPSC-derived R120X RPE cells by western blotting. GAPDH was used as a loading control. ( H ) Sectioned RPE cells were stained for endogenous Gβ1 (green) the membrane marker wheat-germ agglutinin (WGA, red) and DAPI (blue). Gβ1 localizes to the plasma membrane in control RPE cells, but shows a diffuse cytoplasmic staining pattern in R120X RPE cells. Scale bar 10 μm.
    Matrigel Basement Membrane Matrix, supplied by Corning Life Sciences, used in various techniques. Bioz Stars score: 92/100, based on 464 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson phenol red free matrigel
    CXCL12 (SDF-1α) enhances granulation tissue formation that mimics the stromal reaction and angiogenesis in a <t>Matrigel</t> implantation model. A: Typical appearance of stromal formation and angiogenesis formed around a Matrigel. Scale bars = 10 mm.
    Phenol Red Free Matrigel, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 547 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phenol red free matrigel/product/Becton Dickinson
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    91
    Becton Dickinson matrigel solution
    LINC00152 promotes migration and invasion of TSCC cells. Transwell chambers with or without <t>Matrigel</t> were used to detect the migration and invasion of TSCC CAL-27 and SCC-9 cells as indicated (n=3). Magnification, ×40. Data are presented as the mean ± standard deviation. **P
    Matrigel Solution, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 91/100, based on 603 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Charles River Laboratories matrigel
    CCL7 induces migration and invasion of colon cancer cells via CCR3 A. Quantitation of E-cadherin expression on the surface of HCT116 cells treated with or without recombinant CCL7 (200 ng/ml) using FACS analysis. B. Expression of E-cadherin, N-cadherin, and vimentin in HCT116 cells stably transfected with GFP or CCL7 were measured by western blotting (left panel). Expression of CCL7 and EMT markers in negative siRNA control-treated or CCL7 specific siRNAs-treated HCT116 cell extracts (right panel). Actin was used as a loading control. C. A wound healing assay was performed by creating a wound on a confluent monolayer of stable GFP/CCL7 overexpressing cells using l-Dish 35-mm high culture inserts. D. Transwell <t>matrigel</t> invasion assays of HCT116 cells after transfection with 100 nM of negative siRNA control or CCL7 specific siRNAs. E. Cell migration and F. Invasion was measured using a trans-well migration chamber (left panels). Mean fluorescence intensity (MFI) of invaded area is presented in bar graphs (right panels). G. HCT116 cells were treated with 20 nM CCR3 inhibitor SB 328437. Representative images of invaded cells are shown (left panels). MFI values are presented in bar graphs (right panels). H. Expression of EMT markers in HCT116 cells stably transfected with GFP or CCL7 with or without treatment with 20 nM SB 328437 (CCR3 inhibitor) was measured by western blotting. Actin was used as a loading control. Columns: means ± SEs. ** P
    Matrigel, supplied by Charles River Laboratories, used in various techniques. Bioz Stars score: 92/100, based on 720 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Psoralidin inhibits CTPE xenografts tumour growth in nude mice. CTPE cells in a 50- μ l (final volume) of phosphate-buffered saline and Matrigel (1 : 1) were subcutaneously injected into separate flanks of mice. Mice received vehicle (sesame oil; circles) or Pso (20 mg kg −1 Bw; squares) orally for 5 days per week up to 5 weeks. Tumour volumes (mm 3 ) (upper panel) and mouse body weight (lower panel) were measured twice and once a week, respectively. Student’s t -test was used to calculate statistical significance between Pso-treated and -untreated animals at each time point. * P

    Journal: British Journal of Cancer

    Article Title: Inhibition of autophagy prevents cadmium-induced prostate carcinogenesis

    doi: 10.1038/bjc.2017.143

    Figure Lengend Snippet: Psoralidin inhibits CTPE xenografts tumour growth in nude mice. CTPE cells in a 50- μ l (final volume) of phosphate-buffered saline and Matrigel (1 : 1) were subcutaneously injected into separate flanks of mice. Mice received vehicle (sesame oil; circles) or Pso (20 mg kg −1 Bw; squares) orally for 5 days per week up to 5 weeks. Tumour volumes (mm 3 ) (upper panel) and mouse body weight (lower panel) were measured twice and once a week, respectively. Student’s t -test was used to calculate statistical significance between Pso-treated and -untreated animals at each time point. * P

    Article Snippet: Invasion assays The effects of Pso on the invasive properties were determined using Boyden chambers equipped with 8-μ m pore size polyethylene terephthalate membranes coated with Matrigel (BD Biosciences, San Jose, CA, USA).

    Techniques: Mouse Assay, Injection

    3D culture and marker analysis on structures generated by LA7 cells. Panels ( a , b ): LA7-single cell generated tubules in collagen gel ( a ); tubules containing lumen, upper left part of the panel ( b ). Cells were stained with the Hoechst nuclear dye and analyzed with confocal microscopy ( b ). Panel ( c ): LA7-single cell generated cyst in Matrigel. Panel ( d ): cells collected from LA7-derived tubules express luminal (K18), alveolar (β-casein) and myoepithelial (K14) markers. Lane 1: undifferentiated LA7 cells, lane 2: cells derived from a single LA7-generated tubules

    Journal: Cytotechnology

    Article Title: A rat mammary gland cancer cell with stem cell properties of self-renewal and multi-lineage differentiation

    doi: 10.1007/s10616-008-9173-9

    Figure Lengend Snippet: 3D culture and marker analysis on structures generated by LA7 cells. Panels ( a , b ): LA7-single cell generated tubules in collagen gel ( a ); tubules containing lumen, upper left part of the panel ( b ). Cells were stained with the Hoechst nuclear dye and analyzed with confocal microscopy ( b ). Panel ( c ): LA7-single cell generated cyst in Matrigel. Panel ( d ): cells collected from LA7-derived tubules express luminal (K18), alveolar (β-casein) and myoepithelial (K14) markers. Lane 1: undifferentiated LA7 cells, lane 2: cells derived from a single LA7-generated tubules

    Article Snippet: Matrigel preparation LA7 cells were embedded in Growth Factor Reduced Matrigel (Becton Dickinson).

    Techniques: Marker, Generated, Staining, Confocal Microscopy, Derivative Assay

    Effects of KA on HUVECs Differentiation . Effect of KA on matrigel tube formation of endothelial cells. Cells treated with (A) 1% ethanol (B) 10 μg/ml (C) 20 μg/ml and (D) 30 μg/ml (E) 40 μg/ml and (F) 100 μg/ml suramin as a positive control (4X). (G) The Dose response relationship of KA on rat aorta assay. The inhibitory effect was found to be in dose response manner. Data was represented as mean ± SD (n = 3). *** P

    Journal: Cancer Cell International

    Article Title: Antiangiogenic properties of Koetjapic acid, a natural triterpene isolated from Sandoricum koetjaoe Merr

    doi: 10.1186/1475-2867-11-12

    Figure Lengend Snippet: Effects of KA on HUVECs Differentiation . Effect of KA on matrigel tube formation of endothelial cells. Cells treated with (A) 1% ethanol (B) 10 μg/ml (C) 20 μg/ml and (D) 30 μg/ml (E) 40 μg/ml and (F) 100 μg/ml suramin as a positive control (4X). (G) The Dose response relationship of KA on rat aorta assay. The inhibitory effect was found to be in dose response manner. Data was represented as mean ± SD (n = 3). *** P

    Article Snippet: Matrigel matrix (10 mg/ml) was purchased from BD Bioscience, USA.

    Techniques: Positive Control

    In vitro sequential selection of invasive populations from MCF-7 cells using Matrigel Invasion Chambers . (A and B) In each cycle, cells invading the Matrigel were fixed with 10% formalin-neutralized buffer and stained with crystal violet on the underside of the chamber (A), and then the numbers of invading cells were counted under a microscope (B). Data are the means ± SD ( n = 3). Statistical analysis was performed by Student's t test. *, P

    Journal: BMC Cancer

    Article Title: Nuclear ?-catenin and CD44 upregulation characterize invasive cell populations in non-aggressive MCF-7 breast cancer cells

    doi: 10.1186/1471-2407-10-414

    Figure Lengend Snippet: In vitro sequential selection of invasive populations from MCF-7 cells using Matrigel Invasion Chambers . (A and B) In each cycle, cells invading the Matrigel were fixed with 10% formalin-neutralized buffer and stained with crystal violet on the underside of the chamber (A), and then the numbers of invading cells were counted under a microscope (B). Data are the means ± SD ( n = 3). Statistical analysis was performed by Student's t test. *, P

    Article Snippet: In vitro sequential selection of invasive populations from MCF-7 cells BD BioCoat Matrigel Invasion Chambers (for a 6-well plate; BD Biosciences, San Jose, CA) were used to select invasive populations from MCF-7 cells.

