Journal: Development (Cambridge, England)

Article Title: TIE1 dependent lymphatic vascular remodeling is mediated by its second tyrosine kinase domain.

doi: 10.1242/dev.204469

Figure Lengend Snippet: Fig.10. FOXC2 expression was abolished in Tie1 deficient lymphatics but restored

Article Snippet: Primary antibodies for immunohistochemistry Antibody Supplier Catalogue # Application Goat anti-mouse VEGFR3 R&D Systems AF743 1:50 Goat anti-human TIE1 R&D Systems AF619 1:100 Goat anti-mouse TIE2 R&D Systems AF762 1:200 Goat anti-mouse integrinα9 R&D Systems AF3827 1:50 Goat anti-VE-Cadherin R&D Systems AF1002 1:400 Goat anti-human Prox1 R&D Systems AF2727 1:100 Sheep anti-mouse FOXC2 R&D Systems AF6989 1:200 Rabbit anti-GFP Invitrogen A11122 1:200 Rabbit anti-FOXO1 Cell Signaling 2880 1:200 Rabbit anti-Lyve1 abcam Ab14917 1:800 Rabbit anti-Prox1 abcam Ab37128 1:250 Rat anti-mouse CD31 PD Pharmingen 557355 1:200 Rat anti-endomucin abcam ab106100 1:200 Development: doi:10.1242/dev.204469: Supplementary information D ev el o pm en t • S up pl em en ta ry in fo rm at io n

Techniques: Expressing

Journal: Frontiers in Oncology

Article Title: The FENDRR/FOXC2 Axis Contributes to Multidrug Resistance in Gastric Cancer and Correlates With Poor Prognosis

doi: 10.3389/fonc.2021.634579

Figure Lengend Snippet: FENDRR-regulated FOXC2 overexpression promotes GC drug resistance (A) qRT-PCR analysis of FENDRR, FOXF1, FOXL1 and FOXC2 expression in SGC7901/ADR cells infected with FENDRR shRNA or negative controls. (B) Western blot analysis of FOXC2 expression in SGC7901/ADR and SGC7901/VCR cells infected with FENDRR shRNA and in SGC7901 cells transfected with FENDRR vectors and the corresponding negative controls. (C) Expression levels of FENDRR in the nucleus and cytoplasm of the indicated cells was measured by qPCR. U1 and GAPDH were used as nuclear and cytoplasmic controls, respectively. (D) HEK-293 cells were transfected with FOXC2 luciferase reporter constructs and FENDRR-expressing vectors or shRNA and the corresponding negative controls. Luciferase activity values were measured and analyzed. Luciferase values are normalized to empty vector control values. (E) Survival of indicated cells after step-up concentration of ADR treatment for 72 h was evaluated using the CCK-8 assay. (F, G) Colony formation assay with the indicated cells treated with ADR (F) and the apoptotic rate of the indicated cells treated with 5-FU (G) are shown. (H, I) Colony formation assay (H) and apoptosis assay (I) of SGC7901/ADR and SGC7901/VCR cells infected with LV-shFENDRR with FOXC2 vector or control vector after ADR and 5-FU treatment, respectively. (J) LV-shFOXC2-infected cells were transplanted into the right flank of nude mice, and the corresponding control cells were transplanted into the left side. Left, representative images of tumors formed in nude mice (n=5) after ADR treatment are shown, scale bar, 1 cm. Middle, the tumor volumes of different groups were measured at the indicated time points. Right, the tumor weights of different groups measured after tumor isolation. (K) Quantification of the Ki67-positive staining of cells in xenografts from nude mice. ** P < 0.01, * P < 0.05, N.S. , not significant ( P > 0.05), error bars, s.d.

Article Snippet: Briefly, tissue sections were deparaffinized, subjected to antigen retrieval and endogenous peroxidase inactivation, and incubated with primary antibodies against FOXC2 (R&D Systems, AF6989) and Ki-67 (Abcam, ab15580).

