Structured Review

Reliatech human recombinant vegf a165
Vascular alterations after intraocular <t>VEGF-A</t> injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human <t>VEGF-A165</t> (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test
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1) Product Images from "Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1"

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1

Journal: Nature Communications

doi: 10.1038/s41467-017-01738-3

Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test
Figure Legend Snippet: Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test

Techniques Used: Injection, Staining, Concentration Assay, Quantitation Assay, Immunostaining

2) Product Images from "Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis"

Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

Journal: Genes & Development

doi: 10.1101/gad.615311

Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.
Figure Legend Snippet: Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.

Techniques Used: Over Expression, Labeling, Staining, Variant Assay, CTL Assay, Injection

In vivo VEGF-C overexpression. ( A ) Adult mice are infected with either AAV-VEGF-C or control AAVs (AAV-ctl: AAV-EGFP and AAV-HSA) in the vicinity of the SVZ (coordinates from the bregma: anterior, 0.5; lateral, 1.0; depth, 2.1; n = 7 animals per group) and injected intraperitoneally with BrdU the same day. ( B ) Coronal brain cryosections from animals sacrificed at day 2 post-infection are analyzed for the number of BrdU cells (red) in the SVZ. Both the dorsolateral and ventral regions along the striatal wall show more BrdU + cells in AAV-VEGF-C-treated animals as compared with AAV-ctl. Histograms indicate the number of newborn BrdU + cells ( bottom left ) and activated Caspase-3 + cells ( bottom right ) in the SVZ. AAV-VEGF-C promotes the production of newborn cells and inhibits PCD in SVZ cells. ( C ) Coronal cryosections of the periventricular zone immunolabeled with anti-DCX Ab (green). In the striatal SVZ, DCX labeling reflects neuroblast production. AAV-VEGF-C induces a significant expansion of DCX expression in the SVZ compared with controls. ( D , E ) Coronal brain vibratome sections from Vegfr3:YFP adult mice infected with either AAV-VEGF-C or AAV-ctl and sacrificed at day 2. Dividing VEGFR-3-expressing cells (BrdU + YFP + ) are increased in AAV-VEGF-C-treated animals ( D ), while the subventricular expressions of GFAP and YFP are also enhanced ( E ), as compared with controls.(St) Striatum. Bars: B , C , 100 μm; D , E , 20 μm. Error bars indicate SEM. (***) P
Figure Legend Snippet: In vivo VEGF-C overexpression. ( A ) Adult mice are infected with either AAV-VEGF-C or control AAVs (AAV-ctl: AAV-EGFP and AAV-HSA) in the vicinity of the SVZ (coordinates from the bregma: anterior, 0.5; lateral, 1.0; depth, 2.1; n = 7 animals per group) and injected intraperitoneally with BrdU the same day. ( B ) Coronal brain cryosections from animals sacrificed at day 2 post-infection are analyzed for the number of BrdU cells (red) in the SVZ. Both the dorsolateral and ventral regions along the striatal wall show more BrdU + cells in AAV-VEGF-C-treated animals as compared with AAV-ctl. Histograms indicate the number of newborn BrdU + cells ( bottom left ) and activated Caspase-3 + cells ( bottom right ) in the SVZ. AAV-VEGF-C promotes the production of newborn cells and inhibits PCD in SVZ cells. ( C ) Coronal cryosections of the periventricular zone immunolabeled with anti-DCX Ab (green). In the striatal SVZ, DCX labeling reflects neuroblast production. AAV-VEGF-C induces a significant expansion of DCX expression in the SVZ compared with controls. ( D , E ) Coronal brain vibratome sections from Vegfr3:YFP adult mice infected with either AAV-VEGF-C or AAV-ctl and sacrificed at day 2. Dividing VEGFR-3-expressing cells (BrdU + YFP + ) are increased in AAV-VEGF-C-treated animals ( D ), while the subventricular expressions of GFAP and YFP are also enhanced ( E ), as compared with controls.(St) Striatum. Bars: B , C , 100 μm; D , E , 20 μm. Error bars indicate SEM. (***) P

Techniques Used: In Vivo, Over Expression, Mouse Assay, Infection, CTL Assay, Injection, Immunolabeling, Labeling, Expressing

Isolation of NSCs from adult Vegfr3:YFP mice. ( A ) YFP-positive and YFP-negative cells from the periventricular zone of Vegfr3:YFP mice were FACS-sorted. ( B ) The cells were tested for their ability to generate primary neurospheres in the presence of bFGF/EGF (2 × 10 4 cells per well in 24-well plates). Histograms show that YFP-positive and YFP-negative cells form primary neurospheres and include a subset of 0.5% of cell-forming spheres ( n = 6). ( C ) YFP-positive and YFP-negative cells from primary neurospheres are multipotent, differentiating into TuJ1 + neurons, GFAP + astrocytes, and O4 + oligodendrocytes after growth factor removal. Quantification is shown in the histogram ( n = 3). ( D ) YFP + /EGFR + , YFP + /EGFR − , YFP − /EGFR + , and YFP − /EGFR − subpopulations isolated after labeling with EGF-647 are tested for their capacity to self-renew and generate secondary (2 ary NS) and tertiary (3 ary NS) neurospheres. Histogram shows the increase in the number of neurosphere cells (NS-cells) relative to the number of plated cells. Only EGFR + /VEGFR-3 + cells generate numerous secondary neurospheres (2 ary NS) and tertiary neurospheres (3 ary NS) ( n = 4), even forming neurospheres after six passages (6 ary NS) (data not shown), and thus correspond to the majority of NSCs. ( E ) Primary neurospheres cultured in the presence of VEGF-C (50 ng/mL) and 31-C1 (a mouse VEGFR-3 function-blocking Ab, 5 μg/mL). The number of neurosphere formed (NS formed) is expressed as a percentage of the control. The 31-C1 Ab blocks the VEGF-C-induced amplification of neurospheres ( n = 3). ( F ) Dissociated neurosphere cells are counted and TUNEL-labeled in control or VEGF-C-containing (50 ng/mL) medium ( n = 3). ( G ) Increased proliferation of YFP + /EGFR + NSCs following VEGF-C treatment (50 ng/mL). Cells were pulsed with BrdU (10 μM) for 24 h and fixed after 2 d in vitro ( n = 3). Bar: C , 20 μm. Error bars indicate SEM. (*) P
Figure Legend Snippet: Isolation of NSCs from adult Vegfr3:YFP mice. ( A ) YFP-positive and YFP-negative cells from the periventricular zone of Vegfr3:YFP mice were FACS-sorted. ( B ) The cells were tested for their ability to generate primary neurospheres in the presence of bFGF/EGF (2 × 10 4 cells per well in 24-well plates). Histograms show that YFP-positive and YFP-negative cells form primary neurospheres and include a subset of 0.5% of cell-forming spheres ( n = 6). ( C ) YFP-positive and YFP-negative cells from primary neurospheres are multipotent, differentiating into TuJ1 + neurons, GFAP + astrocytes, and O4 + oligodendrocytes after growth factor removal. Quantification is shown in the histogram ( n = 3). ( D ) YFP + /EGFR + , YFP + /EGFR − , YFP − /EGFR + , and YFP − /EGFR − subpopulations isolated after labeling with EGF-647 are tested for their capacity to self-renew and generate secondary (2 ary NS) and tertiary (3 ary NS) neurospheres. Histogram shows the increase in the number of neurosphere cells (NS-cells) relative to the number of plated cells. Only EGFR + /VEGFR-3 + cells generate numerous secondary neurospheres (2 ary NS) and tertiary neurospheres (3 ary NS) ( n = 4), even forming neurospheres after six passages (6 ary NS) (data not shown), and thus correspond to the majority of NSCs. ( E ) Primary neurospheres cultured in the presence of VEGF-C (50 ng/mL) and 31-C1 (a mouse VEGFR-3 function-blocking Ab, 5 μg/mL). The number of neurosphere formed (NS formed) is expressed as a percentage of the control. The 31-C1 Ab blocks the VEGF-C-induced amplification of neurospheres ( n = 3). ( F ) Dissociated neurosphere cells are counted and TUNEL-labeled in control or VEGF-C-containing (50 ng/mL) medium ( n = 3). ( G ) Increased proliferation of YFP + /EGFR + NSCs following VEGF-C treatment (50 ng/mL). Cells were pulsed with BrdU (10 μM) for 24 h and fixed after 2 d in vitro ( n = 3). Bar: C , 20 μm. Error bars indicate SEM. (*) P

Techniques Used: Isolation, Mouse Assay, FACS, Labeling, Cell Culture, Blocking Assay, Amplification, TUNEL Assay, In Vitro

Proposed model for VEGF-C and VEGFR-3 function in the adult SVZ. VEGFR-3 is expressed by a subpopulation of astrocytes (B) and almost all NSCs (B1), but not by the majority of TAPs (C), neuroblasts (A), and endothelial cells (bv). EGFR is expressed by the subpopulation of VEGFR-3 NSCs. VEGF-C produced by SVZ astrocytes and also other cell types promotes activation of VEGFR-3-expressing cells (i.e., niche astrocytes and NSCs), which increase in number following overexpression of VEGF-C. This enhances the number of TAPs and neuroblasts.
Figure Legend Snippet: Proposed model for VEGF-C and VEGFR-3 function in the adult SVZ. VEGFR-3 is expressed by a subpopulation of astrocytes (B) and almost all NSCs (B1), but not by the majority of TAPs (C), neuroblasts (A), and endothelial cells (bv). EGFR is expressed by the subpopulation of VEGFR-3 NSCs. VEGF-C produced by SVZ astrocytes and also other cell types promotes activation of VEGFR-3-expressing cells (i.e., niche astrocytes and NSCs), which increase in number following overexpression of VEGF-C. This enhances the number of TAPs and neuroblasts.

Techniques Used: Produced, Activation Assay, Expressing, Over Expression

3) Product Images from "Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer"

Article Title: Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer

Journal: Pathology Oncology Research

doi: 10.1007/s12253-012-9516-x

Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )
Figure Legend Snippet: Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )

Techniques Used: Expressing

4) Product Images from "Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model"

Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/j.1582-4934.2009.00820.x

ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).
Figure Legend Snippet: ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).

Techniques Used: Enzyme-linked Immunosorbent Assay, In Vitro, Infection

Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Filtration, Mouse Assay, SDS Page, Enzyme-linked Immunosorbent Assay

Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.
Figure Legend Snippet: Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.

Techniques Used: Mouse Assay, Injection, H&E Stain, Immunohistochemistry

Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Enzyme-linked Immunosorbent Assay, Mouse Assay

5) Product Images from "Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model"

Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/j.1582-4934.2009.00820.x

ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).
Figure Legend Snippet: ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).

Techniques Used: Enzyme-linked Immunosorbent Assay, In Vitro, Infection

Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Filtration, Mouse Assay, SDS Page, Enzyme-linked Immunosorbent Assay

Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.
Figure Legend Snippet: Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.