    Techniques: In Vitro, Selection, Staining, Microscopy

    KSHV miRNAs mediate KSHV-induced angiogenesis by activating the AKT pathway. (A) The miRNA cluster was required for KSHV-induced angiogenesis. Capillary tube formation activity of uninfected MSCa (Mock), LTC-KMSCa, LTC-ΔmiRMSCa, and LTC-ΔmiRMSCa complemented with vector control or the miRNA cluster. Equal numbers of cells were seeded on top of the Matrigel. Images of tube formation were captured at 8 h postseeding (left panel). Magnification, ×100. The average number of branches per field of view ± SD at ×40 magnification was quantified (right panel). WT, wild type. (B) LY294002 inhibited capillary tube formation of LTC-KMSCa. LTC-KMSCa were incubated with different concentrations of LY294002, and tube formation was analyzed at 8 h postseeding. Representative images (left panel) and the average number of branches per field of view ± SD (right panel) are shown. (C) AKT phosphorylation at T308 but not S473 was enhanced in LTC-KMSCa (KSHV) compared to uninfected MSCa (Mock). (D) LY294002 suppressed AKT phosphorylation at T308 in LTC-KMSCa in a dose-dependent manner. (E) The miRNA cluster was required for AKT phosphorylation at T308. AKT phosphorylation was examined by Western blotting in uninfected MSCa (Mock), LTC-KMSCa (KSHV), or LTC-ΔmiRMSCa (ΔmiR) complemented with vector control (vector) or the miRNA cluster (miR cluster). *, P

    Journal: mBio

    Article Title: Human Mesenchymal Stem Cells of Diverse Origins Support Persistent Infection with Kaposi’s Sarcoma-Associated Herpesvirus and Manifest Distinct Angiogenic, Invasive, and Transforming Phenotypes

    doi: 10.1128/mBio.02109-15

    Figure Lengend Snippet: KSHV miRNAs mediate KSHV-induced angiogenesis by activating the AKT pathway. (A) The miRNA cluster was required for KSHV-induced angiogenesis. Capillary tube formation activity of uninfected MSCa (Mock), LTC-KMSCa, LTC-ΔmiRMSCa, and LTC-ΔmiRMSCa complemented with vector control or the miRNA cluster. Equal numbers of cells were seeded on top of the Matrigel. Images of tube formation were captured at 8 h postseeding (left panel). Magnification, ×100. The average number of branches per field of view ± SD at ×40 magnification was quantified (right panel). WT, wild type. (B) LY294002 inhibited capillary tube formation of LTC-KMSCa. LTC-KMSCa were incubated with different concentrations of LY294002, and tube formation was analyzed at 8 h postseeding. Representative images (left panel) and the average number of branches per field of view ± SD (right panel) are shown. (C) AKT phosphorylation at T308 but not S473 was enhanced in LTC-KMSCa (KSHV) compared to uninfected MSCa (Mock). (D) LY294002 suppressed AKT phosphorylation at T308 in LTC-KMSCa in a dose-dependent manner. (E) The miRNA cluster was required for AKT phosphorylation at T308. AKT phosphorylation was examined by Western blotting in uninfected MSCa (Mock), LTC-KMSCa (KSHV), or LTC-ΔmiRMSCa (ΔmiR) complemented with vector control (vector) or the miRNA cluster (miR cluster). *, P

    Article Snippet: The migration and invasion assay was performed using a 24-well chamber system with a Matrigel-coated or noncoated membrane according to the manufacturer’s instructions (BD Biosciences).

    Techniques: Activity Assay, Plasmid Preparation, Incubation, Western Blot

    LTC-KMSCs manifest distinct angiogenic, migratory, invasive, and transforming phenotypes. (A) Capillary tube formation of LTC-KMSCs. Equal numbers of uninfected MSCs (Mock) or KMSCs (KSHV) were applied to the top of the Matrigel, and images of tube formation were captured using a phase-contrast microscope at 8 h postseeding (upper panel). Magnification, ×100. An in vitro angiogenic index from the average number of branches per field of view ± SD at ×40 magnification is shown (lower panel). (B and C) Migration and invasiveness of LTC-KMSCs. The migrating (B) or invaded (C) cells were stained with calcein AM. Representative images are shown (upper panel), and the averages ± SDs were calculated based on quantification using a fluorescence microplate reader with 485-nm excitation and 528-nm emission filters. RFU, relative fluorescence units. (D) Colony formation of uninfected MSCs and LTC-KMSCs. Representative cell colonies in soft agar are shown. Colony formation was visualized at day 18 postseeding. Bar, 50 µm. *, P

    Journal: mBio

    Article Title: Human Mesenchymal Stem Cells of Diverse Origins Support Persistent Infection with Kaposi’s Sarcoma-Associated Herpesvirus and Manifest Distinct Angiogenic, Invasive, and Transforming Phenotypes

    doi: 10.1128/mBio.02109-15

    Figure Lengend Snippet: LTC-KMSCs manifest distinct angiogenic, migratory, invasive, and transforming phenotypes. (A) Capillary tube formation of LTC-KMSCs. Equal numbers of uninfected MSCs (Mock) or KMSCs (KSHV) were applied to the top of the Matrigel, and images of tube formation were captured using a phase-contrast microscope at 8 h postseeding (upper panel). Magnification, ×100. An in vitro angiogenic index from the average number of branches per field of view ± SD at ×40 magnification is shown (lower panel). (B and C) Migration and invasiveness of LTC-KMSCs. The migrating (B) or invaded (C) cells were stained with calcein AM. Representative images are shown (upper panel), and the averages ± SDs were calculated based on quantification using a fluorescence microplate reader with 485-nm excitation and 528-nm emission filters. RFU, relative fluorescence units. (D) Colony formation of uninfected MSCs and LTC-KMSCs. Representative cell colonies in soft agar are shown. Colony formation was visualized at day 18 postseeding. Bar, 50 µm. *, P

    Article Snippet: The migration and invasion assay was performed using a 24-well chamber system with a Matrigel-coated or noncoated membrane according to the manufacturer’s instructions (BD Biosciences).

    Techniques: Microscopy, In Vitro, Migration, Staining, Fluorescence

    Actin cytoskeletal and focal adhesion organization in MDA-MB-231 cells invading in CIA. (A) Cells in wound healing assay without Matrigel on 2D surface and cells in CIA with Matrigel overlay were fixed and stained for actin (green), focal adhesion marker phospho-paxillin (red) and DNA (blue). Z-stack confocal images were captured and cell side views are shown to indicate positions of FA/FCs. White arrowheads indicate adhesion complexes. (B) Cells invading in CIA were fixed and stained with actin (red), focal adhesion marker vinculin (green) and DNA (blue). Z-stack confocal images were captured and cell side views are shown to indicate positions of FA/FCs. White arrowheads indicate adhesion complexes. (C) Quantification of adhesion complexes (puncta stained with phospho-paxillin) at the bottom of the cells (within 1 µm range above the glass) and on the cell body or on top of the cells (above 1 µm) under both conditions. All error bars indicate means ± SD; **, P

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: Actin cytoskeletal and focal adhesion organization in MDA-MB-231 cells invading in CIA. (A) Cells in wound healing assay without Matrigel on 2D surface and cells in CIA with Matrigel overlay were fixed and stained for actin (green), focal adhesion marker phospho-paxillin (red) and DNA (blue). Z-stack confocal images were captured and cell side views are shown to indicate positions of FA/FCs. White arrowheads indicate adhesion complexes. (B) Cells invading in CIA were fixed and stained with actin (red), focal adhesion marker vinculin (green) and DNA (blue). Z-stack confocal images were captured and cell side views are shown to indicate positions of FA/FCs. White arrowheads indicate adhesion complexes. (C) Quantification of adhesion complexes (puncta stained with phospho-paxillin) at the bottom of the cells (within 1 µm range above the glass) and on the cell body or on top of the cells (above 1 µm) under both conditions. All error bars indicate means ± SD; **, P

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Multiple Displacement Amplification, Wound Healing Assay, Staining, Marker

    MDA-MB-231 cells form collectively invading chains in CIA. (A) Photos from a time-lapse video of MDA-MB-231 cells invading into Matrigel in CIA showing the formation of long invasion chains at the invading wound edge. Scale bar 50 µm. (B) Cells in a large finger-like chain at the leading edge of a CIA are shown fixed and stained with actin (red), and DNA (blue). Scale bar 20 µm. (C) Actin (red), N-WASP (green) and DAPI (blue) staining of a cell chain at the front of the invading area. White arrowheads indicate puncta of N-WASP co-localizing with filamentous actin. Scale bar 20 µm. (D) Image sequence showing an invading cell actively remodeling the matrix and generating what appear as micro-tunnels (white arrow). Scale bar 20 µm. (E) Staining of actin (red) and DNA (blue) of cell invasion chains in inverted invasion assay and visualized by confocal microscopy. Image showing collective cell invasion chains on single plane (left panel) and the side view of z-stack 3D projection (right panel). Scale bar 20 µm. See also Movies S1 , S2 , S3 .

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: MDA-MB-231 cells form collectively invading chains in CIA. (A) Photos from a time-lapse video of MDA-MB-231 cells invading into Matrigel in CIA showing the formation of long invasion chains at the invading wound edge. Scale bar 50 µm. (B) Cells in a large finger-like chain at the leading edge of a CIA are shown fixed and stained with actin (red), and DNA (blue). Scale bar 20 µm. (C) Actin (red), N-WASP (green) and DAPI (blue) staining of a cell chain at the front of the invading area. White arrowheads indicate puncta of N-WASP co-localizing with filamentous actin. Scale bar 20 µm. (D) Image sequence showing an invading cell actively remodeling the matrix and generating what appear as micro-tunnels (white arrow). Scale bar 20 µm. (E) Staining of actin (red) and DNA (blue) of cell invasion chains in inverted invasion assay and visualized by confocal microscopy. Image showing collective cell invasion chains on single plane (left panel) and the side view of z-stack 3D projection (right panel). Scale bar 20 µm. See also Movies S1 , S2 , S3 .

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Multiple Displacement Amplification, Staining, Sequencing, Invasion Assay, Confocal Microscopy

    Cell invasion in CIA with Matrigel is an MMP dependent process. (A) Addition of 25 µM GM6001 significantly impairs MDA-MB-231 cell invasion in CIA speed and progressed area (arbitrary units) versus time are shown in the graphs. Scale bar 50 µm. (B) siRNA knockdown of MT1-MMP significantly impairs invasion in CIA. Western blot shows a representative knockdown of MT1-MMP. Movies were analyzed in three independent experiments. All error bars indicate means ± SD; **, P

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: Cell invasion in CIA with Matrigel is an MMP dependent process. (A) Addition of 25 µM GM6001 significantly impairs MDA-MB-231 cell invasion in CIA speed and progressed area (arbitrary units) versus time are shown in the graphs. Scale bar 50 µm. (B) siRNA knockdown of MT1-MMP significantly impairs invasion in CIA. Western blot shows a representative knockdown of MT1-MMP. Movies were analyzed in three independent experiments. All error bars indicate means ± SD; **, P

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Multiple Displacement Amplification, Western Blot

    Invadopodia structures in CIA have degradation ability. MDA-MB-231 cells in the CIA assay, showing (A) Cherry-MT1-MMP (red) containing vesicles are delivered to invadopodia structures marked by GFP-N-WASP (green) in CIA. Scale bar 20 µm in the upper panel and 10 µm in the lower panel. (B) DQ collagen is mixed with Matrigel and overlaid on top of the cells in CIA. DQ collagen fluorescence (green) is visualized around some of the actin-rich puncta (blue) with co-localization of N-WASP (red), indicating that they are invadopodia structures in CIA. Scale bar 20 µm.