Techniques: Over Expression, Quantitative RT-PCR, Expressing, Infection, shRNA, Western Blot, Transfection, Luciferase, Construct, Activity Assay, Plasmid Preparation, Control, Concentration Assay, CCK-8 Assay, Colony Assay, Apoptosis Assay, Isolation, Staining

Journal: Frontiers in Oncology

Article Title: The FENDRR/FOXC2 Axis Contributes to Multidrug Resistance in Gastric Cancer and Correlates With Poor Prognosis

doi: 10.3389/fonc.2021.634579

Figure Lengend Snippet: FENDRR increases FOXC2 expression by competitively binding miR-4700-3p (A) Prediction of the potential miRNAs targeting both FENDRR and FOXC2 with two independent miRNA target databases. (B) qRT-PCR analysis of miR-4700-3p expression in the indicated cells. (C) The expression of miR-4700-3p following the downregulation or upregulation of FENDRR expression in the indicated cells. (D) Western blot analysis of FOXC2 expression in SGC7901/ADR and SGC7901/VCR cells transfected with miR-4700-3p mimics or inhibitors and the corresponding negative controls. (E, F) Predicted potential binding sites of miR-4700-3p to FENDRR and FOXC2 are shown (upper panel). The wild-type (wt) and mutant (mut) miR-4700-3p target sequences of FENDRR (E) and FOXC2 (F) were fused to a luciferase reporter and cotransfected into HEK-293 cells with miR-4700-3p mimics or a negative control. Luciferase activity values were measured and normalized to empty vector control values. (G, H) Western blot analysis of FOXC2 expression in the indicated cells. ** P < 0.01, * P < 0.05, N.S. , not significant ( P > 0.05), error bars, s.d.

Article Snippet: Briefly, tissue sections were deparaffinized, subjected to antigen retrieval and endogenous peroxidase inactivation, and incubated with primary antibodies against FOXC2 (R&D Systems, AF6989) and Ki-67 (Abcam, ab15580).

Techniques: Expressing, Binding Assay, Quantitative RT-PCR, Western Blot, Transfection, Mutagenesis, Luciferase, Negative Control, Activity Assay, Plasmid Preparation, Control

Journal: Frontiers in Oncology

Article Title: The FENDRR/FOXC2 Axis Contributes to Multidrug Resistance in Gastric Cancer and Correlates With Poor Prognosis

doi: 10.3389/fonc.2021.634579

Figure Lengend Snippet: miR-4700-3p suppresses drug resistance in GC (A, B) Survival of SGC7901/ADR and SGC7901/VCR cells transfected with miR-4700-3p mimics (A) and SGC7901 cells transfected with miR-4700-3p inhibitors or the corresponding negative controls was evaluated using the CCK-8 assay after step-up concentration of ADR and 5-FU treatment for 72 h. (C, D) The apoptotic rate of the indicated cells treated with 5-FU was shown. (E, F) SGC7901/ADR and SGC7901/VCR cells were cotransfected with miR-4700-3p, FOXC2 vector or their negative controls. Cell survival (E) and apoptosis rate (F) of the indicated cells treated with ADR or 5-FU, respectively, was evaluated. (G) Representative data extracted from TCGA datasets showing correlation between FENDRR (left) and FOXC2 (right) expression with miR-4700-3p expression in GC tissues (n=100). (H) Kaplan-Meier analysis of the correlation between miR-4700-3p expression and overall survival in GC patients included in the TCGA datasets (n=100). ** P < 0.01, * P < 0.05, error bars, s.d.

Article Snippet: Briefly, tissue sections were deparaffinized, subjected to antigen retrieval and endogenous peroxidase inactivation, and incubated with primary antibodies against FOXC2 (R&D Systems, AF6989) and Ki-67 (Abcam, ab15580).

Techniques: Transfection, CCK-8 Assay, Concentration Assay, Plasmid Preparation, Expressing

Journal: Frontiers in Oncology

Article Title: The FENDRR/FOXC2 Axis Contributes to Multidrug Resistance in Gastric Cancer and Correlates With Poor Prognosis

doi: 10.3389/fonc.2021.634579

Figure Lengend Snippet: Positive correlation between FENDRR and FOXC2 expression in human GC tissue samples (A) Representative images of FENDRR and FOXC2 expression in 80 paired GC and adjacent normal tissue samples detected by FISH and IHC, respectively (left). Scale bar, 200 μm (low magnification) or 50 μm (high magnification). Analysis of immunohistochemical staining for FOXC2 in 80 paired CRC specimens and matched adjacent normal tissue samples (right). (B) Correlation between the expression of FENDRR and FOXC2 in 80 GC patients. (C) Kaplan-Meier analysis of the correlation between FOXC2 expression and overall survival in patients with GC. * P < 0.05, error bars, s.d.