Techniques Used: Mouse Assay, Injection, H&E Stain, Immunohistochemistry

Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Enzyme-linked Immunosorbent Assay, Mouse Assay

6) Product Images from "Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation"

Article Title: Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation

Journal: Journal of Clinical Investigation

doi: 10.1172/JCI200522037

VEGF-C and VEGF-D in M. pulmonis –infected airways. Immunohistochemical staining of VEGF-C ( A – C ) and VEGF-D ( D – F ) in mouse airways and lung 14 days after infection. ( A ) VEGF-C (green) in epithelium, peribronchial inflammatory cells,
Figure Legend Snippet: VEGF-C and VEGF-D in M. pulmonis –infected airways. Immunohistochemical staining of VEGF-C ( A – C ) and VEGF-D ( D – F ) in mouse airways and lung 14 days after infection. ( A ) VEGF-C (green) in epithelium, peribronchial inflammatory cells,

Techniques Used: Infection, Immunohistochemistry, Staining

7) Product Images from "Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1"

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1

Journal: Nature Communications

doi: 10.1038/s41467-017-01738-3

Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test
Figure Legend Snippet: Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test

Techniques Used: Injection, Staining, Concentration Assay, Quantitation Assay, Immunostaining

8) Product Images from "Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis"

Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

Journal: Genes & Development

doi: 10.1101/gad.615311

Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.
Figure Legend Snippet: Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.

Techniques Used: Over Expression, Labeling, Staining, Variant Assay, CTL Assay, Injection

9) Product Images from "Outgrowth, proliferation, viability, angiogenesis and phenotype of primary human endothelial cells in different purchasable endothelial culture media: feed wisely"

Article Title: Outgrowth, proliferation, viability, angiogenesis and phenotype of primary human endothelial cells in different purchasable endothelial culture media: feed wisely

Journal: Histochemistry and Cell Biology

doi: 10.1007/s00418-019-01815-2

Effect of different endothelial culture media on tube formation of fpEC into a fibrin matrix. a Tubes formed after 24 h in the various media supplemented with 10% NBCS and 5% human serum, either without stimulation or after stimulation with T (10 ng/ml TNFα) and TFV (10 ng/ml TNFα; 10 ng/ml FGF2; 25 ng/ml VEGF). Cells were imaged using Nikon TiE-2 microscope with Andor Zyla 4.2 sCMOS camera and Nikon Plan Apoλ 4× objective. Scale bar: 500 µm. Arrowheads show the small round tubes formed by fpEC after stimulation with TNFα alone. b Quantification of tube formation (µm/mm 2 ). c Normalisation of all treatments per medium to the mean tube length (µm/mm 2 ) of M199 medium illustrates the similarity of data. Data are given as mean ± SD of four different fpEC isolations, each performed in triplicates. a significant vs respective condition in Endopan medium, NBCS newborn calf serum
Figure Legend Snippet: Effect of different endothelial culture media on tube formation of fpEC into a fibrin matrix. a Tubes formed after 24 h in the various media supplemented with 10% NBCS and 5% human serum, either without stimulation or after stimulation with T (10 ng/ml TNFα) and TFV (10 ng/ml TNFα; 10 ng/ml FGF2; 25 ng/ml VEGF). Cells were imaged using Nikon TiE-2 microscope with Andor Zyla 4.2 sCMOS camera and Nikon Plan Apoλ 4× objective. Scale bar: 500 µm. Arrowheads show the small round tubes formed by fpEC after stimulation with TNFα alone. b Quantification of tube formation (µm/mm 2 ). c Normalisation of all treatments per medium to the mean tube length (µm/mm 2 ) of M199 medium illustrates the similarity of data. Data are given as mean ± SD of four different fpEC isolations, each performed in triplicates. a significant vs respective condition in Endopan medium, NBCS newborn calf serum

Techniques Used: Microscopy

10) Product Images from "Apelin+ Endothelial Niche Cells Control Hematopoiesis and Mediate Vascular Regeneration after Myeloablative Injury"

Article Title: Apelin+ Endothelial Niche Cells Control Hematopoiesis and Mediate Vascular Regeneration after Myeloablative Injury

Journal: Cell Stem Cell

doi: 10.1016/j.stem.2019.10.006

Crosstalk of HSPC with Apln + ECs in BM Vascular Regeneration and Hematopoietic Reconstitution (A) Diagram depicting the transplantation of wild-type Lin – cells into Ctrl and Vegfr2 iΔApln host mice. (B) Bone vessels at 2.5 weeks after transplantation in control or Vegfr2 iΔApln host mice. Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in control (n = 10–11) and Vegfr2 iΔApln (n = 11) host mice. (C) Scheme depicting VEGF-A treatment in combination with transplantation of wild-type Lin – cells. (D) Survival curve of irradiated mice transplanted with 10 4 Lin – cells and intravenous injection of PBS (vehicle; n = 35) or recombinant VEGF-A (n = 20). (E) Bone vessels at 3 weeks after irradiation and treatment with vehicle (PBS + Lin – cells) or VEGF-A (VEGF-A + Lin – cells). Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in vehicle (n = 12–14) and VEGF-A (n = 11)-treated mice. (F) Secondary long-term competitive repopulating assay with donor-derived CD45.2 cells from 1 st transplant of vehicle or VEGF-A-treated recipients. Graphs show percentage of CD45.2 donor-derived myeloid cells, B cells, and T cells (vehicle = 5, VEGF-A = 7). (G) Confocal tile scan overview and high-magnification images of GFP and Emcn signals in the Apln-mTmG diaphysis after infusion of vehicle or VEGF-A. Quantification of GFP + cell relative to Emcn + (vehicle = 3, VEGF-A = 3). (H) Apln transcripts in cultured bEnd.3 cells. Actb was used as control (vehicle = 11, VEGF-A = 11). (I) Quantification of CFU-GM number, CD45% and CD11b% in methylcellulose assays (700 Lin – HSPC were seeded). Vehicle (n = 8) or VEGF-A (4 μg/ml; n = 8) were added to standard culture medium. Error bars, mean ± SEM. p values, two-tailed unpaired Student’s t test. See also Figure S7 .
Figure Legend Snippet: Crosstalk of HSPC with Apln + ECs in BM Vascular Regeneration and Hematopoietic Reconstitution (A) Diagram depicting the transplantation of wild-type Lin – cells into Ctrl and Vegfr2 iΔApln host mice. (B) Bone vessels at 2.5 weeks after transplantation in control or Vegfr2 iΔApln host mice. Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in control (n = 10–11) and Vegfr2 iΔApln (n = 11) host mice. (C) Scheme depicting VEGF-A treatment in combination with transplantation of wild-type Lin – cells. (D) Survival curve of irradiated mice transplanted with 10 4 Lin – cells and intravenous injection of PBS (vehicle; n = 35) or recombinant VEGF-A (n = 20). (E) Bone vessels at 3 weeks after irradiation and treatment with vehicle (PBS + Lin – cells) or VEGF-A (VEGF-A + Lin – cells). Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in vehicle (n = 12–14) and VEGF-A (n = 11)-treated mice. (F) Secondary long-term competitive repopulating assay with donor-derived CD45.2 cells from 1 st transplant of vehicle or VEGF-A-treated recipients. Graphs show percentage of CD45.2 donor-derived myeloid cells, B cells, and T cells (vehicle = 5, VEGF-A = 7). (G) Confocal tile scan overview and high-magnification images of GFP and Emcn signals in the Apln-mTmG diaphysis after infusion of vehicle or VEGF-A. Quantification of GFP + cell relative to Emcn + (vehicle = 3, VEGF-A = 3). (H) Apln transcripts in cultured bEnd.3 cells. Actb was used as control (vehicle = 11, VEGF-A = 11). (I) Quantification of CFU-GM number, CD45% and CD11b% in methylcellulose assays (700 Lin – HSPC were seeded). Vehicle (n = 8) or VEGF-A (4 μg/ml; n = 8) were added to standard culture medium. Error bars, mean ± SEM. p values, two-tailed unpaired Student’s t test. See also Figure S7 .

Techniques Used: Transplantation Assay, Mouse Assay, Irradiation, Injection, Recombinant, Derivative Assay, Cell Culture, Two Tailed Test

11) Product Images from "The histone deacetylase inhibitor trichostatin a decreases lymphangiogenesis by inducing apoptosis and cell cycle arrest via p21-dependent pathways"

Article Title: The histone deacetylase inhibitor trichostatin a decreases lymphangiogenesis by inducing apoptosis and cell cycle arrest via p21-dependent pathways

Journal: BMC Cancer

doi: 10.1186/s12885-016-2807-y

Effects of HDACi on primary human lymphatic endothelial cells. a , b Cells were exposed to increasing concentrations of trichostatin A (TSA), sodium butyrate (NaB) and valproic acid (VPA) alone and ( c ) in the presence or absence of 20 ng/ml VEGF-A and 100 ng/ml VEGF-C for 24 h as indicated. Cell proliferation and viability was measured using the BrdU ( a , c ) and Alamar blue assay ( b ). Average absorbance values (mean ± SE) from 4 wells per experimental condition are displayed; data are expressed as cell proliferation and vialibity in percentage (%) with regard to solvent controls (=100%; ethanol and H 2 O). Results were confirmed in four independent sets of experiments. *p
Figure Legend Snippet: Effects of HDACi on primary human lymphatic endothelial cells. a , b Cells were exposed to increasing concentrations of trichostatin A (TSA), sodium butyrate (NaB) and valproic acid (VPA) alone and ( c ) in the presence or absence of 20 ng/ml VEGF-A and 100 ng/ml VEGF-C for 24 h as indicated. Cell proliferation and viability was measured using the BrdU ( a , c ) and Alamar blue assay ( b ). Average absorbance values (mean ± SE) from 4 wells per experimental condition are displayed; data are expressed as cell proliferation and vialibity in percentage (%) with regard to solvent controls (=100%; ethanol and H 2 O). Results were confirmed in four independent sets of experiments. *p

Techniques Used: Alamar Blue Assay

12) Product Images from "Spatial regulation of VEGF receptor endocytosis in angiogenesis"