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: Invadopodia structures in CIA have degradation ability. MDA-MB-231 cells in the CIA assay, showing (A) Cherry-MT1-MMP (red) containing vesicles are delivered to invadopodia structures marked by GFP-N-WASP (green) in CIA. Scale bar 20 µm in the upper panel and 10 µm in the lower panel. (B) DQ collagen is mixed with Matrigel and overlaid on top of the cells in CIA. DQ collagen fluorescence (green) is visualized around some of the actin-rich puncta (blue) with co-localization of N-WASP (red), indicating that they are invadopodia structures in CIA. Scale bar 20 µm.

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Multiple Displacement Amplification, Fluorescence

    Actin-rich puncta containing cortactin, Arp2/3 complex and N-WASP are apparent in CIA. (A–C) Cells invading under Matrigel in CIA are fixed and stained with proteins previously localized to invadopodia, cortactin (red), N-WASP (green), Arp2/3 component p34-Arc (green) and actin (blue) in a number of cell lines including MDA-MB-231 (A), CHL1 (B), HT1080 (C). (D) Invadopodia in CIA localize at the front of the invading pseudopods (white arrowhead) and branching sites of the pseudopods (blue arrowhead). Quantification shows there are more invadopodia in the front half of the cells than the rear half. Cells were analyzed in three independent experiments. All error bars indicate means ± SD; **, P

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: Actin-rich puncta containing cortactin, Arp2/3 complex and N-WASP are apparent in CIA. (A–C) Cells invading under Matrigel in CIA are fixed and stained with proteins previously localized to invadopodia, cortactin (red), N-WASP (green), Arp2/3 component p34-Arc (green) and actin (blue) in a number of cell lines including MDA-MB-231 (A), CHL1 (B), HT1080 (C). (D) Invadopodia in CIA localize at the front of the invading pseudopods (white arrowhead) and branching sites of the pseudopods (blue arrowhead). Quantification shows there are more invadopodia in the front half of the cells than the rear half. Cells were analyzed in three independent experiments. All error bars indicate means ± SD; **, P

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Staining, Multiple Displacement Amplification

    Invadopodia protrude into the matrix in CIA. (A) MDA-MB-231 cells completely embedded in collagen I also show similar elongated morphology as in CIA. Staining of N-WASP or cortactin (green) and actin (red) show that N-WASP and filamentous actin containing structures resembling invadopodia (white arrowheads) are also present in cells migrating in a pure 3D environment. Scale bar 20 µm. (B) Z-stack projection showing invadopodia-like structures (arrowheads) localize to various positions, including the front of the invading pseudopods facing upward into the Matrigel and at the periphery and dorsal surface. Staining shows N-WASP (green), actin (red) and DAPI (blue) or cortactin (blue). See also Movie S8.

    Journal: PLoS ONE

    Article Title: Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

    doi: 10.1371/journal.pone.0030605

    Figure Lengend Snippet: Invadopodia protrude into the matrix in CIA. (A) MDA-MB-231 cells completely embedded in collagen I also show similar elongated morphology as in CIA. Staining of N-WASP or cortactin (green) and actin (red) show that N-WASP and filamentous actin containing structures resembling invadopodia (white arrowheads) are also present in cells migrating in a pure 3D environment. Scale bar 20 µm. (B) Z-stack projection showing invadopodia-like structures (arrowheads) localize to various positions, including the front of the invading pseudopods facing upward into the Matrigel and at the periphery and dorsal surface. Staining shows N-WASP (green), actin (red) and DAPI (blue) or cortactin (blue). See also Movie S8.

    Article Snippet: After cells adhere, the stopper is removed and 250 µl of 50% BD Matrigel™ (4.5 mg/ml) in PBS was overlaid onto the cell monolayer seeded in the inner circle of the dish to create a matrix barrier (0.8 mm high) against the cellular surface and allowed to polymerize for 2 hours prior to adding growth medium on the top of the set Matrigel.

    Techniques: Multiple Displacement Amplification, Staining

    Rhus coriaria inhibits the invasion, downregulates MMP-9 and prostaglandin E 2 (PgE2). ( A ) MDA-MB-231 cells were incubated for 24 h with or without RCE (10 and 50 μg/mL). Cells that invaded into the matrigel were scored as described in Materials and Methods. DAPI-stained nuclei of invading cells were photographed at X 100 magnification under an inverted microscope (Nikon Ti-U, Nikon). ( B ) Quantification of invaded MDA-MB-231 into the matrigel. Values represented in percent were calculated from three independent experiments and are represented as mean ± SEM. Statistical analysis was performed using one-way ANOVA ( *p

    Journal: Scientific Reports

    Article Title: Rhus coriaria suppresses angiogenesis, metastasis and tumor growth of breast cancer through inhibition of STAT3, NFκB and nitric oxide pathways

    doi: 10.1038/srep21144

    Figure Lengend Snippet: Rhus coriaria inhibits the invasion, downregulates MMP-9 and prostaglandin E 2 (PgE2). ( A ) MDA-MB-231 cells were incubated for 24 h with or without RCE (10 and 50 μg/mL). Cells that invaded into the matrigel were scored as described in Materials and Methods. DAPI-stained nuclei of invading cells were photographed at X 100 magnification under an inverted microscope (Nikon Ti-U, Nikon). ( B ) Quantification of invaded MDA-MB-231 into the matrigel. Values represented in percent were calculated from three independent experiments and are represented as mean ± SEM. Statistical analysis was performed using one-way ANOVA ( *p

    Article Snippet: Matrigel invasion assays Invasiveness of the MDA-MB-231 cells treated with the RCE was tested using BD Matrigel Invasion Chamber (8-μm pore size; BD Biosciences, Bedfrord, MA, USA) according to manufacturer’s instructions.

    Techniques: Multiple Displacement Amplification, Incubation, Staining, Inverted Microscopy

    Effect of Rhus coriaria on the formation of capillary-like structures by HUVECs in vitro. ( A ) Patterns of angiogenesis induced by human umbilical vein endothelial cells (HUVECs) cultured on Matrigel matrix in 96-well plates with or without RCE. ( B ) Quantification of tubular morphogenesis induced in HUVECs cells cultured with or without RCE. Tube formation was determined by the length of tube-like structures containing connected cells. LY (20 μM) was used as positive controls for angiogenesis inhibition in this assay. Data are mean ± S.E.M. from three independent experiments. Statistical analysis was performed using one-way ANOVA ( *p

    Journal: Scientific Reports

    Article Title: Rhus coriaria suppresses angiogenesis, metastasis and tumor growth of breast cancer through inhibition of STAT3, NFκB and nitric oxide pathways

    doi: 10.1038/srep21144

    Figure Lengend Snippet: Effect of Rhus coriaria on the formation of capillary-like structures by HUVECs in vitro. ( A ) Patterns of angiogenesis induced by human umbilical vein endothelial cells (HUVECs) cultured on Matrigel matrix in 96-well plates with or without RCE. ( B ) Quantification of tubular morphogenesis induced in HUVECs cells cultured with or without RCE. Tube formation was determined by the length of tube-like structures containing connected cells. LY (20 μM) was used as positive controls for angiogenesis inhibition in this assay. Data are mean ± S.E.M. from three independent experiments. Statistical analysis was performed using one-way ANOVA ( *p

    Article Snippet: Matrigel invasion assays Invasiveness of the MDA-MB-231 cells treated with the RCE was tested using BD Matrigel Invasion Chamber (8-μm pore size; BD Biosciences, Bedfrord, MA, USA) according to manufacturer’s instructions.

    Techniques: In Vitro, Cell Culture, Inhibition

    Exosomes from hypoxia increases microvascular tube formation in a dose-dependent manner. hPMEC were incubated in Matrigel in absence or presence of different exosomal protein concentration from pMSC exposed to 1%, 3% or 8% O 2 . (A) Quantitative analysis of the total tube formation. (B) Concentration response from data in A. insert: half-maximal stimulatory concentration (SC 50 ) at 16 h. Values are mean ± SEM. In A, * *p

    Journal: PLoS ONE

    Article Title: Exosomal Signaling during Hypoxia Mediates Microvascular Endothelial Cell Migration and Vasculogenesis

    doi: 10.1371/journal.pone.0068451

    Figure Lengend Snippet: Exosomes from hypoxia increases microvascular tube formation in a dose-dependent manner. hPMEC were incubated in Matrigel in absence or presence of different exosomal protein concentration from pMSC exposed to 1%, 3% or 8% O 2 . (A) Quantitative analysis of the total tube formation. (B) Concentration response from data in A. insert: half-maximal stimulatory concentration (SC 50 ) at 16 h. Values are mean ± SEM. In A, * *p

    Article Snippet: For the tube formation assay, 48-well culture plates on ice were incubated with 144 µl of chilled BD Matrigel matrix (10 mg/ml) per well at 37°C for 60 min. hPMEC (6×104 ) were resuspended in culture medium with the indicated concentration of pMSC-derived exosomes (5, 10 or 20 µg/ml) and incubated for up to 24 h at 37°C.