Article Snippet: Briefly, tissue sections were deparaffinized, subjected to antigen retrieval and endogenous peroxidase inactivation, and incubated with primary antibodies against FOXC2 (R&D Systems, AF6989) and Ki-67 (Abcam, ab15580).

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Frontiers in Oncology

Article Title: The FENDRR/FOXC2 Axis Contributes to Multidrug Resistance in Gastric Cancer and Correlates With Poor Prognosis

doi: 10.3389/fonc.2021.634579

Figure Lengend Snippet: Correlation of FOXC2 expression and patients’ clinicopathological variables in GC tissues.

Article Snippet: Briefly, tissue sections were deparaffinized, subjected to antigen retrieval and endogenous peroxidase inactivation, and incubated with primary antibodies against FOXC2 (R&D Systems, AF6989) and Ki-67 (Abcam, ab15580).

Techniques: Expressing

Journal: Developmental biology

Article Title: Loss of Flow Responsive Tie1 Results in Impaired Aortic Valve Remodeling

doi: 10.1016/j.ydbio.2019.07.011

Figure Lengend Snippet: Key Resource Table (Antibodies for IHC)

Article Snippet: Ultrastructural analysis of endothelial morphology was examined with transmission electron microscopy (EM) as described ( Odelin et al., 2014 ). table ft1 table-wrap mode="anchored" t5 caption a7 Antibodies SOURCE IDENTIFIER Working concentration Rabbit anti-pHH3 Upstate Cat#06–570; PRID:AB_310177 1:100 Rat anti-BrdU Abcam Cat#ab6326; PRID:AB_305426 1:100 Rabbit anti Cleaved Caspase-3 (Asp175) Cell Signaling Cat#9664; PRID:AB_2070042 1:400 Goat anti Tie1 R&D Systems Cat#AF619; PRID:AB_355481 1:50 Goat anti-mouse Tie2 R&D Systems Cat#AF762; PRID:AB_2203220 1:400 Rat anti-mouse CD31 Pharmingen, Cat#557355; PRID:AB_396660 1:200 Mouse anti Collagen III Abcam Cat#ab6310; PRID:AB_305413 1:400 Rabbit anti Collagen I Abcam Cat#ab34710; PRID:AB_731684 1:100 Rabbit anti-Erg1 Abcam Cat#ab92513; PRID:AB_2630401 1:100 Goat anti-VE-Cadherin R&D Systems Cat#AF1002; PRID:AB_2077789 1:400 Rabbit anti-Versican (GAGβ) Chemicon Cat#AB1033; PRID:AB_90462 1:1500 Hyaluronic acid binding protein, biotinylated (HABP) Millipore Cat#385911 1:100 Goat anti-Sox9 Cell Signaling Cat#82630 1:100 Rabbit anti Versican V0,V1 Neo (DPEAAE) ThermoFisher Cat#PA1-1748A; PRID:AB_2304324 1:1500 Sheep anti-Foxc2 R&D Systems Cat#AF6989; PRID:AB_10973139 1:200 NL557 Conjugated donkey anti-sheep IgG R&D Systems Cat#NL010; PRID:AB_884220 1:600 Open in a separate window Key Resource Table (Antibodies for IHC)

Techniques: Concentration Assay, Binding Assay

Journal: Developmental biology

Article Title: Loss of Flow Responsive Tie1 Results in Impaired Aortic Valve Remodeling

doi: 10.1016/j.ydbio.2019.07.011

Figure Lengend Snippet: Polarized expression of Tie1 and other flow sensitive endothelial markers in the remodeling aortic valves. A, Tie1 (red), Cdh5 (red), CD31 (green) and Tie2 (red) expression in remodeling aortic valves was detected by immunostaining Tie1fl/fl hearts at E14.5, E16.5, and E18.5. B and C, AoV sections of P0 Tie1fl/fl heart were co-immunostained for Tie1 (red) and CD31 (green) or Cdh5 (red) and CD31 (green). D, P0 Tie1+/lacz heart AoV sections were stained for (β-Gal, β-galactosidase) with X-gal (blue) and then with Tie2 antibody (red, E). DAPI (blue) denotes nuclei. Scale bars: 50 μm. From E16.5, Tie1 and Cdh5 begin to have higher expression on the arterial sides (arrows); in contrast, CD31 and Tie2 have higher expression on the ventricular sides (arrowheads). F, Quantification of relative expression of Tie1, Cdh5, CD31 and Tie2 in remodeling aortic valves: arterial side (predicted to face oscillatory shear stress) vs. ventricular side (predicted to face high laminar shear stress). *P<0.05, **P<0.0,1***P<0.001 (all compared with E14.5).