Article Title: Spatial regulation of VEGF receptor endocytosis in angiogenesis

Journal: Nature cell biology

doi: 10.1038/ncb2679

Endothelial Dab2 and PAR-3 regulate angiogenic vessel growth a , Overview of the P6 control and Dab2 iΔEC retinal vasculature. Anti-Dab2 (white/red) staining is shown in top panels. ECs, Isolectin B4 (IB4). Bottom panels show higher magnification of the angiogenic front. b , Defects in the P6 Pard3 iΔEC retinal vasculature. Anti-PAR-3 and IB4 (green) staining in middle panels show successful deletion of PAR-3 in Pard3 iΔEC vessels. Residual round signals correspond to autofluorescent blood cells. Bottom panels show higher magnification of the angiogenic front. c, d , Quantitation of filopodia number and length, tip cell number, EC-covered area, EC proliferation and vessels branch points in Dab2 mutant (iΔEC) ( c ) or Pard3 mutant ( d ) retinas compared to the corresponding control littermates (Ctrl). Dab2 mutant n=7, Pard3 mutant n=3. Percentage of reduction is indicated. Data represent the means±s.d. P values, two-tailed Student’s t-test. e , Reduced uptake of labelled VEGF-A (red) at the Dab2 iΔEC or Pard3 iΔEC angiogenic front compared to control littermates. Green arrowheads indicate VEGF-A spots, white arrowheads marks ECs with no or little uptake. f , Statistical analysis of internalised Alexa-coupled VEGF-A in the angiogenic front and central plexus. Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test.
Figure Legend Snippet: Endothelial Dab2 and PAR-3 regulate angiogenic vessel growth a , Overview of the P6 control and Dab2 iΔEC retinal vasculature. Anti-Dab2 (white/red) staining is shown in top panels. ECs, Isolectin B4 (IB4). Bottom panels show higher magnification of the angiogenic front. b , Defects in the P6 Pard3 iΔEC retinal vasculature. Anti-PAR-3 and IB4 (green) staining in middle panels show successful deletion of PAR-3 in Pard3 iΔEC vessels. Residual round signals correspond to autofluorescent blood cells. Bottom panels show higher magnification of the angiogenic front. c, d , Quantitation of filopodia number and length, tip cell number, EC-covered area, EC proliferation and vessels branch points in Dab2 mutant (iΔEC) ( c ) or Pard3 mutant ( d ) retinas compared to the corresponding control littermates (Ctrl). Dab2 mutant n=7, Pard3 mutant n=3. Percentage of reduction is indicated. Data represent the means±s.d. P values, two-tailed Student’s t-test. e , Reduced uptake of labelled VEGF-A (red) at the Dab2 iΔEC or Pard3 iΔEC angiogenic front compared to control littermates. Green arrowheads indicate VEGF-A spots, white arrowheads marks ECs with no or little uptake. f , Statistical analysis of internalised Alexa-coupled VEGF-A in the angiogenic front and central plexus. Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test.

Techniques Used: Staining, Quantitation Assay, Mutagenesis, Two Tailed Test

Increased VEGF uptake and sprouting in the Prkci iΔEC central retina a, b , VEGF-A (red, a ) or VEGF-C ( b ) uptake in the Prkci iΔEC central plexus. Green arrowheads indicate ligand spots. Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white). c , Statistical analysis of internalised VEGFs as shown in ( a ) and ( b ). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test. d , Phenotype of the Isolectin B4-stained (IB4) P6 Prkci iΔEC retinal vasculature. Higher magnification of the angiogenic front (middle) and central plexus (bottom) are shown. e–g , Quantitation of vessels branch points, EC area and proliferation ( e , n=3), the number and length of filopodia and the number of distal sprout tips at the angiogenic front ( f , n=6), and the number of ectopic sprouts and filopodia ( g , n=6) in the central retina of Prkci mutants (iΔEC) compared to control (Ctrl) retinas. Data represent the means±s.d. P values, two-tailed Student’s t-test
Figure Legend Snippet: Increased VEGF uptake and sprouting in the Prkci iΔEC central retina a, b , VEGF-A (red, a ) or VEGF-C ( b ) uptake in the Prkci iΔEC central plexus. Green arrowheads indicate ligand spots. Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white). c , Statistical analysis of internalised VEGFs as shown in ( a ) and ( b ). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test. d , Phenotype of the Isolectin B4-stained (IB4) P6 Prkci iΔEC retinal vasculature. Higher magnification of the angiogenic front (middle) and central plexus (bottom) are shown. e–g , Quantitation of vessels branch points, EC area and proliferation ( e , n=3), the number and length of filopodia and the number of distal sprout tips at the angiogenic front ( f , n=6), and the number of ectopic sprouts and filopodia ( g , n=6) in the central retina of Prkci mutants (iΔEC) compared to control (Ctrl) retinas. Data represent the means±s.d. P values, two-tailed Student’s t-test

Techniques Used: Two Tailed Test, Staining, Quantitation Assay

Spatial differences in VEGF uptake in the retina a , Quantitation of Alexa-labelled VEGF-A in retinal ECs at 10 and 45 min after injection. The formation of intracellular spots was blocked by MiTMAB. Data represent the means±s.d. of 6 independent experiments. P values, ANOVA. b , VEGF-A uptake in blood vessels of Flk iΔEC mice. Green arrowheads indicate internalised Alexa dye-coupled VEGF-A (red). Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed student t test. c , Spatial distribution of VEGF-A or VEGF-C (red) uptake in retinal vessels. Green arrowheads indicate internalised label spots. Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white).
Figure Legend Snippet: Spatial differences in VEGF uptake in the retina a , Quantitation of Alexa-labelled VEGF-A in retinal ECs at 10 and 45 min after injection. The formation of intracellular spots was blocked by MiTMAB. Data represent the means±s.d. of 6 independent experiments. P values, ANOVA. b , VEGF-A uptake in blood vessels of Flk iΔEC mice. Green arrowheads indicate internalised Alexa dye-coupled VEGF-A (red). Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed student t test. c , Spatial distribution of VEGF-A or VEGF-C (red) uptake in retinal vessels. Green arrowheads indicate internalised label spots. Cell nuclei, DAPI (blue); ECs, Isolectin B4 (IB4, white).

Techniques Used: Quantitation Assay, Injection, Mouse Assay, Two Tailed Test

Dab2 and PAR-3 control VEGF receptor internalisation a , Alexa546-labelled VEGF-A or VEGF-C (red) accumulated in the perinuclear region of control mouse ECs at 30 min after stimulation, which was strongly reduced after knockdown of Dab2 or Pard3 . Actin, Phalloidin (green); nuclei, DAPI (blue). b, c , Quantitation of Alexa546-positive peri-nuclear VEGF-A ( b ) or VEGF-C ( c ) spots. Two different siRNAs were used for Dab2 and Pard3 in ( b ). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test. At least 100 cells were scored in each experiment. d, e , Biochemical detection of biotinylated (surface) VEGFR2 and VEGFR3 in stimulated control and Dab2 ( d ) or Pard3 ( e ) KD cells. Antibodies used for immunoblotting and molecular weight marker are indicated. f , Activation of Rac1 in control and Dab2 or Pard3 KD mouse ECs stimulated with VEGF-A or VEGF-C for 5 min, as indicated.
Figure Legend Snippet: Dab2 and PAR-3 control VEGF receptor internalisation a , Alexa546-labelled VEGF-A or VEGF-C (red) accumulated in the perinuclear region of control mouse ECs at 30 min after stimulation, which was strongly reduced after knockdown of Dab2 or Pard3 . Actin, Phalloidin (green); nuclei, DAPI (blue). b, c , Quantitation of Alexa546-positive peri-nuclear VEGF-A ( b ) or VEGF-C ( c ) spots. Two different siRNAs were used for Dab2 and Pard3 in ( b ). Data represent the means±s.d. of 6 independent experiments. P values, two-tailed Student’s t-test. At least 100 cells were scored in each experiment. d, e , Biochemical detection of biotinylated (surface) VEGFR2 and VEGFR3 in stimulated control and Dab2 ( d ) or Pard3 ( e ) KD cells. Antibodies used for immunoblotting and molecular weight marker are indicated. f , Activation of Rac1 in control and Dab2 or Pard3 KD mouse ECs stimulated with VEGF-A or VEGF-C for 5 min, as indicated.

Techniques Used: Quantitation Assay, Two Tailed Test, Molecular Weight, Marker, Activation Assay

Negative regulation of VEGF receptor internalisation by aPKC a , Phosphorylation of a GST-Dab2 fusion protein by recombinant PKCλ reduced its interaction with VEGFR3 and VEGFR2 in pull down assays. Densiometric (DM) values are shown below bands. ATP for the kinase activation was added (+) or absent (−), as indicated. CBB, Coomassie Brilliant Blue staining of GST fusion proteins. b , Western blot showing reduced Dab2 phoshorylation at Serine 24 (pS24) in cultured ECs after 30 min incubation with 5µM aPKC inhibitor. c , Association of Dab2 with immunoprecipitated VEGFR3 (anti-R3) was enhanced in Prkci iΔEC lung lysate compared to control littermates. No specific bands were immunoprecipitated with IgG. VEGFR3-associated Dab2 lacked detectable phosphorylation in Ser24 (pS24). d , Effect of aPKC inhibition on VEGF-A- or VEGF-C-induced VEGF receptor internalisation at indicated time points. Cells were preincubated with 5 µM of aPKC inhibitor for 30 min. Surface VEGFR2 (R2) and VEGFR3 (R3) were identified by biotinylation. e , Increased activation of MAP kinase (p-ERK1/2) by VEGF-A or VEGF-C after aPKC inhibition in cultured mouse ECs. Effects on AKT phosphorylation were comparably modest. Total ERK1/2 and AKT are shown as loading controls. Molecular weight marker (kD) is indicated. f , Anti-aPKC (total), anti-phospho-aPKC (p-aPKC, Thr560) and Isolectin B4 (IB4) staining of the P6 control and Prkci iΔEC retinal vasculature. g , Anti-aPKC immunostaining (red) labels ECs (IB4, green) at the angiogenic front and in the central retinal plexus. h , Phospho-aPKC (p-aPKC, Thr560) immunosignals were weak at the angiogenic front in comparison to vessels of the central plexus. Higher magnification of p-aPKC signals and merged channels in insets is shown on the right.
Figure Legend Snippet: Negative regulation of VEGF receptor internalisation by aPKC a , Phosphorylation of a GST-Dab2 fusion protein by recombinant PKCλ reduced its interaction with VEGFR3 and VEGFR2 in pull down assays. Densiometric (DM) values are shown below bands. ATP for the kinase activation was added (+) or absent (−), as indicated. CBB, Coomassie Brilliant Blue staining of GST fusion proteins. b , Western blot showing reduced Dab2 phoshorylation at Serine 24 (pS24) in cultured ECs after 30 min incubation with 5µM aPKC inhibitor. c , Association of Dab2 with immunoprecipitated VEGFR3 (anti-R3) was enhanced in Prkci iΔEC lung lysate compared to control littermates. No specific bands were immunoprecipitated with IgG. VEGFR3-associated Dab2 lacked detectable phosphorylation in Ser24 (pS24). d , Effect of aPKC inhibition on VEGF-A- or VEGF-C-induced VEGF receptor internalisation at indicated time points. Cells were preincubated with 5 µM of aPKC inhibitor for 30 min. Surface VEGFR2 (R2) and VEGFR3 (R3) were identified by biotinylation. e , Increased activation of MAP kinase (p-ERK1/2) by VEGF-A or VEGF-C after aPKC inhibition in cultured mouse ECs. Effects on AKT phosphorylation were comparably modest. Total ERK1/2 and AKT are shown as loading controls. Molecular weight marker (kD) is indicated. f , Anti-aPKC (total), anti-phospho-aPKC (p-aPKC, Thr560) and Isolectin B4 (IB4) staining of the P6 control and Prkci iΔEC retinal vasculature. g , Anti-aPKC immunostaining (red) labels ECs (IB4, green) at the angiogenic front and in the central retinal plexus. h , Phospho-aPKC (p-aPKC, Thr560) immunosignals were weak at the angiogenic front in comparison to vessels of the central plexus. Higher magnification of p-aPKC signals and merged channels in insets is shown on the right.