    Techniques: Incubation, Protein Concentration, Concentration Assay

    Effect of Jagged1 siRNA on AngII-induced impairment of VEGF production and capillary-like tube formation of CMVECs. ( A ) VEGF production. Cultured CMVECs were treated as described under Figure 5B . Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 3 independent experiments (n=3). ( B ) Capillary-like tube formation. CMVECs transfected with #2 siRNA sequence of Jagged1 (si-Jag1) or scramble RNA (Scram) were seeded onto the matrigel in plates containing 1µM AngII or not. Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Representative photographs are shown (scale bar: 100µm). ( C ) Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). * p

    Journal: PLoS ONE

    Article Title: Regulation of p53 by Jagged1 Contributes to Angiotensin II-Induced Impairment of Myocardial Angiogenesis

    doi: 10.1371/journal.pone.0076529

    Figure Lengend Snippet: Effect of Jagged1 siRNA on AngII-induced impairment of VEGF production and capillary-like tube formation of CMVECs. ( A ) VEGF production. Cultured CMVECs were treated as described under Figure 5B . Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 3 independent experiments (n=3). ( B ) Capillary-like tube formation. CMVECs transfected with #2 siRNA sequence of Jagged1 (si-Jag1) or scramble RNA (Scram) were seeded onto the matrigel in plates containing 1µM AngII or not. Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Representative photographs are shown (scale bar: 100µm). ( C ) Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). * p

    Article Snippet: Capillary-like tube formation 200 µl growth factor-reduced Matrigel matrix (BD) was homogenized, layered into a 24-well plate on a cooled planar surface and allowed to solidify at 37 °C [ ].

    Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay, Transfection, Sequencing, Microscopy

    AngII-induced impairment of angiogenetic responses and accumulation and phosphorylation of p53 in cultured CMVECs. ( A ) Formation of capillary-like tubes. CMVECs were seeded onto the matrigel in plates containing 1 µM AngII (AngII) or vehicle (Control). Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Left panel : Representative photomicrographs of capillary-like tubes (scale bar: 100 µm). Right panel : Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 15 independent experiments (n=15). ( B ) Measurement of VEGF protein levels in culture medium. Cultured CMVECs were incubated with AngII (1 µM) or vehicle (Control) for 18 hours. Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). ( C ) Cytosolic p53 and p-p53 protein expression. CMVECs were treated as described under (B) and then collected and lysed for Western blot analyses of p53 and p-p53. β-actin was used as a loading control. ( D ) Nucleus p-p53 and Hif-1α expression. Nucleus proteins isolated from CMVECs treated as described under (B) were subjected to Western blot analyses for p-p53 and Hif-1α. TFIIB was used as a loading control. Representative immunoblots are shown. The expression of p53, p-p53 or Hif-1α was quantified as folds of β-actin, p53 or TFIIB, respectively. Data represent mean ± S.E.M. obtained from 3 independent experiments (n=3). * p

    Journal: PLoS ONE

    Article Title: Regulation of p53 by Jagged1 Contributes to Angiotensin II-Induced Impairment of Myocardial Angiogenesis

    doi: 10.1371/journal.pone.0076529

    Figure Lengend Snippet: AngII-induced impairment of angiogenetic responses and accumulation and phosphorylation of p53 in cultured CMVECs. ( A ) Formation of capillary-like tubes. CMVECs were seeded onto the matrigel in plates containing 1 µM AngII (AngII) or vehicle (Control). Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Left panel : Representative photomicrographs of capillary-like tubes (scale bar: 100 µm). Right panel : Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 15 independent experiments (n=15). ( B ) Measurement of VEGF protein levels in culture medium. Cultured CMVECs were incubated with AngII (1 µM) or vehicle (Control) for 18 hours. Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). ( C ) Cytosolic p53 and p-p53 protein expression. CMVECs were treated as described under (B) and then collected and lysed for Western blot analyses of p53 and p-p53. β-actin was used as a loading control. ( D ) Nucleus p-p53 and Hif-1α expression. Nucleus proteins isolated from CMVECs treated as described under (B) were subjected to Western blot analyses for p-p53 and Hif-1α. TFIIB was used as a loading control. Representative immunoblots are shown. The expression of p53, p-p53 or Hif-1α was quantified as folds of β-actin, p53 or TFIIB, respectively. Data represent mean ± S.E.M. obtained from 3 independent experiments (n=3). * p

    Article Snippet: Capillary-like tube formation 200 µl growth factor-reduced Matrigel matrix (BD) was homogenized, layered into a 24-well plate on a cooled planar surface and allowed to solidify at 37 °C [ ].

    Techniques: Cell Culture, Microscopy, Incubation, Enzyme-linked Immunosorbent Assay, Expressing, Western Blot, Isolation

    Effect of p53 inhibitor on AngII-induced impairment of angiogenetic responses in cultured CMVECs. ( A ) Formation of capillary-like tubes. CMVECs treated with PFT-α or with vehicle (DMSO) for 30 min were seeded onto the matrigel in plates containing 1 µM AngII with or withour PFT-α. CMVECs treated with DMSO and seeded onto the matrigel without AngII served as the control (DMSO). Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Left panel : Representative photomicrographs of capillary-like tubes (scale bar: 100 µm). Right panel : Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 15 independent experiments (n=15). ( B ) Measurement of VEGF protein levels in culture medium. CMVECs were pretreated with PFT-α or DMSO for 30 min and then incubated with AngII (1 µM) for 18 hours. CMVECs treated with DMSO but without AngII served as the control (DMSO). Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). ( C ) Cytosolic p53 and p-p53 protein expression. CMVECs treated as described under (B) were collected and lysed for Western blot analyses for p53 and p-p53. β-Actin expression served as a loading control. ( D ) Nucleus p-p53 and Hif-1α expression. Nucleus proteins isolated from CMVECs treated as described under (B) were subjected to Western blot analyses for p-p53 and Hif-1α. TFIIB was used as a loading control. Representative immunoblots are shown. The expression of p53, p-p53 or Hif-1α was quantified as folds of β-actin or of TFIIB, respectively. Data represent mean ± S.E.M. obtained from 3 independent experiments (n=3). * p

    Journal: PLoS ONE

    Article Title: Regulation of p53 by Jagged1 Contributes to Angiotensin II-Induced Impairment of Myocardial Angiogenesis

    doi: 10.1371/journal.pone.0076529

    Figure Lengend Snippet: Effect of p53 inhibitor on AngII-induced impairment of angiogenetic responses in cultured CMVECs. ( A ) Formation of capillary-like tubes. CMVECs treated with PFT-α or with vehicle (DMSO) for 30 min were seeded onto the matrigel in plates containing 1 µM AngII with or withour PFT-α. CMVECs treated with DMSO and seeded onto the matrigel without AngII served as the control (DMSO). Eighteen hours later, the formation of capillary-like tubes was observed under an optical microscope. Left panel : Representative photomicrographs of capillary-like tubes (scale bar: 100 µm). Right panel : Quantitative analysis for capillary-like tube formation. Capillary-like tubes were counted in randomly selected 5 fields for each plate. Data are expressed as mean ± S.E.M. obtained from 15 independent experiments (n=15). ( B ) Measurement of VEGF protein levels in culture medium. CMVECs were pretreated with PFT-α or DMSO for 30 min and then incubated with AngII (1 µM) for 18 hours. CMVECs treated with DMSO but without AngII served as the control (DMSO). Culture medium was collected and VEGF proteins were measured by ELISA. Data are expressed as mean ± S.E.M. obtained from 5 independent experiments (n=5). ( C ) Cytosolic p53 and p-p53 protein expression. CMVECs treated as described under (B) were collected and lysed for Western blot analyses for p53 and p-p53. β-Actin expression served as a loading control. ( D ) Nucleus p-p53 and Hif-1α expression. Nucleus proteins isolated from CMVECs treated as described under (B) were subjected to Western blot analyses for p-p53 and Hif-1α. TFIIB was used as a loading control. Representative immunoblots are shown. The expression of p53, p-p53 or Hif-1α was quantified as folds of β-actin or of TFIIB, respectively. Data represent mean ± S.E.M. obtained from 3 independent experiments (n=3). * p

    Article Snippet: Capillary-like tube formation 200 µl growth factor-reduced Matrigel matrix (BD) was homogenized, layered into a 24-well plate on a cooled planar surface and allowed to solidify at 37 °C [ ].

    Techniques: Cell Culture, Microscopy, Incubation, Enzyme-linked Immunosorbent Assay, Expressing, Western Blot, Isolation

    Effect of Thr-328-Ect2 phospho-mutants on anchorage-independent growth and invasion of NSCLC cells. A , immunoblot analysis of H1703 NSCLC cells stably transduced with either nontarget ( NT ) or Ect2 RNAi lentivirus and stably transfected with either empty vector ( V ) or one encoding WT-Ect2 ( WT ), T328A-Ect2 ( T328A ), or T328D-Ect2 ( T328D ) mutants. B , effect of WT and Thr-328-Ect2 mutants on anchorage-independent growth in soft agar. Expression of either WT or T328D-Ect2 restores soft agar colony growth to Ect2-RNAi cells, whereas expression of T328A-Ect2 does not. C , effect of WT and Thr-328-Ect2 mutants on cellular invasion through Matrigel. Expression of either WT or T328D-Ect2 restores cellular invasion to Ect2-RNAi cells, whereas expression of T328A-Ect2 does not. Data in B and C represent the mean ± the S.E., n = 4, and are presented as % NT control; * indicates significantly different from NT cells; ** indicates significantly different from Ect2 RNAi cells expressing control empty vector, p

    Journal: The Journal of Biological Chemistry

    Article Title: Oncogenic Activity of Ect2 Is Regulated through Protein Kinase C?-mediated Phosphorylation *

    doi: 10.1074/jbc.M110.196113

    Figure Lengend Snippet: Effect of Thr-328-Ect2 phospho-mutants on anchorage-independent growth and invasion of NSCLC cells. A , immunoblot analysis of H1703 NSCLC cells stably transduced with either nontarget ( NT ) or Ect2 RNAi lentivirus and stably transfected with either empty vector ( V ) or one encoding WT-Ect2 ( WT ), T328A-Ect2 ( T328A ), or T328D-Ect2 ( T328D ) mutants. B , effect of WT and Thr-328-Ect2 mutants on anchorage-independent growth in soft agar. Expression of either WT or T328D-Ect2 restores soft agar colony growth to Ect2-RNAi cells, whereas expression of T328A-Ect2 does not. C , effect of WT and Thr-328-Ect2 mutants on cellular invasion through Matrigel. Expression of either WT or T328D-Ect2 restores cellular invasion to Ect2-RNAi cells, whereas expression of T328A-Ect2 does not. Data in B and C represent the mean ± the S.E., n = 4, and are presented as % NT control; * indicates significantly different from NT cells; ** indicates significantly different from Ect2 RNAi cells expressing control empty vector, p

    Article Snippet: Matrigel-coated plates were obtained from BD Biosciences.