Article Snippet: Ultrastructural analysis of endothelial morphology was examined with transmission electron microscopy (EM) as described ( Odelin et al., 2014 ). table ft1 table-wrap mode="anchored" t5 caption a7 Antibodies SOURCE IDENTIFIER Working concentration Rabbit anti-pHH3 Upstate Cat#06–570; PRID:AB_310177 1:100 Rat anti-BrdU Abcam Cat#ab6326; PRID:AB_305426 1:100 Rabbit anti Cleaved Caspase-3 (Asp175) Cell Signaling Cat#9664; PRID:AB_2070042 1:400 Goat anti Tie1 R&D Systems Cat#AF619; PRID:AB_355481 1:50 Goat anti-mouse Tie2 R&D Systems Cat#AF762; PRID:AB_2203220 1:400 Rat anti-mouse CD31 Pharmingen, Cat#557355; PRID:AB_396660 1:200 Mouse anti Collagen III Abcam Cat#ab6310; PRID:AB_305413 1:400 Rabbit anti Collagen I Abcam Cat#ab34710; PRID:AB_731684 1:100 Rabbit anti-Erg1 Abcam Cat#ab92513; PRID:AB_2630401 1:100 Goat anti-VE-Cadherin R&D Systems Cat#AF1002; PRID:AB_2077789 1:400 Rabbit anti-Versican (GAGβ) Chemicon Cat#AB1033; PRID:AB_90462 1:1500 Hyaluronic acid binding protein, biotinylated (HABP) Millipore Cat#385911 1:100 Goat anti-Sox9 Cell Signaling Cat#82630 1:100 Rabbit anti Versican V0,V1 Neo (DPEAAE) ThermoFisher Cat#PA1-1748A; PRID:AB_2304324 1:1500 Sheep anti-Foxc2 R&D Systems Cat#AF6989; PRID:AB_10973139 1:200 NL557 Conjugated donkey anti-sheep IgG R&D Systems Cat#NL010; PRID:AB_884220 1:600 Open in a separate window Key Resource Table (Antibodies for IHC)

Techniques: Expressing, Immunostaining, Staining

Journal: Developmental biology

Article Title: Loss of Flow Responsive Tie1 Results in Impaired Aortic Valve Remodeling

doi: 10.1016/j.ydbio.2019.07.011

Figure Lengend Snippet: Nfatc1enCre mediates loxP excision exclusively in the valvular endocardium. A-D, X-gal staining of R26fslz;Nfatc1enCre heart sections shows that Nfatc1enCre-mediated recombination is restricted to the endothelium of the developing aortic valves (arrows) but not the endothelium of the artery (solid arrowheads), mesenchyme derived from EMT or ventricular endocardium (open arrowheads) at E12.5 (A), at E14.5 (B) and at P0 (C), and lacZ expression remains in the majority of the valvular endothelial cells throughout adulthood (D). E, Representative images of immuno-staining of Tie2-cko AoVs at E14.5 with anti-Tie1 antibody showing efficient specific Tie1 deletion in valvular endothelium (arrows) without change of normal expression of Tie1 in arterial endothelium (solid arrowheads) or ventricular endocardium (open arrowheads). F, Nfatc1enCre-mediated Tie1 deletion was not very efficient in a few (2 out of 9) mutant embryos. G and H, the same sections in E and F were co-stained for CD31, showing the integrity of endothelial cells. Yellow arrows indicate the mosaic Tie1 expression in the mutant valvular endothelium. DAPI, blue. Scale bars: 50μm in A-C, H-I and 200μm in D. I, The efficiency of Cre mediated Tie1 deletion in AoVs was assessed with qRT-PCR on valve leaflets of Tie1fl/fl (control) and Tie1fl/lz;Nfatc1enCre (Tie1-cko) mice at E14.5 and at P0. For normalization, Tie1 mRNA level in control mice at E14.5 was set as 1. ***P<0.001.