Techniques Used: Recombinant, Activation Assay, Staining, Western Blot, Cell Culture, Incubation, Immunoprecipitation, Inhibition, Molecular Weight, Marker, Immunostaining

13) Product Images from "Paracrine Signals From Liver Sinusoidal Endothelium Regulate Hepatitis C Virus Replication"

Article Title: Paracrine Signals From Liver Sinusoidal Endothelium Regulate Hepatitis C Virus Replication

Journal: Hepatology (Baltimore, Md.)

doi: 10.1002/hep.26571

Endothelial BMP4 expression is regulated by way of VEGFR-2. LSEC were starved of VEGF overnight and stimulated with VEGF-A, PlGF, or VEGF-E (all 10 ng/mL) for 8 hours, lysed, and RNA prepared for qRT-PCR analysis of BMP4 mRNA (A). Conditioned media (CM) from LSEC treated with the above ligands was collected and used to treat Huh-7.5 cells for 18 hours prior to infecting with HCV JFH-1 (B). Data are mean ±1 SD of n = 4 donor LSEC. Statistical comparisons were made with the Kruskal-Wallis test with Dunn's correction as appropriate and where ** P
Figure Legend Snippet: Endothelial BMP4 expression is regulated by way of VEGFR-2. LSEC were starved of VEGF overnight and stimulated with VEGF-A, PlGF, or VEGF-E (all 10 ng/mL) for 8 hours, lysed, and RNA prepared for qRT-PCR analysis of BMP4 mRNA (A). Conditioned media (CM) from LSEC treated with the above ligands was collected and used to treat Huh-7.5 cells for 18 hours prior to infecting with HCV JFH-1 (B). Data are mean ±1 SD of n = 4 donor LSEC. Statistical comparisons were made with the Kruskal-Wallis test with Dunn's correction as appropriate and where ** P

Techniques Used: Expressing, Quantitative RT-PCR

14) Product Images from "MP0250, a VEGF and HGF neutralizing DARPin® molecule shows high anti-tumor efficacy in mouse xenograft and patient-derived tumor models"

Article Title: MP0250, a VEGF and HGF neutralizing DARPin® molecule shows high anti-tumor efficacy in mouse xenograft and patient-derived tumor models

Journal: Oncotarget

doi: 10.18632/oncotarget.21738

Tumor growth inhibition in U87MG and A673 xenograft models Tumor growth inhibition in the A673 rhabdomyosarcoma xenograft model ( A, B ) and the U87MG glioblastoma model ( C, D ). Figures 2A and 2C show the anti-tumor response to MP0250, the anti-HGF DARPin ® molecule and the anti-VEGF DARPin ® molecule. Figure 2B and 2D show a dose response of MP0250. Tumor growth is plotted as mean +/- SEM.
Figure Legend Snippet: Tumor growth inhibition in U87MG and A673 xenograft models Tumor growth inhibition in the A673 rhabdomyosarcoma xenograft model ( A, B ) and the U87MG glioblastoma model ( C, D ). Figures 2A and 2C show the anti-tumor response to MP0250, the anti-HGF DARPin ® molecule and the anti-VEGF DARPin ® molecule. Figure 2B and 2D show a dose response of MP0250. Tumor growth is plotted as mean +/- SEM.

Techniques Used: Inhibition

Tumor growth inhibition in syngeneic models and anti-angiogenic effect of MP0250 Tumor growth inhibition in the orthotopic renal cancer model (RENCA-LN model) ( A, B ) and the MC38 colorectal cancer model ( C, D ). Luciferase-transfected RENCA cells were orthotopically implanted into the left kidney of BalbB mice. Tumor growth was monitored by detection of luciferase activity in vivo during the study (Figure 3A ) and determination of tumor volume at the end of the study (Figure 3B ). MP0250 was compared to sorafenib at doses indicated in the figures. Figure 3C shows the time course of the anti-tumor response to MP0250 and the HGF inhibitor and the VEGF inhibitor. Figure 3D shows the tumor volumes at the end of the study. ( E ) shows the anti-angiogenic effect of the compounds in the MC38 effect demonstrated by immuno-histochemistry for CD31. Tumor growth is plotted as mean +/− SEM.
Figure Legend Snippet: Tumor growth inhibition in syngeneic models and anti-angiogenic effect of MP0250 Tumor growth inhibition in the orthotopic renal cancer model (RENCA-LN model) ( A, B ) and the MC38 colorectal cancer model ( C, D ). Luciferase-transfected RENCA cells were orthotopically implanted into the left kidney of BalbB mice. Tumor growth was monitored by detection of luciferase activity in vivo during the study (Figure 3A ) and determination of tumor volume at the end of the study (Figure 3B ). MP0250 was compared to sorafenib at doses indicated in the figures. Figure 3C shows the time course of the anti-tumor response to MP0250 and the HGF inhibitor and the VEGF inhibitor. Figure 3D shows the tumor volumes at the end of the study. ( E ) shows the anti-angiogenic effect of the compounds in the MC38 effect demonstrated by immuno-histochemistry for CD31. Tumor growth is plotted as mean +/− SEM.

Techniques Used: Inhibition, Luciferase, Transfection, Mouse Assay, Activity Assay, In Vivo, Immunohistochemistry

MP0250 and its Inhibition of VEGF- and HGF-induced cellular functions Model of MP0250, which is composed of two human serum albumin (HSA) DARPin ® molecules flanking a VEGF and a HGF binding DARPin ® molecule ( A ). The binding of MP0250 to VEGF was determined by quantification of free hVEGF in solution after addition of increasing concentrations of MP0250 ( B ). Inhibition of VEGF-induced HUVEC proliferation was analyzed in the absence (grey triangle) or presence of increasing concentrations of MP0250. Cell growth was quantified by OD measurement representing BrdU uptake. Error bars indicate the standard deviation of independent duplicates ( C ). Competition of binding of VEGF-A to sVEGFR2-Fc ( D ) and sVEGFR1-Fc ( E ) in the presence of increasing concentrations of MP0250 (filled diamond). As a control, a non-binding DARPin ® molecule was titrated (filled grey circle).The HTRF signal was detected. Error bars indicate the standard deviation of independent triplicates (1d, 1e). Inhibition of cMET-phosphorylation in A549 cells by MP0250. Inhibition of phosphorylation as measured by ELISA measurement (OD450-620) versus the concentration of the inhibitor ( F ). Inhibition of U87MG proliferation by MP0250; error bars indicate the standard deviation of independent duplicates ( G ). Dashed black lines in the figures indicate IC50s. Data shown in the figure represents one experiment out of independent experiments as outlined in the Materials and Methods section.
Figure Legend Snippet: MP0250 and its Inhibition of VEGF- and HGF-induced cellular functions Model of MP0250, which is composed of two human serum albumin (HSA) DARPin ® molecules flanking a VEGF and a HGF binding DARPin ® molecule ( A ). The binding of MP0250 to VEGF was determined by quantification of free hVEGF in solution after addition of increasing concentrations of MP0250 ( B ). Inhibition of VEGF-induced HUVEC proliferation was analyzed in the absence (grey triangle) or presence of increasing concentrations of MP0250. Cell growth was quantified by OD measurement representing BrdU uptake. Error bars indicate the standard deviation of independent duplicates ( C ). Competition of binding of VEGF-A to sVEGFR2-Fc ( D ) and sVEGFR1-Fc ( E ) in the presence of increasing concentrations of MP0250 (filled diamond). As a control, a non-binding DARPin ® molecule was titrated (filled grey circle).The HTRF signal was detected. Error bars indicate the standard deviation of independent triplicates (1d, 1e). Inhibition of cMET-phosphorylation in A549 cells by MP0250. Inhibition of phosphorylation as measured by ELISA measurement (OD450-620) versus the concentration of the inhibitor ( F ). Inhibition of U87MG proliferation by MP0250; error bars indicate the standard deviation of independent duplicates ( G ). Dashed black lines in the figures indicate IC50s. Data shown in the figure represents one experiment out of independent experiments as outlined in the Materials and Methods section.

Techniques Used: Inhibition, Binding Assay, Standard Deviation, Enzyme-linked Immunosorbent Assay, Concentration Assay

15) Product Images from "Alternatively spliced VEGF receptor-2 is an essential endogenous inhibitor of lymphatic vessels"

Article Title: Alternatively spliced VEGF receptor-2 is an essential endogenous inhibitor of lymphatic vessels

Journal: Nature medicine

doi: 10.1038/nm.2018

Endogenous Vegf-c and sVegfr-2 selectively modulate corneal lymphangiogenesis. (a) Representative color photographs of sutured wild-type mouse corneas (top). Representative flat mounts of sutured corneas. Lyve-1 + , lymphatic vessel - green;Cd31 + /Lyve-1 – , blood vessel – red. (b) Quantification of corneal vessel area of sutured corneas of wild-type mice ( n = 8). Lyve-1 + , lymphatic vessel - white bars; Cd31 + /Lyve-1 – , blood vessel – black bars. (c) ELISA quantification of Vegf-c/VEGF-C in wild-type mouse corneas 2 and 4 days after suture placement or implantation of pellets containing 160 ng recombinant human VEGF-C. (d) Immunobloting of sVegfr-2 secreted by transfected human embryonic kidney cells (HEK), Chinese hamster ovary cells (CHO), and mouse corneal epithelial cells (KEPI) under reducing (RED) and non-reducing (NR) conditions. M, marker; lane 1. pNull; lane 2. psVegfr-2 (RED); lane 3. psVegfr-2 (NR). (e) Silver staining of polyacrylamide gel loaded with Vegfr-2/Fc and recombinant sVegfr-2 (rsVegfr-2) and resolved under reducing versus non-reducing conditions. Dim, dimer; Mon, monomer; M, marker. (f) Corneal area occupied by blood vessels (Cd31 + /Lyve-1 – ) following suture injury in wild-type mice. (g) Representative immunoblot of phosphorylated Vegfr-2 (phosVegfr-2) of porcine aortic endothelial cells stably transfected with Vegfr-2 (PAE-KDR). Reblotting for total Vegfr-2 and Vinculin are shown. ϕ, no Vegf-a. (h) Corneal area occupied by blood vessels (Cd31 + /Lyve-1 – ) following pVegf-a injection. NS, not significant; *, P
Figure Legend Snippet: Endogenous Vegf-c and sVegfr-2 selectively modulate corneal lymphangiogenesis. (a) Representative color photographs of sutured wild-type mouse corneas (top). Representative flat mounts of sutured corneas. Lyve-1 + , lymphatic vessel - green;Cd31 + /Lyve-1 – , blood vessel – red. (b) Quantification of corneal vessel area of sutured corneas of wild-type mice ( n = 8). Lyve-1 + , lymphatic vessel - white bars; Cd31 + /Lyve-1 – , blood vessel – black bars. (c) ELISA quantification of Vegf-c/VEGF-C in wild-type mouse corneas 2 and 4 days after suture placement or implantation of pellets containing 160 ng recombinant human VEGF-C. (d) Immunobloting of sVegfr-2 secreted by transfected human embryonic kidney cells (HEK), Chinese hamster ovary cells (CHO), and mouse corneal epithelial cells (KEPI) under reducing (RED) and non-reducing (NR) conditions. M, marker; lane 1. pNull; lane 2. psVegfr-2 (RED); lane 3. psVegfr-2 (NR). (e) Silver staining of polyacrylamide gel loaded with Vegfr-2/Fc and recombinant sVegfr-2 (rsVegfr-2) and resolved under reducing versus non-reducing conditions. Dim, dimer; Mon, monomer; M, marker. (f) Corneal area occupied by blood vessels (Cd31 + /Lyve-1 – ) following suture injury in wild-type mice. (g) Representative immunoblot of phosphorylated Vegfr-2 (phosVegfr-2) of porcine aortic endothelial cells stably transfected with Vegfr-2 (PAE-KDR). Reblotting for total Vegfr-2 and Vinculin are shown. ϕ, no Vegf-a. (h) Corneal area occupied by blood vessels (Cd31 + /Lyve-1 – ) following pVegf-a injection. NS, not significant; *, P