    Techniques: Stable Transfection, Transduction, Transfection, Plasmid Preparation, Expressing

    Mitotic Ect2 phosphorylation sites are not necessary for the oncogenic activity of Ect2. A , immunoblot analysis demonstrating expression of S-tagged wild-type Ect2 ( WT ), T328A Ect2 ( T328A ), T341A Ect2 ( T341A ), and T412A Ect2 ( T412A ) in Ect2 RNAi NSCLC cells. B , effect of WT, T328A, T341A, and T412A Ect2 mutants on anchorage-independent growth in soft agar. Only the T328A Ect2 mutant fails to restore soft agar colony growth to Ect2-RNAi cells. C , effect of WT, T328A, T341A, and T412A Ect2 mutants on cellular invasion through Matrigel. Only the T328A Ect2 mutant fails to restore soft agar colony growth to Ect2-RNAi cells. Data in B and C represent the mean ± S.E., n = 4, and are presented as % NT control; * indicates significantly different from NT cells; ** indicates significantly different from Ect2 RNAi cells expressing control empty vector; p

    Journal: The Journal of Biological Chemistry

    Article Title: Oncogenic Activity of Ect2 Is Regulated through Protein Kinase C?-mediated Phosphorylation *

    doi: 10.1074/jbc.M110.196113

    Figure Lengend Snippet: Mitotic Ect2 phosphorylation sites are not necessary for the oncogenic activity of Ect2. A , immunoblot analysis demonstrating expression of S-tagged wild-type Ect2 ( WT ), T328A Ect2 ( T328A ), T341A Ect2 ( T341A ), and T412A Ect2 ( T412A ) in Ect2 RNAi NSCLC cells. B , effect of WT, T328A, T341A, and T412A Ect2 mutants on anchorage-independent growth in soft agar. Only the T328A Ect2 mutant fails to restore soft agar colony growth to Ect2-RNAi cells. C , effect of WT, T328A, T341A, and T412A Ect2 mutants on cellular invasion through Matrigel. Only the T328A Ect2 mutant fails to restore soft agar colony growth to Ect2-RNAi cells. Data in B and C represent the mean ± S.E., n = 4, and are presented as % NT control; * indicates significantly different from NT cells; ** indicates significantly different from Ect2 RNAi cells expressing control empty vector; p

    Article Snippet: Matrigel-coated plates were obtained from BD Biosciences.

    Techniques: Activity Assay, Expressing, Mutagenesis, Plasmid Preparation

    MBECs express endothelial-specific genes and display endothelial functional characteristics . (A) Primarily passaged MBECs were collected, and total RNA was isolated and prepared for reverse-transcriptase PCR analysis for expression of multiple endothelial cell genes. VE-cadherin, VWF, VEGFR-2, and PECAM (CD31) were all expressed by MBECs, confirming their endothelial cell lineage. (B) Primary MBECs (5 × 10 4 ) were plated in 24-well tissue culture plates precoated with Matrigel to analyze for evidence of capillary tube forming capability. As shown, MBECs readily formed tubelike structures, consistent with an endothelial cell lineage.

    Journal: Blood

    Article Title: Transplantation of vascular endothelial cells mediates the hematopoietic recovery and survival of lethally irradiated mice

    doi: 10.1182/blood-2006-05-022640

    Figure Lengend Snippet: MBECs express endothelial-specific genes and display endothelial functional characteristics . (A) Primarily passaged MBECs were collected, and total RNA was isolated and prepared for reverse-transcriptase PCR analysis for expression of multiple endothelial cell genes. VE-cadherin, VWF, VEGFR-2, and PECAM (CD31) were all expressed by MBECs, confirming their endothelial cell lineage. (B) Primary MBECs (5 × 10 4 ) were plated in 24-well tissue culture plates precoated with Matrigel to analyze for evidence of capillary tube forming capability. As shown, MBECs readily formed tubelike structures, consistent with an endothelial cell lineage.

    Article Snippet: Briefly, 300 μL/well Matrigel was placed into a precooled 24-well plate (Corning) and incubated at 37°C for 30 minutes to solidify.

    Techniques: Functional Assay, Isolation, Polymerase Chain Reaction, Expressing

    ID4 inhibits myoepithelial differentiation of organoids. a) Schematic diagram of 3D Matrigel organoid assay. ID4-GFP+ basal cells were FACS purified from adult (10-11 weeks) ID4-GFP reporter mice. Exon 1 and 2 of Id4 are floxed and a GFP reporter cassette introduced. Basal cells are reprogrammed in culture using ROCK inhibitor Y-27632 and irradiated NIH-3T3 feeder cells. Adenoviral Cre is used to knock out ID4 as shown by western blotting. Single cells are then seeded on top of a Matrigel plug and grown for 6 days followed by immunofluorescent staining and quantification. Organoids grown from conditionally reprogrammed basal cells were treated with control GFP adenovirus (ID4 WT) or with Cre Adeno virus (ID4 KO) then stained for ID4 (b) and α-SMA (d). Scale bar = 10 μm. Fluorescence was quantified for ID4 (c) and α-SMA (e) in approximately 10 organoids per experiment. n=4. Unpaired two-tailed students t-test. *** p

    Journal: bioRxiv

    Article Title: Inhibitor of Differentiation 4 (ID4) represses myoepithelial differentiation of mammary stem cells through its interaction with HEB

    doi: 10.1101/2020.04.06.026963

    Figure Lengend Snippet: ID4 inhibits myoepithelial differentiation of organoids. a) Schematic diagram of 3D Matrigel organoid assay. ID4-GFP+ basal cells were FACS purified from adult (10-11 weeks) ID4-GFP reporter mice. Exon 1 and 2 of Id4 are floxed and a GFP reporter cassette introduced. Basal cells are reprogrammed in culture using ROCK inhibitor Y-27632 and irradiated NIH-3T3 feeder cells. Adenoviral Cre is used to knock out ID4 as shown by western blotting. Single cells are then seeded on top of a Matrigel plug and grown for 6 days followed by immunofluorescent staining and quantification. Organoids grown from conditionally reprogrammed basal cells were treated with control GFP adenovirus (ID4 WT) or with Cre Adeno virus (ID4 KO) then stained for ID4 (b) and α-SMA (d). Scale bar = 10 μm. Fluorescence was quantified for ID4 (c) and α-SMA (e) in approximately 10 organoids per experiment. n=4. Unpaired two-tailed students t-test. *** p

    Article Snippet: Tissue culture flasks coated with Growth Factor-reduced Matrigel (Corning) diluted 1:60 in PBS for 30 min to 1 hr at 37°C.

    Techniques: FACS, Purification, Mouse Assay, Irradiation, Knock-Out, Western Blot, Staining, Fluorescence, Two Tailed Test

    BAF53a promotes proliferation, migration and invasion of glioma cells. The overexpression and knockdown interfered efficiency of BAF53a detected by real-time PCR (A) and western blotting (B), results showed shBAF53a-2 had the best knockdown effect and the overexpression effect was also satisfactory. (C) The MTT assays showed BAF53a overexpression promoted glioma cell proliferation, while BAF53a knockdown inhibited glioma cell proliferation. (D) The colony formation assays also showed BAF53a expression promoted glioma cell proliferation. (E) The Transwell assay showed BAF53a expression facilitated migration of glioma cells. (F) The wound healing assay showed BAF53a expression provided the faster wound healing capacities of glioma cells. (G) The Transwell-Matrigel chamber assay showed BAF53a expression facilitated invasion of glioma cells.

    Journal: Oncology Reports

    Article Title: BAF53a is a potential prognostic biomarker and promotes invasion and epithelial-mesenchymal transition of glioma cells

    doi: 10.3892/or.2017.6019

    Figure Lengend Snippet: BAF53a promotes proliferation, migration and invasion of glioma cells. The overexpression and knockdown interfered efficiency of BAF53a detected by real-time PCR (A) and western blotting (B), results showed shBAF53a-2 had the best knockdown effect and the overexpression effect was also satisfactory. (C) The MTT assays showed BAF53a overexpression promoted glioma cell proliferation, while BAF53a knockdown inhibited glioma cell proliferation. (D) The colony formation assays also showed BAF53a expression promoted glioma cell proliferation. (E) The Transwell assay showed BAF53a expression facilitated migration of glioma cells. (F) The wound healing assay showed BAF53a expression provided the faster wound healing capacities of glioma cells. (G) The Transwell-Matrigel chamber assay showed BAF53a expression facilitated invasion of glioma cells.

    Article Snippet: The Matrigel matrix was purchased from Corning Life Sciences (354248; Corning, MA, USA).

    Techniques: Migration, Over Expression, Real-time Polymerase Chain Reaction, Western Blot, MTT Assay, Expressing, Transwell Assay, Wound Healing Assay, Boyden Chamber Assay

    Effect of DMSA-Fe 2 O 3 on tube network formed by HAECs cultured on Matrigel within 14 h. ( a ) HAECs can form a capillary-like network on Matrigel-coated wells within 14 h. ( b ) An obvious failure to form networks by HAECs in the presence of 0.01 mg/ml DMSA-Fe 2 O 3 . ( c ) Few tube networks by HAECs in the presence of 0.02 mg/ml DMSA-Fe 2 O 3 . ( d ) The high urea solution (6M urea) was used as a positive control for the inhibition of tube formation.