Article Snippet: Ultrastructural analysis of endothelial morphology was examined with transmission electron microscopy (EM) as described ( Odelin et al., 2014 ). table ft1 table-wrap mode="anchored" t5 caption a7 Antibodies SOURCE IDENTIFIER Working concentration Rabbit anti-pHH3 Upstate Cat#06–570; PRID:AB_310177 1:100 Rat anti-BrdU Abcam Cat#ab6326; PRID:AB_305426 1:100 Rabbit anti Cleaved Caspase-3 (Asp175) Cell Signaling Cat#9664; PRID:AB_2070042 1:400 Goat anti Tie1 R&D Systems Cat#AF619; PRID:AB_355481 1:50 Goat anti-mouse Tie2 R&D Systems Cat#AF762; PRID:AB_2203220 1:400 Rat anti-mouse CD31 Pharmingen, Cat#557355; PRID:AB_396660 1:200 Mouse anti Collagen III Abcam Cat#ab6310; PRID:AB_305413 1:400 Rabbit anti Collagen I Abcam Cat#ab34710; PRID:AB_731684 1:100 Rabbit anti-Erg1 Abcam Cat#ab92513; PRID:AB_2630401 1:100 Goat anti-VE-Cadherin R&D Systems Cat#AF1002; PRID:AB_2077789 1:400 Rabbit anti-Versican (GAGβ) Chemicon Cat#AB1033; PRID:AB_90462 1:1500 Hyaluronic acid binding protein, biotinylated (HABP) Millipore Cat#385911 1:100 Goat anti-Sox9 Cell Signaling Cat#82630 1:100 Rabbit anti Versican V0,V1 Neo (DPEAAE) ThermoFisher Cat#PA1-1748A; PRID:AB_2304324 1:1500 Sheep anti-Foxc2 R&D Systems Cat#AF6989; PRID:AB_10973139 1:200 NL557 Conjugated donkey anti-sheep IgG R&D Systems Cat#NL010; PRID:AB_884220 1:600 Open in a separate window Key Resource Table (Antibodies for IHC)

Techniques: Staining, Derivative Assay, Expressing, Immunostaining, Mutagenesis, Quantitative RT-PCR

Journal: Developmental biology

Article Title: Loss of Flow Responsive Tie1 Results in Impaired Aortic Valve Remodeling

doi: 10.1016/j.ydbio.2019.07.011

Figure Lengend Snippet: Key Resource Table (Antibodies for IHC)

Article Snippet: Ultrastructural analysis of endothelial morphology was examined with transmission electron microscopy (EM) as described ( Odelin et al., 2014 ). table ft1 table-wrap mode="anchored" t5 caption a7 Antibodies SOURCE IDENTIFIER Working concentration Rabbit anti-pHH3 Upstate Cat#06–570; PRID:AB_310177 1:100 Rat anti-BrdU Abcam Cat#ab6326; PRID:AB_305426 1:100 Rabbit anti Cleaved Caspase-3 (Asp175) Cell Signaling Cat#9664; PRID:AB_2070042 1:400 Goat anti Tie1 R&D Systems Cat#AF619; PRID:AB_355481 1:50 Goat anti-mouse Tie2 R&D Systems Cat#AF762; PRID:AB_2203220 1:400 Rat anti-mouse CD31 Pharmingen, Cat#557355; PRID:AB_396660 1:200 Mouse anti Collagen III Abcam Cat#ab6310; PRID:AB_305413 1:400 Rabbit anti Collagen I Abcam Cat#ab34710; PRID:AB_731684 1:100 Rabbit anti-Erg1 Abcam Cat#ab92513; PRID:AB_2630401 1:100 Goat anti-VE-Cadherin R&D Systems Cat#AF1002; PRID:AB_2077789 1:400 Rabbit anti-Versican (GAGβ) Chemicon Cat#AB1033; PRID:AB_90462 1:1500 Hyaluronic acid binding protein, biotinylated (HABP) Millipore Cat#385911 1:100 Goat anti-Sox9 Cell Signaling Cat#82630 1:100 Rabbit anti Versican V0,V1 Neo (DPEAAE) ThermoFisher Cat#PA1-1748A; PRID:AB_2304324 1:1500 Sheep anti-Foxc2 R&D Systems Cat#AF6989; PRID:AB_10973139 1:200 NL557 Conjugated donkey anti-sheep IgG R&D Systems Cat#NL010; PRID:AB_884220 1:600 Open in a separate window Key Resource Table (Antibodies for IHC)

Techniques: Concentration Assay, Binding Assay