Techniques Used: Mouse Assay, Enzyme-linked Immunosorbent Assay, Recombinant, Western Blot, Transfection, Marker, Silver Staining, Stable Transfection, Injection

sVEGFR-2 exists in humans and inhibits human lymphangioma cell proliferation. (a) PCR cloning of open-reading-frame of VEGFR2 from human umbilical vein endothelial cells. (b) sVEGFR-2 immunolocalization (brown) in the human cornea using AA21129. Cell nuclei stained blue by hematoxylin. (c,d) Proliferation of lymphatic endothelial cells isolated from two children with lymphangioma, stimulated by VEGF-C and treated with sVEGFR-2 or bovine serum albumin (BSA), quantified by BrdU uptake. ( n = 6–9); c , Patient #1 is a 4-month old child; d , Patient #2 is a 10-month-old child. NS, not significant; *, P
Figure Legend Snippet: sVEGFR-2 exists in humans and inhibits human lymphangioma cell proliferation. (a) PCR cloning of open-reading-frame of VEGFR2 from human umbilical vein endothelial cells. (b) sVEGFR-2 immunolocalization (brown) in the human cornea using AA21129. Cell nuclei stained blue by hematoxylin. (c,d) Proliferation of lymphatic endothelial cells isolated from two children with lymphangioma, stimulated by VEGF-C and treated with sVEGFR-2 or bovine serum albumin (BSA), quantified by BrdU uptake. ( n = 6–9); c , Patient #1 is a 4-month old child; d , Patient #2 is a 10-month-old child. NS, not significant; *, P

Techniques Used: Polymerase Chain Reaction, Clone Assay, Staining, Isolation

Loss of endogenous sVegfr-2, which antagonizes Vegf-c, leads to spontaneous corneal lymphangiogenesis. (a) sVegfr2 mRNA detection (purple-blue) by in situ hybridization in the mouse cornea. Epi, epithelium; Str, stroma. ( b) sVegfr-2 immunolocalization (brown) in the mouse cornea using AA21127. Cell nuclei stained blue by hematoxylin. ( c) Representative corneal flat mounts of a LeCre/Vegfr2 loxP/loxP mouse ( n = 30). Lyve-1 + (green) lymphatic and Cd31 + (red) Lyve-1 – blood vessels are shown. ( d,e) Immunofluorescent detection of Lyve-1 + (green) Prox1 + (red) lymphatic vessels in corneal whole mounts ( d ) and cross sections ( e ) of LeCre/Vegfr2 loxP/loxP mice. (f) Transmission electron microscopy of lymphatic vessel in cornea of LeCre/Vegfr2 loxP/loxP mice. Inset (right). ( g ) Quantification of vessel area in corneal flat mounts ( n = 20). Error bars depict s.e.m. (h) Representative western blot of samples of recombinant sVegfr-2 (rsVegfr-2) incubated with recombinant Vegf-c (rVegf-c) and immunoprecipitated with either anti-Vegfr-2 (3 rd lane from the left) antibody or isotype control IgG (4 th lane from the left) and immunoblotted with anti-Vegf-c antibody. sVegfr-2 and Vegf-c resolution shown on first two lanes from the left. (i) Representative immunoblot of Vegf-c-induced Vegfr-3 phosphorylation of mouse lymphatic endothelial cells. Vegfr-3 re-blot is shown. ϕ, media only. (j) Proliferation of human lymphatic microvascular endothelial cells, quantified by BrdU uptake. ( f,j) , NS, not significant ;* P
Figure Legend Snippet: Loss of endogenous sVegfr-2, which antagonizes Vegf-c, leads to spontaneous corneal lymphangiogenesis. (a) sVegfr2 mRNA detection (purple-blue) by in situ hybridization in the mouse cornea. Epi, epithelium; Str, stroma. ( b) sVegfr-2 immunolocalization (brown) in the mouse cornea using AA21127. Cell nuclei stained blue by hematoxylin. ( c) Representative corneal flat mounts of a LeCre/Vegfr2 loxP/loxP mouse ( n = 30). Lyve-1 + (green) lymphatic and Cd31 + (red) Lyve-1 – blood vessels are shown. ( d,e) Immunofluorescent detection of Lyve-1 + (green) Prox1 + (red) lymphatic vessels in corneal whole mounts ( d ) and cross sections ( e ) of LeCre/Vegfr2 loxP/loxP mice. (f) Transmission electron microscopy of lymphatic vessel in cornea of LeCre/Vegfr2 loxP/loxP mice. Inset (right). ( g ) Quantification of vessel area in corneal flat mounts ( n = 20). Error bars depict s.e.m. (h) Representative western blot of samples of recombinant sVegfr-2 (rsVegfr-2) incubated with recombinant Vegf-c (rVegf-c) and immunoprecipitated with either anti-Vegfr-2 (3 rd lane from the left) antibody or isotype control IgG (4 th lane from the left) and immunoblotted with anti-Vegf-c antibody. sVegfr-2 and Vegf-c resolution shown on first two lanes from the left. (i) Representative immunoblot of Vegf-c-induced Vegfr-3 phosphorylation of mouse lymphatic endothelial cells. Vegfr-3 re-blot is shown. ϕ, media only. (j) Proliferation of human lymphatic microvascular endothelial cells, quantified by BrdU uptake. ( f,j) , NS, not significant ;* P

Techniques Used: In Situ Hybridization, Staining, Mouse Assay, Transmission Assay, Electron Microscopy, Western Blot, Recombinant, Incubation, Immunoprecipitation

16) Product Images from "Engineering Blood and Lymphatic Microvascular Networks in Fibrin Matrices"

Article Title: Engineering Blood and Lymphatic Microvascular Networks in Fibrin Matrices

Journal: Frontiers in Bioengineering and Biotechnology

doi: 10.3389/fbioe.2017.00025

Schematic representation of the separation of human dermal microvascular endothelial cells (HDMEC) into lymphatic endothelial cells (LEC) and blood vascular endothelial cells (BEC) . Scale bar represents 100 µm. (A) Brightfield microscopy image of HDMEC before MACS with representative FACS plot showing the presence of podoplanin-positive cells. Arrows indicate LEC islands within BEC populations. (B) Brightfield microcopy images and representative FACS plots of a pure LEC population. The population is positive for lymphatic markers podoplanin, LYVE-1, and vascular endothelial growth factor receptor 3 (VEGFR3). (C) Brightfield microcopy images and representative FACS plots of a pure BEC population. The population is largely negative for lymphatic markers podoplanin and LYVE-1, but positive for VEGFR3.
Figure Legend Snippet: Schematic representation of the separation of human dermal microvascular endothelial cells (HDMEC) into lymphatic endothelial cells (LEC) and blood vascular endothelial cells (BEC) . Scale bar represents 100 µm. (A) Brightfield microscopy image of HDMEC before MACS with representative FACS plot showing the presence of podoplanin-positive cells. Arrows indicate LEC islands within BEC populations. (B) Brightfield microcopy images and representative FACS plots of a pure LEC population. The population is positive for lymphatic markers podoplanin, LYVE-1, and vascular endothelial growth factor receptor 3 (VEGFR3). (C) Brightfield microcopy images and representative FACS plots of a pure BEC population. The population is largely negative for lymphatic markers podoplanin and LYVE-1, but positive for VEGFR3.

Techniques Used: Microscopy, Magnetic Cell Separation, FACS

17) Product Images from "Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model"

Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

Journal: Journal of Cellular and Molecular Medicine

doi: 10.1111/j.1582-4934.2009.00820.x

ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).
Figure Legend Snippet: ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).

Techniques Used: Enzyme-linked Immunosorbent Assay, In Vitro, Infection

Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Filtration, Mouse Assay, SDS Page, Enzyme-linked Immunosorbent Assay

Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.
Figure Legend Snippet: Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.