    Journal: Nanoscale Research Letters

    Article Title: The cytotoxicity evaluation of magnetic iron oxide nanoparticles on human aortic endothelial cells

    doi: 10.1186/1556-276X-8-215

    Figure Lengend Snippet: Effect of DMSA-Fe 2 O 3 on tube network formed by HAECs cultured on Matrigel within 14 h. ( a ) HAECs can form a capillary-like network on Matrigel-coated wells within 14 h. ( b ) An obvious failure to form networks by HAECs in the presence of 0.01 mg/ml DMSA-Fe 2 O 3 . ( c ) Few tube networks by HAECs in the presence of 0.02 mg/ml DMSA-Fe 2 O 3 . ( d ) The high urea solution (6M urea) was used as a positive control for the inhibition of tube formation.

    Article Snippet: In this study, the Matrigel basement membrane matrix was used as extracellular matrix support to observe whether angiogenesis of HAEC can be intervened by DMSA-Fe2 O3 or not.

    Techniques: Cell Culture, Positive Control, Inhibition

    Length of tube networks formed by HAEC cultured on Matrigel. Image-Pro plus 6.0 for Windows software was used to measure the length of tube networks (pixels). The stained cells were inspected under a light microscope at ×100 magnification and captured more than three pictures from different fields. The average data from the same well was calculated as its quantitative value. Data are expressed as mean ± SD. ** p

    Journal: Nanoscale Research Letters

    Article Title: The cytotoxicity evaluation of magnetic iron oxide nanoparticles on human aortic endothelial cells

    doi: 10.1186/1556-276X-8-215

    Figure Lengend Snippet: Length of tube networks formed by HAEC cultured on Matrigel. Image-Pro plus 6.0 for Windows software was used to measure the length of tube networks (pixels). The stained cells were inspected under a light microscope at ×100 magnification and captured more than three pictures from different fields. The average data from the same well was calculated as its quantitative value. Data are expressed as mean ± SD. ** p

    Article Snippet: In this study, the Matrigel basement membrane matrix was used as extracellular matrix support to observe whether angiogenesis of HAEC can be intervened by DMSA-Fe2 O3 or not.

    Techniques: Cell Culture, Software, Staining, Light Microscopy

    Formation of tubular network by MDA-MB-231 cells was tested on matrigel in presence and absence of ephrinB2-Fc. Cells were treated with (a) PBS, (b) 3 μ g/mL ephrinB2-Fc and (c) 10 μ g/mL ephrinB2-Fc. Treatment with ephrinB2-Fc was expected to totally abrogate tubular formation and form aggregated colonies, yet it was surprisingly observed that tubular formation was not inhibited by ephrinB2-Fc treatment compared to PBS. Figures were inspected at 100x magnification and scale bars represent 20 μ m.

    Journal: Disease Markers

    Article Title: EphB4 Tyrosine Kinase Stimulation Inhibits Growth of MDA-MB-231 Breast Cancer Cells in a Dose and Time Dependent Manner

    doi: 10.1155/2013/857895

    Figure Lengend Snippet: Formation of tubular network by MDA-MB-231 cells was tested on matrigel in presence and absence of ephrinB2-Fc. Cells were treated with (a) PBS, (b) 3 μ g/mL ephrinB2-Fc and (c) 10 μ g/mL ephrinB2-Fc. Treatment with ephrinB2-Fc was expected to totally abrogate tubular formation and form aggregated colonies, yet it was surprisingly observed that tubular formation was not inhibited by ephrinB2-Fc treatment compared to PBS. Figures were inspected at 100x magnification and scale bars represent 20 μ m.

    Article Snippet: Alamar-blue was obtained from AbdSerotec, and matrigel was purchased from Invitrogen.

    Techniques: Multiple Displacement Amplification

    c-MET protein and mRNA levels are highly upregulated in invasive CRC daughter cell lines BD BioCoat Matrigel invasion chambers were used to isolate invasive subpopulations from a panel of CRC cells. ( A ) Migration and invasion assays of invasive subpopulations of HCT116, HKH-2 and LoVo sublines were compared with the parental cells using the quantitative xCELLigence system. ( B ) qRT-PCR assessment of MET gene expression levels in parental and invasive cell models. ( C ) Western blot analysis of pc-MET 1234/1235 and c-MET protein expression levels in parental (Par) and invasive (Inv) cell models. ( D ) HGF protein levels in the culture media of parental (Par) and invasive (Inv) subpopulations were measured by ELISA [Positive control: CCD-18Co (18Co)]. *= p

    Journal: Oncotarget

    Article Title: Transcriptional upregulation of c-MET is associated with invasion and tumor budding in colorectal cancer

    doi: 10.18632/oncotarget.12933

    Figure Lengend Snippet: c-MET protein and mRNA levels are highly upregulated in invasive CRC daughter cell lines BD BioCoat Matrigel invasion chambers were used to isolate invasive subpopulations from a panel of CRC cells. ( A ) Migration and invasion assays of invasive subpopulations of HCT116, HKH-2 and LoVo sublines were compared with the parental cells using the quantitative xCELLigence system. ( B ) qRT-PCR assessment of MET gene expression levels in parental and invasive cell models. ( C ) Western blot analysis of pc-MET 1234/1235 and c-MET protein expression levels in parental (Par) and invasive (Inv) cell models. ( D ) HGF protein levels in the culture media of parental (Par) and invasive (Inv) subpopulations were measured by ELISA [Positive control: CCD-18Co (18Co)]. *= p

    Article Snippet: For classical migration/invasion assays, Corning Transwell PET migration Chambers (Corning) and BD BioCoat Matrigel Invasion Chambers (BD Biosciences) were used as described previously [ , ].

    Techniques: Migration, Quantitative RT-PCR, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Positive Control

    MMP activity influences LPS- and IL-10-stimulated macrophage-mediated cancer cell invasion and angiogenesis. a AGS cells were incubated in BD BioCoat TM Matrigel TM Invasion Chambers for 24 h with RPMI medium (-) or macrophages stimulated for 72 h with 10 ng/ml LPS (LPSmac)) or with 10 ng/ml IL-10 (IL-10mac) and supplied or not with a pharmacological inhibitor of matrix metalloproteases, Galardin (10 μM). Invasive cells were determined as described in Materials and Methods. Bars represent mean values of independent experiments performed with, at least, 4 different donors; flags indicate standard deviations. *, significantly different from AGS in RPMI medium at p

    Journal: BMC Cancer

    Article Title: Matrix metalloproteases as maestros for the dual role of LPS- and IL-10-stimulated macrophages in cancer cell behaviour

    doi: 10.1186/s12885-015-1466-8

    Figure Lengend Snippet: MMP activity influences LPS- and IL-10-stimulated macrophage-mediated cancer cell invasion and angiogenesis. a AGS cells were incubated in BD BioCoat TM Matrigel TM Invasion Chambers for 24 h with RPMI medium (-) or macrophages stimulated for 72 h with 10 ng/ml LPS (LPSmac)) or with 10 ng/ml IL-10 (IL-10mac) and supplied or not with a pharmacological inhibitor of matrix metalloproteases, Galardin (10 μM). Invasive cells were determined as described in Materials and Methods. Bars represent mean values of independent experiments performed with, at least, 4 different donors; flags indicate standard deviations. *, significantly different from AGS in RPMI medium at p

    Article Snippet: Invasion assays Invasion assays were performed as previously [ ], using BD BioCoat™ Matrigel™ Invasion Chambers (BD Biosciences, Madrid, Spain) and AGS or RKO cells in the upper compartment, and LPS- (LPSmac) or IL-10-stimulated macrophages (IL-10mac) in the lower compartment.

    Techniques: Activity Assay, Incubation

    IL-10-stimulated macrophages are more efficient in stimulating cancer cell invasion. a AGS human gastric or b RKO human colorectal cancer cells were incubated in BD BioCoat TM Matrigel TM Invasion Chambers for 24 h with RPMI medium (-), or human macrophages differentiated for 10 days and stimulated for 72 h with 10 ng/ml LPS (LPSmac) or 10 ng/ml IL-10 (IL-10mac). Invasive cells and invasive ratio were determined as described in Materials and Methods. Bars represent mean values of independent experiments performed with, at least, 4 different donors; flags indicate standard deviations

    Journal: BMC Cancer

    Article Title: Matrix metalloproteases as maestros for the dual role of LPS- and IL-10-stimulated macrophages in cancer cell behaviour

    doi: 10.1186/s12885-015-1466-8

    Figure Lengend Snippet: IL-10-stimulated macrophages are more efficient in stimulating cancer cell invasion. a AGS human gastric or b RKO human colorectal cancer cells were incubated in BD BioCoat TM Matrigel TM Invasion Chambers for 24 h with RPMI medium (-), or human macrophages differentiated for 10 days and stimulated for 72 h with 10 ng/ml LPS (LPSmac) or 10 ng/ml IL-10 (IL-10mac). Invasive cells and invasive ratio were determined as described in Materials and Methods. Bars represent mean values of independent experiments performed with, at least, 4 different donors; flags indicate standard deviations

    Article Snippet: Invasion assays Invasion assays were performed as previously [ ], using BD BioCoat™ Matrigel™ Invasion Chambers (BD Biosciences, Madrid, Spain) and AGS or RKO cells in the upper compartment, and LPS- (LPSmac) or IL-10-stimulated macrophages (IL-10mac) in the lower compartment.