Techniques Used: Mouse Assay, Injection, H&E Stain, Immunohistochemistry

Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P
Figure Legend Snippet: Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P

Techniques Used: Enzyme-linked Immunosorbent Assay, Mouse Assay

18) Product Images from "Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1"

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1

Journal: Nature Communications

doi: 10.1038/s41467-017-01738-3

Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test
Figure Legend Snippet: Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test

Techniques Used: Injection, Staining, Concentration Assay, Quantitation Assay, Immunostaining

Endothelial changes after pericyte depletion. a – f Maximum intensity projection of confocal images from control and DTR iPC P6 retinas stained for IB4 (red) in combination with VEGF-A a , VEGFR2 b , VEGFR3 c , Tie2 d , Esm1 e , and Dll4 f (all in white), as indicated. Note local increase of VEGFR2, VEGFR3, and Esm1 (arrowheads in b , c , e ) but not Tie2 or VEGF-A at the edge of the vessel plexus. Dll4 expression in DTR iPC sprouts is increased in some regions (arrowheads) but absent in others (arrows). Scale bar, 100 µm. g – j Quantitation of VEGF-A immunosignals area and intensity g , signal intensity for VEGFR2 h and VEGFR3 i and proportion of Esm1+ area with respect to vascular area j in the P6 control and DTR iPC angiogenic front. Error bars, s.e.m. p -values, Student’s t -test
Figure Legend Snippet: Endothelial changes after pericyte depletion. a – f Maximum intensity projection of confocal images from control and DTR iPC P6 retinas stained for IB4 (red) in combination with VEGF-A a , VEGFR2 b , VEGFR3 c , Tie2 d , Esm1 e , and Dll4 f (all in white), as indicated. Note local increase of VEGFR2, VEGFR3, and Esm1 (arrowheads in b , c , e ) but not Tie2 or VEGF-A at the edge of the vessel plexus. Dll4 expression in DTR iPC sprouts is increased in some regions (arrowheads) but absent in others (arrows). Scale bar, 100 µm. g – j Quantitation of VEGF-A immunosignals area and intensity g , signal intensity for VEGFR2 h and VEGFR3 i and proportion of Esm1+ area with respect to vascular area j in the P6 control and DTR iPC angiogenic front. Error bars, s.e.m. p -values, Student’s t -test

Techniques Used: Staining, Expressing, Quantitation Assay

19) Product Images from "Genetic Heterogeneity of Lymphangiogenesis in Different Mouse Strains"

Article Title: Genetic Heterogeneity of Lymphangiogenesis in Different Mouse Strains

Journal: The American Journal of Pathology

doi: 10.2353/ajpath.2010.090794

Strain-dependency of lymphangiogenesis induced by VEGF-C pellets in the murine corneal micropocket assay. Lymphangiogenesis induced by the lymphangiogenic growth factor (VEGF-C) differed significantly between Balb/cAnNCrl and FVB/NCrl ( P
Figure Legend Snippet: Strain-dependency of lymphangiogenesis induced by VEGF-C pellets in the murine corneal micropocket assay. Lymphangiogenesis induced by the lymphangiogenic growth factor (VEGF-C) differed significantly between Balb/cAnNCrl and FVB/NCrl ( P

Techniques Used:

20) Product Images from "Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer"

Article Title: Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer

Journal: Pathology Oncology Research

doi: 10.1007/s12253-012-9516-x

Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )
Figure Legend Snippet: Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )

Techniques Used: Expressing

21) Product Images from "Apelin+ Endothelial Niche Cells Control Hematopoiesis and Mediate Vascular Regeneration after Myeloablative Injury"

Article Title: Apelin+ Endothelial Niche Cells Control Hematopoiesis and Mediate Vascular Regeneration after Myeloablative Injury

Journal: Cell Stem Cell

doi: 10.1016/j.stem.2019.10.006

Crosstalk of HSPC with Apln + ECs in BM Vascular Regeneration and Hematopoietic Reconstitution (A) Diagram depicting the transplantation of wild-type Lin – cells into Ctrl and Vegfr2 iΔApln host mice. (B) Bone vessels at 2.5 weeks after transplantation in control or Vegfr2 iΔApln host mice. Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in control (n = 10–11) and Vegfr2 iΔApln (n = 11) host mice. (C) Scheme depicting VEGF-A treatment in combination with transplantation of wild-type Lin – cells. (D) Survival curve of irradiated mice transplanted with 10 4 Lin – cells and intravenous injection of PBS (vehicle; n = 35) or recombinant VEGF-A (n = 20). (E) Bone vessels at 3 weeks after irradiation and treatment with vehicle (PBS + Lin – cells) or VEGF-A (VEGF-A + Lin – cells). Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in vehicle (n = 12–14) and VEGF-A (n = 11)-treated mice. (F) Secondary long-term competitive repopulating assay with donor-derived CD45.2 cells from 1 st transplant of vehicle or VEGF-A-treated recipients. Graphs show percentage of CD45.2 donor-derived myeloid cells, B cells, and T cells (vehicle = 5, VEGF-A = 7). (G) Confocal tile scan overview and high-magnification images of GFP and Emcn signals in the Apln-mTmG diaphysis after infusion of vehicle or VEGF-A. Quantification of GFP + cell relative to Emcn + (vehicle = 3, VEGF-A = 3). (H) Apln transcripts in cultured bEnd.3 cells. Actb was used as control (vehicle = 11, VEGF-A = 11). (I) Quantification of CFU-GM number, CD45% and CD11b% in methylcellulose assays (700 Lin – HSPC were seeded). Vehicle (n = 8) or VEGF-A (4 μg/ml; n = 8) were added to standard culture medium. Error bars, mean ± SEM. p values, two-tailed unpaired Student’s t test. See also Figure S7 .
Figure Legend Snippet: Crosstalk of HSPC with Apln + ECs in BM Vascular Regeneration and Hematopoietic Reconstitution (A) Diagram depicting the transplantation of wild-type Lin – cells into Ctrl and Vegfr2 iΔApln host mice. (B) Bone vessels at 2.5 weeks after transplantation in control or Vegfr2 iΔApln host mice. Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in control (n = 10–11) and Vegfr2 iΔApln (n = 11) host mice. (C) Scheme depicting VEGF-A treatment in combination with transplantation of wild-type Lin – cells. (D) Survival curve of irradiated mice transplanted with 10 4 Lin – cells and intravenous injection of PBS (vehicle; n = 35) or recombinant VEGF-A (n = 20). (E) Bone vessels at 3 weeks after irradiation and treatment with vehicle (PBS + Lin – cells) or VEGF-A (VEGF-A + Lin – cells). Quantification of Emcn + area, percentage of LSK cells, number of BMNCs, B220 + , CD11b + , and CD8 + cells in vehicle (n = 12–14) and VEGF-A (n = 11)-treated mice. (F) Secondary long-term competitive repopulating assay with donor-derived CD45.2 cells from 1 st transplant of vehicle or VEGF-A-treated recipients. Graphs show percentage of CD45.2 donor-derived myeloid cells, B cells, and T cells (vehicle = 5, VEGF-A = 7). (G) Confocal tile scan overview and high-magnification images of GFP and Emcn signals in the Apln-mTmG diaphysis after infusion of vehicle or VEGF-A. Quantification of GFP + cell relative to Emcn + (vehicle = 3, VEGF-A = 3). (H) Apln transcripts in cultured bEnd.3 cells. Actb was used as control (vehicle = 11, VEGF-A = 11). (I) Quantification of CFU-GM number, CD45% and CD11b% in methylcellulose assays (700 Lin – HSPC were seeded). Vehicle (n = 8) or VEGF-A (4 μg/ml; n = 8) were added to standard culture medium. Error bars, mean ± SEM. p values, two-tailed unpaired Student’s t test. See also Figure S7 .

Techniques Used: Transplantation Assay, Mouse Assay, Irradiation, Injection, Recombinant, Derivative Assay, Cell Culture, Two Tailed Test

22) Product Images from "Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation"

Article Title: Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation

Journal: Journal of Clinical Investigation

doi: 10.1172/JCI200522037

VEGF-C and VEGF-D in M. pulmonis –infected airways. Immunohistochemical staining of VEGF-C ( A – C ) and VEGF-D ( D – F ) in mouse airways and lung 14 days after infection. ( A ) VEGF-C (green) in epithelium, peribronchial inflammatory cells,
Figure Legend Snippet: VEGF-C and VEGF-D in M. pulmonis –infected airways. Immunohistochemical staining of VEGF-C ( A – C ) and VEGF-D ( D – F ) in mouse airways and lung 14 days after infection. ( A ) VEGF-C (green) in epithelium, peribronchial inflammatory cells,

Techniques Used: Infection, Immunohistochemistry, Staining

23) Product Images from "The histone deacetylase inhibitor trichostatin a decreases lymphangiogenesis by inducing apoptosis and cell cycle arrest via p21-dependent pathways"

Article Title: The histone deacetylase inhibitor trichostatin a decreases lymphangiogenesis by inducing apoptosis and cell cycle arrest via p21-dependent pathways

Journal: BMC Cancer

doi: 10.1186/s12885-016-2807-y

Effects of HDACi on primary human lymphatic endothelial cells. a , b Cells were exposed to increasing concentrations of trichostatin A (TSA), sodium butyrate (NaB) and valproic acid (VPA) alone and ( c ) in the presence or absence of 20 ng/ml VEGF-A and 100 ng/ml VEGF-C for 24 h as indicated. Cell proliferation and viability was measured using the BrdU ( a , c ) and Alamar blue assay ( b ). Average absorbance values (mean ± SE) from 4 wells per experimental condition are displayed; data are expressed as cell proliferation and vialibity in percentage (%) with regard to solvent controls (=100%; ethanol and H 2 O). Results were confirmed in four independent sets of experiments. *p
Figure Legend Snippet: Effects of HDACi on primary human lymphatic endothelial cells. a , b Cells were exposed to increasing concentrations of trichostatin A (TSA), sodium butyrate (NaB) and valproic acid (VPA) alone and ( c ) in the presence or absence of 20 ng/ml VEGF-A and 100 ng/ml VEGF-C for 24 h as indicated. Cell proliferation and viability was measured using the BrdU ( a , c ) and Alamar blue assay ( b ). Average absorbance values (mean ± SE) from 4 wells per experimental condition are displayed; data are expressed as cell proliferation and vialibity in percentage (%) with regard to solvent controls (=100%; ethanol and H 2 O). Results were confirmed in four independent sets of experiments. *p

Techniques Used: Alamar Blue Assay

Related Articles

Incubation:

Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model
Article Snippet: .. For IP of VEGF-A from cell lysates and liver lysates after adenovirus treatment 1–6 mg protein was incubated over 2 hr (cell lysate) or 16 hr (liver lysates) with 1 μg anti-VEGF-A antibody (MAB clone 3C5, Reliatech, Wolfenbuettel, Germany). .. For pull-down lysates were supplemented with 50 μl anti-mouse IgG agarose (Sigma) and incubated over night at 4°C.

Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model
Article Snippet: .. The membrane was blocked with 50% low-fat milk in TBS buffer and incubated with an antigen-affinity-purified VEGF-A antibody at 1 μg/ml (#2668, Reliatech) in TBS/10% low-fat milk overnight at 4°C. .. To visualize the signals, the membrane was incubated with a goat-anti rabbit IgG conjugated with horseradish peroxidase (dil.

Recombinant:

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1
Article Snippet: .. Twenty-four hours after seeding, cells were stimulated with the same media used for culture supplemented with 200 pM of human recombinant VEGF-A165 (Reliatech, 300-076), which was prepared freshly every day. .. After 1, 2, 3, or 4 days of stimulation, cells were washed once with D-PBS (Sigma, D8537) and lysed in 400 µl of RLT Plus buffer (Qiagen) supplemented with β-mercaptoethanol (10 μl ml−1 ).

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1
Article Snippet: .. VEGF-A intraocular injections Intraocular injection of human recombinant VEGF-A165 (Reliatech, 300-076) was performed in P6 pups that were anesthesized by intraperitoneal injection of xylazine (Bayer, Rompun 2%; 10 mg kg−1 ) and ketamine (Zoetis, Ketavet 100 mg ml−1 ; 100 mg kg−1 ) dissolved in saline. .. The eyelids were carefully separated using a scalpel and 0.5 µl of VEGF-A165 at a concentration of 5 μg μl−1 were injected into the vitreous humor using glass capillary pipettes with a micromanipulator (Nanoject II, Drummond Scientific).

Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis
Article Snippet: .. For stimulation with VEGFs, cells were plated in CCM containing EGF and bFGF alone or with recombinant rat VEGF-A or VEGF-C (50 ng/mL; Reliatech). .. For blocking VEGFR-3 function, rat 31C1-IgG2a Abs (5 μg/mL) were added to FACS-sorted cells cultured in the presence or absence of VEGF-C. For proliferation assays, cells were pulsed with BrdU (10 μM) for 24 h, and fixed after 2 d in vitro with 4% PFA in PBS for 15 min at room temperature.

Injection:

Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1
Article Snippet: .. VEGF-A intraocular injections Intraocular injection of human recombinant VEGF-A165 (Reliatech, 300-076) was performed in P6 pups that were anesthesized by intraperitoneal injection of xylazine (Bayer, Rompun 2%; 10 mg kg−1 ) and ketamine (Zoetis, Ketavet 100 mg ml−1 ; 100 mg kg−1 ) dissolved in saline. .. The eyelids were carefully separated using a scalpel and 0.5 µl of VEGF-A165 at a concentration of 5 μg μl−1 were injected into the vitreous humor using glass capillary pipettes with a micromanipulator (Nanoject II, Drummond Scientific).

Expressing:

Article Title: Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer
Article Snippet: .. In order to examine the expression of VEGF-C, VEGF-D and Lyve-1, mouse antibodies anti-VEGF-C (2 μg/ml), anti-VEGF-D (2 μg/ml) and anti-Lyve-1 (1 μg/ml) were used (ReliaTech GmbH, Braunschweig, Germany). ..

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  • 91
    Reliatech human recombinant vegf a165
    Vascular alterations after intraocular <t>VEGF-A</t> injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human <t>VEGF-A165</t> (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test
    Human Recombinant Vegf A165, supplied by Reliatech, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Reliatech vegf c
    Effects of <t>AAV-VEGF-A</t> and <t>AAV-VEGF-C</t> overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.
    Vegf C, supplied by Reliatech, used in various techniques. Bioz Stars score: 92/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Reliatech mouse antibodies anti vegf c
    Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of <t>VEGF-C</t> expression ( d )
    Mouse Antibodies Anti Vegf C, supplied by Reliatech, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Reliatech anti vegf a antibody
    ELISA quantification of in vitro complex formation between sFlt-1-Fc and <t>VEGF</t> in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).
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    Image Search Results


    Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test

    Journal: Nature Communications

    Article Title: Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1

    doi: 10.1038/s41467-017-01738-3

    Figure Lengend Snippet: Vascular alterations after intraocular VEGF-A injection. a Morphology of IB4-stained P6 wild-type retinal vessels at 4 h after administration of human VEGF-A165 (0.5 µl at a concentration of 5 μg μl −1 ). Note blunt appearance of the vessel front after VEGF-A injection but not for vehicle (PBS) control. Scale bar, 200 µm. b Quantitation of sprouts and filopodia at the front of the P6 vessel plexus after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. c PDGFRβ+ (green) pericytes are unaffected by short-term VEGF-A administration, whereas VEGFR2 immunosignals (white) are increased in IB4+ (red) ECs (arrowheads). Images shown correspond to insets in a . Scale bar, 100 µm. d Quantitation of VEGFR2 immunosignals intensity in the peripheral plexus of P6 retinas after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test. e Confocal images showing increased Esm1 immunostaining (white) in IB4+ (red) ECs in the peripheral plexus (arrowheads) after VEGF-A injection in P6 pups. Scale bar, 200 µm. f VEGF-A165 injection-mediated increase of Esm1 immunosignals (normalized to IB4+ EC area) in the peripheral capillary plexus but not at the edge of the angiogenic front in comparison to PBS-injected controls at P6. Error bars, s.e.m. p -values, Student’s t -test. NS, not statistically significant. g Short-term VEGF-A165 administration leads to clustering of Erg1+ (green) and IB4+ (red) ECs, as indicated, in thick sprout-like structures of P6 retinas. Panels in the center and on the right (scale bar, 20 µm) show higher magnification of the insets on the left (scale bar, 100 µm). Dashed lines in panels on the right outline IB4+ vessels. h Quantitation of EC density in the leading front vessel and emerging sprouts of the P6 angiogenic front after injection of VEGF-A165 or vehicle control. Error bars, s.e.m. p -values, Student’s t -test

    Article Snippet: Twenty-four hours after seeding, cells were stimulated with the same media used for culture supplemented with 200 pM of human recombinant VEGF-A165 (Reliatech, 300-076), which was prepared freshly every day.

    Techniques: Injection, Staining, Concentration Assay, Quantitation Assay, Immunostaining

    Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.

    Journal: Genes & Development

    Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

    doi: 10.1101/gad.615311

    Figure Lengend Snippet: Effects of AAV-VEGF-A and AAV-VEGF-C overexpression on the adult SVZ. Blood vessel pattern in the striatal wall, as shown by labeling of endothelial cells (CD31, green) and pericytes (PDGFR-β, red), while astrocytes are stained with anti-GFAP Ab (blue). AAV-VEGF-C-treated and AAV-VEGF-C156-treated (a VEGF-C variant that cannot bind to VEGFR-2) animals display a vascular network similar to AAV-ctl. In contrast, AAV-VEGF-A induced robust angiogenesis, which is attested to by a dense network of CD31 + endothelial cells and PDGFRβ + pericytes at the site of injection. Note that the number of astroglial cells increased in AAV-VEGF-C- and AAV-VEGF-C156-treated animals compared with controls. Bar, 20 μm.

    Article Snippet: For stimulation with VEGFs, cells were plated in CCM containing EGF and bFGF alone or with recombinant rat VEGF-A or VEGF-C (50 ng/mL; Reliatech).

    Techniques: Over Expression, Labeling, Staining, Variant Assay, CTL Assay, Injection

    In vivo VEGF-C overexpression. ( A ) Adult mice are infected with either AAV-VEGF-C or control AAVs (AAV-ctl: AAV-EGFP and AAV-HSA) in the vicinity of the SVZ (coordinates from the bregma: anterior, 0.5; lateral, 1.0; depth, 2.1; n = 7 animals per group) and injected intraperitoneally with BrdU the same day. ( B ) Coronal brain cryosections from animals sacrificed at day 2 post-infection are analyzed for the number of BrdU cells (red) in the SVZ. Both the dorsolateral and ventral regions along the striatal wall show more BrdU + cells in AAV-VEGF-C-treated animals as compared with AAV-ctl. Histograms indicate the number of newborn BrdU + cells ( bottom left ) and activated Caspase-3 + cells ( bottom right ) in the SVZ. AAV-VEGF-C promotes the production of newborn cells and inhibits PCD in SVZ cells. ( C ) Coronal cryosections of the periventricular zone immunolabeled with anti-DCX Ab (green). In the striatal SVZ, DCX labeling reflects neuroblast production. AAV-VEGF-C induces a significant expansion of DCX expression in the SVZ compared with controls. ( D , E ) Coronal brain vibratome sections from Vegfr3:YFP adult mice infected with either AAV-VEGF-C or AAV-ctl and sacrificed at day 2. Dividing VEGFR-3-expressing cells (BrdU + YFP + ) are increased in AAV-VEGF-C-treated animals ( D ), while the subventricular expressions of GFAP and YFP are also enhanced ( E ), as compared with controls.(St) Striatum. Bars: B , C , 100 μm; D , E , 20 μm. Error bars indicate SEM. (***) P

    Journal: Genes & Development

    Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

    doi: 10.1101/gad.615311

    Figure Lengend Snippet: In vivo VEGF-C overexpression. ( A ) Adult mice are infected with either AAV-VEGF-C or control AAVs (AAV-ctl: AAV-EGFP and AAV-HSA) in the vicinity of the SVZ (coordinates from the bregma: anterior, 0.5; lateral, 1.0; depth, 2.1; n = 7 animals per group) and injected intraperitoneally with BrdU the same day. ( B ) Coronal brain cryosections from animals sacrificed at day 2 post-infection are analyzed for the number of BrdU cells (red) in the SVZ. Both the dorsolateral and ventral regions along the striatal wall show more BrdU + cells in AAV-VEGF-C-treated animals as compared with AAV-ctl. Histograms indicate the number of newborn BrdU + cells ( bottom left ) and activated Caspase-3 + cells ( bottom right ) in the SVZ. AAV-VEGF-C promotes the production of newborn cells and inhibits PCD in SVZ cells. ( C ) Coronal cryosections of the periventricular zone immunolabeled with anti-DCX Ab (green). In the striatal SVZ, DCX labeling reflects neuroblast production. AAV-VEGF-C induces a significant expansion of DCX expression in the SVZ compared with controls. ( D , E ) Coronal brain vibratome sections from Vegfr3:YFP adult mice infected with either AAV-VEGF-C or AAV-ctl and sacrificed at day 2. Dividing VEGFR-3-expressing cells (BrdU + YFP + ) are increased in AAV-VEGF-C-treated animals ( D ), while the subventricular expressions of GFAP and YFP are also enhanced ( E ), as compared with controls.(St) Striatum. Bars: B , C , 100 μm; D , E , 20 μm. Error bars indicate SEM. (***) P

    Article Snippet: For stimulation with VEGFs, cells were plated in CCM containing EGF and bFGF alone or with recombinant rat VEGF-A or VEGF-C (50 ng/mL; Reliatech).