    Techniques: Incubation

    Role of miR-200b subfamily and miR-205 on prostate cancer proliferation and invasion ( A ) Expression levels of miR-200a, miR-200b, miR-429, miR-205, and ERG in commonly used prostate cancer cell lines, including LNCaP, VCaP, 22RV1, PC3, DU145. Mature miRNAs expression levels were measured by qRT-PCR, normalized against RNU48 internal control. ( B ) Expression of miRNAs in PC3 stable cell lines, including PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( C ) The effect of miRNAs on PC3 cell growth as determined by MTT assay. ( D ) PC3 stable cell Matrigel invasion assay using BD Biocoat Matrigel Invasion Chambers. Representative pictures of invaded cells stained with crystal violet. Four stable cell lines, PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( E ) Quantification of invasion. Invaded cells were stained with crystal violet, which was then solubilized in 1% SDS. The absorbance was measured at 595 nm. Error bars, mean ± SEM. * P

    Journal: Oncotarget

    Article Title: The tumor suppressive miR-200b subfamily is an ERG target gene in human prostate tumors

    doi: 10.18632/oncotarget.9366

    Figure Lengend Snippet: Role of miR-200b subfamily and miR-205 on prostate cancer proliferation and invasion ( A ) Expression levels of miR-200a, miR-200b, miR-429, miR-205, and ERG in commonly used prostate cancer cell lines, including LNCaP, VCaP, 22RV1, PC3, DU145. Mature miRNAs expression levels were measured by qRT-PCR, normalized against RNU48 internal control. ( B ) Expression of miRNAs in PC3 stable cell lines, including PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( C ) The effect of miRNAs on PC3 cell growth as determined by MTT assay. ( D ) PC3 stable cell Matrigel invasion assay using BD Biocoat Matrigel Invasion Chambers. Representative pictures of invaded cells stained with crystal violet. Four stable cell lines, PC3-vec, PC3-miR-200b/a/429, PC3-miR-205, and PC3-miR-200b/a/429/205. ( E ) Quantification of invasion. Invaded cells were stained with crystal violet, which was then solubilized in 1% SDS. The absorbance was measured at 595 nm. Error bars, mean ± SEM. * P

    Article Snippet: The Matrigel invasion assay was performed using the BD Biocoat Matrigel Invasion Chamber (BD Biosciences) following the manufacturer's instructions, and the pictures were taken using the EVOS XL Cell Imaging system (ThermoFisher).

    Techniques: Expressing, Quantitative RT-PCR, Stable Transfection, MTT Assay, Invasion Assay, Staining

    Invasion, attachment, and proliferation assay of glioblastoma cells . All assays were performed with four different glioblastoma cell lines: U87, U251, U373, and C6 (Top to bottom). Values in (A) and (B) are expressed as a percentage of corresponding parental line data. (A) Invasion: IM-3 subpopulations were compared to their parental lines. (B) Attachment: Cells adherent to Matrigel ® after 4 h were stained with hematoxylin QS and counted. (C) Proliferation: The rate of cell proliferation was evaluated with CellTiter 96 Aqueous One Solution (Promega) at five different time points, 4 h, 24 h, 48 h, 72 h and 96 h after seeding cells. Triangles - IM-3 lines, squares - parental. Data are represented as means ± SD (n = 3). P, parental cells; IM3, IM3 cells; IM3, IM3 cell line; **, p

    Journal: BMC Cancer

    Article Title: Serial selection for invasiveness increases expression of miR-143/miR-145 in glioblastoma cell lines

    doi: 10.1186/1471-2407-12-143

    Figure Lengend Snippet: Invasion, attachment, and proliferation assay of glioblastoma cells . All assays were performed with four different glioblastoma cell lines: U87, U251, U373, and C6 (Top to bottom). Values in (A) and (B) are expressed as a percentage of corresponding parental line data. (A) Invasion: IM-3 subpopulations were compared to their parental lines. (B) Attachment: Cells adherent to Matrigel ® after 4 h were stained with hematoxylin QS and counted. (C) Proliferation: The rate of cell proliferation was evaluated with CellTiter 96 Aqueous One Solution (Promega) at five different time points, 4 h, 24 h, 48 h, 72 h and 96 h after seeding cells. Triangles - IM-3 lines, squares - parental. Data are represented as means ± SD (n = 3). P, parental cells; IM3, IM3 cells; IM3, IM3 cell line; **, p

    Article Snippet: Boyden chamber migration assay of knockdown miRNA Transfected cells A suspension of 300,000 cells/mL in serum-free medium of parental and IM3 cells transfected with LNA antagomirs (against miR-143 and miR-145 or a control LNA antagomir) was plated in the upper chamber of a Boyden manifold over a Matrigel-coated membrane (24-well insert; pore size, 8 um; BD Biosciences).

    Techniques: Proliferation Assay, Staining

    Selection of highly invasive glioblastoma cell line subpopulations (IM3) . (A) Schematic diagram of our serial selection algorithm (Matrigel Invasion Chamber, BD Bioscience). (B) Serial selection of cells resulted in highly invasive subpopulations of each cell type. Cells (P, parental cells) were invaded through 8 μm Matrigel ® pores three times for selection of invasive cells (IM3).

    Journal: BMC Cancer

    Article Title: Serial selection for invasiveness increases expression of miR-143/miR-145 in glioblastoma cell lines

    doi: 10.1186/1471-2407-12-143

    Figure Lengend Snippet: Selection of highly invasive glioblastoma cell line subpopulations (IM3) . (A) Schematic diagram of our serial selection algorithm (Matrigel Invasion Chamber, BD Bioscience). (B) Serial selection of cells resulted in highly invasive subpopulations of each cell type. Cells (P, parental cells) were invaded through 8 μm Matrigel ® pores three times for selection of invasive cells (IM3).

    Article Snippet: Boyden chamber migration assay of knockdown miRNA Transfected cells A suspension of 300,000 cells/mL in serum-free medium of parental and IM3 cells transfected with LNA antagomirs (against miR-143 and miR-145 or a control LNA antagomir) was plated in the upper chamber of a Boyden manifold over a Matrigel-coated membrane (24-well insert; pore size, 8 um; BD Biosciences).

    Techniques: Selection

    Establishment of mouse prostate organoid cultures A) Overview of the isolation of the prostate from the mouse urogenital system (for a detailed isolation protocol see 25 ). The procedure refers to pictures I – IX. B) Schematic representation of the anatomy of the mouse prostate. AP, anterior prostate; DLP, dorsolateral prostate; VP, ventral prostate. C) Example of how to plate the matrigel disc in a well of a tissue culture plate. D) Representative pictures of organoids growing from mouse prostate tissue 1 and 7 days after plating. Scale bars, 100 micron.

    Journal: Nature protocols

    Article Title: Organoid culture systems for prostate epithelial tissue and prostate cancer tissue

    doi: 10.1038/nprot.2016.006

    Figure Lengend Snippet: Establishment of mouse prostate organoid cultures A) Overview of the isolation of the prostate from the mouse urogenital system (for a detailed isolation protocol see 25 ). The procedure refers to pictures I – IX. B) Schematic representation of the anatomy of the mouse prostate. AP, anterior prostate; DLP, dorsolateral prostate; VP, ventral prostate. C) Example of how to plate the matrigel disc in a well of a tissue culture plate. D) Representative pictures of organoids growing from mouse prostate tissue 1 and 7 days after plating. Scale bars, 100 micron.

    Article Snippet: Reagents Collagenase Type II (Life Technologies, cat. no. 17101-015) TrypLE Express (Life Technologies, cat. no. 12605-010) Dulbeccos Modified Eagle Medium (DMEM; Life Technologies, cat. no. 31966) Advanced DMEM/F12 (adDMEM/F12; Life Technologies, cat. no. 12634-034) GlutaMAX 100× (Life Technologies, cat. no. 35050-068) Penicillin-streptomycin (Life Technologies, cat. no. 15140-122) Hepes (Life Technologies, cat. no. 15630-056) Zeocin (Life Technologies, cat. no. R250-01) Phosphate buffered saline Matrigel, Growth Factor Reduced (GFR), Phenol Red-free (BD, cat. no. 356231) B27 supplement 50× (Life Technologies, cat. no. 17504-044) Nicotinamide (Sigma-Aldrich, cat. no. N0636) N-acetylcysteine (Sigma-Aldrich, cat. no. A9165) A83-01 (Tocris Bioscience, cat. no. 2939) Y-27632 (Abmole Bioscience, cat. no. M1817) Human FGF-10 (PeproTech, cat. no. 100-26) Human FGF-2 (PeproTech, cat. no. 100-18B) Human EGF (PeproTech, cat. no. AF-100-15) Recombinant human Noggin (Peprotech, cat. no. 120-10C) R-spondin 1-conditioned medium; home made from the 293T-HA-RspoI-Fc cell line (derived from Calvin Kuo lab), or recombinant R-spondin 1 protein (R & D Systems, cat. no. 4645-RS-025) Prostaglandin E2 (Tocris Bioscience, cat. no. 2296) SB202190 (Sigma-Aldrich, cat. no. S7076) (DiHydro)Testosterone (5α-Androstan-17β-ol-3-one) (Sigma-Aldrich, cat. no. A8380) Fetal bovine serum (Sigma-Aldrich, cat. no. F7524) Deoxyribonuclease I (DNase I) from bovine pancreas (Sigma-Aldrich, cat. no. D5025) DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride; Life Technologies, cat. no. D1306) RosetteSep® Human CD45 Depletion Cocktail (Stem Cell Technologies, cat. no. 15122) Ficoll-Paque™ PLUS (GE Healthcare Life Sciences, cat. no. 17-1440-02) Recovery Cell Culture Freezing medium (Life Technologies, cat. no. 12648-010) RNeasy mini kit (Qiagen, cat. no. 74104) Reliaprep gDNA tissue miniprep system (Promega, cat. no. A2052) GoScript Reverse Transcriptase (Promega, cat. no. A5003) Oligo(dT) 15 Primer (Promega, cat. no. C1101) Rec.