    Techniques: In Vivo, Over Expression, Mouse Assay, Infection, CTL Assay, Injection, Immunolabeling, Labeling, Expressing

    Isolation of NSCs from adult Vegfr3:YFP mice. ( A ) YFP-positive and YFP-negative cells from the periventricular zone of Vegfr3:YFP mice were FACS-sorted. ( B ) The cells were tested for their ability to generate primary neurospheres in the presence of bFGF/EGF (2 × 10 4 cells per well in 24-well plates). Histograms show that YFP-positive and YFP-negative cells form primary neurospheres and include a subset of 0.5% of cell-forming spheres ( n = 6). ( C ) YFP-positive and YFP-negative cells from primary neurospheres are multipotent, differentiating into TuJ1 + neurons, GFAP + astrocytes, and O4 + oligodendrocytes after growth factor removal. Quantification is shown in the histogram ( n = 3). ( D ) YFP + /EGFR + , YFP + /EGFR − , YFP − /EGFR + , and YFP − /EGFR − subpopulations isolated after labeling with EGF-647 are tested for their capacity to self-renew and generate secondary (2 ary NS) and tertiary (3 ary NS) neurospheres. Histogram shows the increase in the number of neurosphere cells (NS-cells) relative to the number of plated cells. Only EGFR + /VEGFR-3 + cells generate numerous secondary neurospheres (2 ary NS) and tertiary neurospheres (3 ary NS) ( n = 4), even forming neurospheres after six passages (6 ary NS) (data not shown), and thus correspond to the majority of NSCs. ( E ) Primary neurospheres cultured in the presence of VEGF-C (50 ng/mL) and 31-C1 (a mouse VEGFR-3 function-blocking Ab, 5 μg/mL). The number of neurosphere formed (NS formed) is expressed as a percentage of the control. The 31-C1 Ab blocks the VEGF-C-induced amplification of neurospheres ( n = 3). ( F ) Dissociated neurosphere cells are counted and TUNEL-labeled in control or VEGF-C-containing (50 ng/mL) medium ( n = 3). ( G ) Increased proliferation of YFP + /EGFR + NSCs following VEGF-C treatment (50 ng/mL). Cells were pulsed with BrdU (10 μM) for 24 h and fixed after 2 d in vitro ( n = 3). Bar: C , 20 μm. Error bars indicate SEM. (*) P

    Journal: Genes & Development

    Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

    doi: 10.1101/gad.615311

    Figure Lengend Snippet: Isolation of NSCs from adult Vegfr3:YFP mice. ( A ) YFP-positive and YFP-negative cells from the periventricular zone of Vegfr3:YFP mice were FACS-sorted. ( B ) The cells were tested for their ability to generate primary neurospheres in the presence of bFGF/EGF (2 × 10 4 cells per well in 24-well plates). Histograms show that YFP-positive and YFP-negative cells form primary neurospheres and include a subset of 0.5% of cell-forming spheres ( n = 6). ( C ) YFP-positive and YFP-negative cells from primary neurospheres are multipotent, differentiating into TuJ1 + neurons, GFAP + astrocytes, and O4 + oligodendrocytes after growth factor removal. Quantification is shown in the histogram ( n = 3). ( D ) YFP + /EGFR + , YFP + /EGFR − , YFP − /EGFR + , and YFP − /EGFR − subpopulations isolated after labeling with EGF-647 are tested for their capacity to self-renew and generate secondary (2 ary NS) and tertiary (3 ary NS) neurospheres. Histogram shows the increase in the number of neurosphere cells (NS-cells) relative to the number of plated cells. Only EGFR + /VEGFR-3 + cells generate numerous secondary neurospheres (2 ary NS) and tertiary neurospheres (3 ary NS) ( n = 4), even forming neurospheres after six passages (6 ary NS) (data not shown), and thus correspond to the majority of NSCs. ( E ) Primary neurospheres cultured in the presence of VEGF-C (50 ng/mL) and 31-C1 (a mouse VEGFR-3 function-blocking Ab, 5 μg/mL). The number of neurosphere formed (NS formed) is expressed as a percentage of the control. The 31-C1 Ab blocks the VEGF-C-induced amplification of neurospheres ( n = 3). ( F ) Dissociated neurosphere cells are counted and TUNEL-labeled in control or VEGF-C-containing (50 ng/mL) medium ( n = 3). ( G ) Increased proliferation of YFP + /EGFR + NSCs following VEGF-C treatment (50 ng/mL). Cells were pulsed with BrdU (10 μM) for 24 h and fixed after 2 d in vitro ( n = 3). Bar: C , 20 μm. Error bars indicate SEM. (*) P

    Article Snippet: For stimulation with VEGFs, cells were plated in CCM containing EGF and bFGF alone or with recombinant rat VEGF-A or VEGF-C (50 ng/mL; Reliatech).

    Techniques: Isolation, Mouse Assay, FACS, Labeling, Cell Culture, Blocking Assay, Amplification, TUNEL Assay, In Vitro

    Proposed model for VEGF-C and VEGFR-3 function in the adult SVZ. VEGFR-3 is expressed by a subpopulation of astrocytes (B) and almost all NSCs (B1), but not by the majority of TAPs (C), neuroblasts (A), and endothelial cells (bv). EGFR is expressed by the subpopulation of VEGFR-3 NSCs. VEGF-C produced by SVZ astrocytes and also other cell types promotes activation of VEGFR-3-expressing cells (i.e., niche astrocytes and NSCs), which increase in number following overexpression of VEGF-C. This enhances the number of TAPs and neuroblasts.

    Journal: Genes & Development

    Article Title: Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

    doi: 10.1101/gad.615311

    Figure Lengend Snippet: Proposed model for VEGF-C and VEGFR-3 function in the adult SVZ. VEGFR-3 is expressed by a subpopulation of astrocytes (B) and almost all NSCs (B1), but not by the majority of TAPs (C), neuroblasts (A), and endothelial cells (bv). EGFR is expressed by the subpopulation of VEGFR-3 NSCs. VEGF-C produced by SVZ astrocytes and also other cell types promotes activation of VEGFR-3-expressing cells (i.e., niche astrocytes and NSCs), which increase in number following overexpression of VEGF-C. This enhances the number of TAPs and neuroblasts.

    Article Snippet: For stimulation with VEGFs, cells were plated in CCM containing EGF and bFGF alone or with recombinant rat VEGF-A or VEGF-C (50 ng/mL; Reliatech).

    Techniques: Produced, Activation Assay, Expressing, Over Expression

    Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )

    Journal: Pathology Oncology Research

    Article Title: Angiotensin II Type 1 Receptor (AT-1R) Expression Correlates with VEGF-A and VEGF-D Expression in Invasive Ductal Breast Cancer

    doi: 10.1007/s12253-012-9516-x

    Figure Lengend Snippet: Kaplan-Meier’s diagram of survival for 102 patients, as related to intensity of AT-1R expression ( a ), primary tumour size ( b ), presence lymph node metastases ( c ) and intensity of VEGF-C expression ( d )

    Article Snippet: In order to examine the expression of VEGF-C, VEGF-D and Lyve-1, mouse antibodies anti-VEGF-C (2 μg/ml), anti-VEGF-D (2 μg/ml) and anti-Lyve-1 (1 μg/ml) were used (ReliaTech GmbH, Braunschweig, Germany).

    Techniques: Expressing

    ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

    doi: 10.1111/j.1582-4934.2009.00820.x

    Figure Lengend Snippet: ELISA quantification of in vitro complex formation between sFlt-1-Fc and VEGF in cell supernatants. Five hundred fifty-nine cells were infected with increasing amounts of Ad-sFlt-1in the absence (white bars) or in the presence of Ad-VEGF with a MOI of 10 (grey bars) or with MOI of 20 (black bars). The amounts of sFlt-1 in the supernatant 72 hrs after infection were diluted and measured by an ELISA for mouse Flt-1. Indicated are the mean values from duplicates (mean ± SEM).

    Article Snippet: For IP of VEGF-A from cell lysates and liver lysates after adenovirus treatment 1–6 mg protein was incubated over 2 hr (cell lysate) or 16 hr (liver lysates) with 1 μg anti-VEGF-A antibody (MAB clone 3C5, Reliatech, Wolfenbuettel, Germany).

    Techniques: Enzyme-linked Immunosorbent Assay, In Vitro, Infection

    Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

    doi: 10.1111/j.1582-4934.2009.00820.x

    Figure Lengend Snippet: Functionality of glomerular filtration of sFlt-1-treated mice after a 10 days time period. (A) SDS-PAGE analysis was performed with 1 μl of representative mouse urine samples. Lane 1 contains urine from untreated mice, lane 2 urine from eGFP mice (2 × 10 9 pfu) and lane 3 shows urine from mice treated with Adv for VEGF (1 × 10 8 pfu). Lane 4 and 5 contain urine from mice treated with AdvsFlt-1 (7,5 × 10 9 pfu (H) and 3.5 × 10 9 pfu (L), respectively) and lane 6 urine from a rescue experiment. The presence of large amounts of albumin around 67 kD is identified in the AdvsFlt-1-treated mouse and demonstrated damage of the kidney filter. (B) Mouse albumin was measured by ELISA in 24-h urine from untreated ( n = 6), GFP (2 × 10 9 pfu, n = 2), VEGF (1 × 10 8 pfu, n = 3) and msFlt-1- (3,5 × 10 9 pfu, n = 3) treated animals. For rescue mouse were co-treated with msFlt-1 and VEGF ( n = 2). Data are mean (±SEM) of several mice used for each experiment. * P

    Article Snippet: For IP of VEGF-A from cell lysates and liver lysates after adenovirus treatment 1–6 mg protein was incubated over 2 hr (cell lysate) or 16 hr (liver lysates) with 1 μg anti-VEGF-A antibody (MAB clone 3C5, Reliatech, Wolfenbuettel, Germany).

    Techniques: Filtration, Mouse Assay, SDS Page, Enzyme-linked Immunosorbent Assay

    Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

    doi: 10.1111/j.1582-4934.2009.00820.x

    Figure Lengend Snippet: Renal histology in rescue mice. Renal histology in mice after VEGF (1 × 10 8 pfu) and sFlt-1 (3.5 × 10 9 pfu) treatment, and in sFlt-1+VEGF groups. VEGF-treated mice show no significant lesions. Soluble Flt-1-injected mice show severe endotheliosis with occlusion of capillary lumens. Soluble Flt-1 + VEGF-treated animals show mild effects of glomerular endotheliosis and much less accumulation of fibrin and albumin seen in sFlt-1-treated animals. HE haematoxylin and eosin stain, PAS periodic acid Schiff reaction; IHC for fibrin and albumin was counterstained with haematoxylin; scale bar 50 μm.

    Article Snippet: For IP of VEGF-A from cell lysates and liver lysates after adenovirus treatment 1–6 mg protein was incubated over 2 hr (cell lysate) or 16 hr (liver lysates) with 1 μg anti-VEGF-A antibody (MAB clone 3C5, Reliatech, Wolfenbuettel, Germany).

    Techniques: Mouse Assay, Injection, H&E Stain, Immunohistochemistry

    Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Reduction of circulating soluble Flt-1 alleviates preeclampsia-like symptoms in a mouse model

    doi: 10.1111/j.1582-4934.2009.00820.x

    Figure Lengend Snippet: Quantitative measurement of mouse sFlt-1 by ELISA and detection of VEGF-A and VEGF-R in mice. In mice either untreated ( n = 4) or treated with Adv for eGFP ( n = 4) or AdvVEGF ( n = 6) endogenous sFlt-1 concentrations are under the detection level in plasma (A) and in urine (B). Mice treated with high concentrations of Adv for sFlt-1 (5–7,5 × 10 9 pfu, n = 10) show extremly high concentrations and mice treated with lower levels, which were also used for rescue experiments (2,5–3,5 × 10 9 pfu, n = 10) have a reduced level for sFlt-1 in plasma and urine. Rescue experiments with AdsFlt-1 and AdVEGF (1 × 10 8 , n = 4) have significant reduced levels in plasma und urine. Data are mean (±SEM) of several mice used for each experiment. * P

    Article Snippet: For IP of VEGF-A from cell lysates and liver lysates after adenovirus treatment 1–6 mg protein was incubated over 2 hr (cell lysate) or 16 hr (liver lysates) with 1 μg anti-VEGF-A antibody (MAB clone 3C5, Reliatech, Wolfenbuettel, Germany).

    Techniques: Enzyme-linked Immunosorbent Assay, Mouse Assay