    Techniques: Isolation

    Generation of induced pluripotent stem cells (iPSCs) and retinal pigment epithelium cells (RPE) from RP2 R120X fibroblasts. ( A ) Schematic representation of iPSC and RPE cell generation from fibroblasts. A skin biopsy was taken and cut into small pieces using scalpel blades. The cells were then cultured in DMEM and outgrowing fibroblasts from the skin biopsy were isolated manually. To reprogram fibroblasts into iPSCs, cells were electroporated with plasmids containing Oct4, Sox2, Myc, Lin28 and Klf4, plated onto matrigel-coated dishes and cultured in mTESR medium. Forming iPSC colonies were isolated manually. iPSC were differentiated into RPE cells by culturing in HESC—bFGF medium. Pigmented RPE colonies were excised manually using crescent blades. ( B ) iPSC derived RPE from controls and the RP2 R120X patient showed nuclear (DAPI, blue) localization of the iPSC markers Nanog (red) and Oct4 (red). Scale bar 10 μm. ( C ) Differentiated control and R120X iPSC-derived RPE cells display the classic cobblestone pigmented morphology. Scale bar 10 μm. ( D ) Sectioned RPE cells were stained for RPE and differentiation markers: ATP1B1 (green), Col4 (red), Bestrophin (green), MerTK (red), Pmel17 (green), ZO1 (red), mitf (green) or ( E ) RPE65 (green). Scale bar 10 μm ( F ) RP2 protein is undetectable in iPSC-derived R120X RPE cells at the plasma membrane and the basal body by ICC. Scale bar 10 μm. ( G ) RP2 protein is undetectable in the iPSC-derived R120X RPE cells by western blotting. GAPDH was used as a loading control. ( H ) Sectioned RPE cells were stained for endogenous Gβ1 (green) the membrane marker wheat-germ agglutinin (WGA, red) and DAPI (blue). Gβ1 localizes to the plasma membrane in control RPE cells, but shows a diffuse cytoplasmic staining pattern in R120X RPE cells. Scale bar 10 μm.

    Journal: Human Molecular Genetics

    Article Title: Translational read-through of the RP2 Arg120stop mutation in patient iPSC-derived retinal pigment epithelium cells

    doi: 10.1093/hmg/ddu509

    Figure Lengend Snippet: Generation of induced pluripotent stem cells (iPSCs) and retinal pigment epithelium cells (RPE) from RP2 R120X fibroblasts. ( A ) Schematic representation of iPSC and RPE cell generation from fibroblasts. A skin biopsy was taken and cut into small pieces using scalpel blades. The cells were then cultured in DMEM and outgrowing fibroblasts from the skin biopsy were isolated manually. To reprogram fibroblasts into iPSCs, cells were electroporated with plasmids containing Oct4, Sox2, Myc, Lin28 and Klf4, plated onto matrigel-coated dishes and cultured in mTESR medium. Forming iPSC colonies were isolated manually. iPSC were differentiated into RPE cells by culturing in HESC—bFGF medium. Pigmented RPE colonies were excised manually using crescent blades. ( B ) iPSC derived RPE from controls and the RP2 R120X patient showed nuclear (DAPI, blue) localization of the iPSC markers Nanog (red) and Oct4 (red). Scale bar 10 μm. ( C ) Differentiated control and R120X iPSC-derived RPE cells display the classic cobblestone pigmented morphology. Scale bar 10 μm. ( D ) Sectioned RPE cells were stained for RPE and differentiation markers: ATP1B1 (green), Col4 (red), Bestrophin (green), MerTK (red), Pmel17 (green), ZO1 (red), mitf (green) or ( E ) RPE65 (green). Scale bar 10 μm ( F ) RP2 protein is undetectable in iPSC-derived R120X RPE cells at the plasma membrane and the basal body by ICC. Scale bar 10 μm. ( G ) RP2 protein is undetectable in the iPSC-derived R120X RPE cells by western blotting. GAPDH was used as a loading control. ( H ) Sectioned RPE cells were stained for endogenous Gβ1 (green) the membrane marker wheat-germ agglutinin (WGA, red) and DAPI (blue). Gβ1 localizes to the plasma membrane in control RPE cells, but shows a diffuse cytoplasmic staining pattern in R120X RPE cells. Scale bar 10 μm.

    Article Snippet: On Day 7 the cells were dissociated using TrypLE™ Express and plated onto 6-well plates coated with Corning® Matrigel® basement membrane matrix (Corning) at a density of 2 × 105 cells per well and maintained overnight in fibroblast growth medium.

    Techniques: Cell Culture, Isolation, Derivative Assay, Staining, Immunocytochemistry, Western Blot, Marker, Whole Genome Amplification

    CXCL12 (SDF-1α) enhances granulation tissue formation that mimics the stromal reaction and angiogenesis in a Matrigel implantation model. A: Typical appearance of stromal formation and angiogenesis formed around a Matrigel. Scale bars = 10 mm.

    Journal: The American Journal of Pathology

    Article Title: COX-2 and Prostaglandin EP3/EP4 Signaling Regulate the Tumor Stromal Proangiogenic Microenvironment via CXCL12-CXCR4 Chemokine Systems

    doi: 10.2353/ajpath.2010.090607

    Figure Lengend Snippet: CXCL12 (SDF-1α) enhances granulation tissue formation that mimics the stromal reaction and angiogenesis in a Matrigel implantation model. A: Typical appearance of stromal formation and angiogenesis formed around a Matrigel. Scale bars = 10 mm.

    Article Snippet: Under light ether anesthesia, mice were subcutaneously injected in the back with 0.25 ml of phenol-red-free Matrigel (Japan BD Biosciences, Tokyo, Japan), diluted 1:1 in saline.

    Techniques:

    LINC00152 promotes migration and invasion of TSCC cells. Transwell chambers with or without Matrigel were used to detect the migration and invasion of TSCC CAL-27 and SCC-9 cells as indicated (n=3). Magnification, ×40. Data are presented as the mean ± standard deviation. **P

    Journal: Oncology Letters

    Article Title: Long non-coding RNA LINC00152 acts as a sponge of miRNA-193b-3p to promote tongue squamous cell carcinoma progression

    doi: 10.3892/ol.2020.11293

    Figure Lengend Snippet: LINC00152 promotes migration and invasion of TSCC cells. Transwell chambers with or without Matrigel were used to detect the migration and invasion of TSCC CAL-27 and SCC-9 cells as indicated (n=3). Magnification, ×40. Data are presented as the mean ± standard deviation. **P

    Article Snippet: For the invasion assay, the Transwell chamber was precoated with Matrigel solution (BD Biosciences), and ~4×105 cells were seeded into the upper chamber.

    Techniques: Migration, Standard Deviation

    CCL7 induces migration and invasion of colon cancer cells via CCR3 A. Quantitation of E-cadherin expression on the surface of HCT116 cells treated with or without recombinant CCL7 (200 ng/ml) using FACS analysis. B. Expression of E-cadherin, N-cadherin, and vimentin in HCT116 cells stably transfected with GFP or CCL7 were measured by western blotting (left panel). Expression of CCL7 and EMT markers in negative siRNA control-treated or CCL7 specific siRNAs-treated HCT116 cell extracts (right panel). Actin was used as a loading control. C. A wound healing assay was performed by creating a wound on a confluent monolayer of stable GFP/CCL7 overexpressing cells using l-Dish 35-mm high culture inserts. D. Transwell matrigel invasion assays of HCT116 cells after transfection with 100 nM of negative siRNA control or CCL7 specific siRNAs. E. Cell migration and F. Invasion was measured using a trans-well migration chamber (left panels). Mean fluorescence intensity (MFI) of invaded area is presented in bar graphs (right panels). G. HCT116 cells were treated with 20 nM CCR3 inhibitor SB 328437. Representative images of invaded cells are shown (left panels). MFI values are presented in bar graphs (right panels). H. Expression of EMT markers in HCT116 cells stably transfected with GFP or CCL7 with or without treatment with 20 nM SB 328437 (CCR3 inhibitor) was measured by western blotting. Actin was used as a loading control. Columns: means ± SEs. ** P

    Journal: Oncotarget

    Article Title: Crosstalk between CCL7 and CCR3 promotes metastasis of colon cancer cells via ERK-JNK signaling pathways

    doi: 10.18632/oncotarget.9209

    Figure Lengend Snippet: CCL7 induces migration and invasion of colon cancer cells via CCR3 A. Quantitation of E-cadherin expression on the surface of HCT116 cells treated with or without recombinant CCL7 (200 ng/ml) using FACS analysis. B. Expression of E-cadherin, N-cadherin, and vimentin in HCT116 cells stably transfected with GFP or CCL7 were measured by western blotting (left panel). Expression of CCL7 and EMT markers in negative siRNA control-treated or CCL7 specific siRNAs-treated HCT116 cell extracts (right panel). Actin was used as a loading control. C. A wound healing assay was performed by creating a wound on a confluent monolayer of stable GFP/CCL7 overexpressing cells using l-Dish 35-mm high culture inserts. D. Transwell matrigel invasion assays of HCT116 cells after transfection with 100 nM of negative siRNA control or CCL7 specific siRNAs. E. Cell migration and F. Invasion was measured using a trans-well migration chamber (left panels). Mean fluorescence intensity (MFI) of invaded area is presented in bar graphs (right panels). G. HCT116 cells were treated with 20 nM CCR3 inhibitor SB 328437. Representative images of invaded cells are shown (left panels). MFI values are presented in bar graphs (right panels). H. Expression of EMT markers in HCT116 cells stably transfected with GFP or CCL7 with or without treatment with 20 nM SB 328437 (CCR3 inhibitor) was measured by western blotting. Actin was used as a loading control. Columns: means ± SEs. ** P

    Article Snippet: Briefly, cells overexpressing CCL7 were suspended in 50 μl PBS supplemented with 50% matrigel and injected subcutaneously into the flanks of 6-week-old female BALB/c nu/nu mice (Charles River Laboratories, Wilmington, DE, USA).

    Techniques: Migration, Quantitation Assay, Expressing, Recombinant, FACS, Stable Transfection, Transfection, Western Blot, Wound Healing Assay, Fluorescence