gene exp vegfa mm01281449 m1  (Thermo Fisher)


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    Thermo Fisher gene exp vegfa mm01281449 m1
    NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , <t>vegfa</t> , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p
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    Images

    1) Product Images from "VEGFD Protects Retinal Ganglion Cells and, consequently, Capillaries against Excitotoxic Injury"

    Article Title: VEGFD Protects Retinal Ganglion Cells and, consequently, Capillaries against Excitotoxic Injury

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1016/j.omtm.2019.12.009

    NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , vegfa , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p
    Figure Legend Snippet: NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , vegfa , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p

    Techniques Used: Expressing, Quantitative RT-PCR, Injection, Immunolabeling, Mouse Assay, Labeling

    2) Product Images from "Hypoxia potentiates the cytotoxic effect of piperlongumine in pheochromocytoma models"

    Article Title: Hypoxia potentiates the cytotoxic effect of piperlongumine in pheochromocytoma models

    Journal: Oncotarget

    doi: 10.18632/oncotarget.9643

    Piperlongumine activates apoptosis and necroptosis, and inhibits cell migration and invasion A. RIP1-IP was performed on cell lysates from MPC cells treated with the indicated concentrations of PL at 21% and 1% O 2 for 24 hours and probed for RIP1 (top lane) and RIP3 (bottom lane). A representative image (n=3) is shown. B. MTT cells were treated with indicated concentrations of PL at 21% and 1% O 2 for 24 hours. Total cell lysates were analyzed by Western blot for cleaved PARP, cleaved caspase 3 and caspase 3. β-tubulin was used as a loading control. The representative image (n=3) is shown. C. 1.5 × 10 5 MPC cells were plated in the upper part of transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. D. 1.5 × 10 5 MPC cells were plated in the upper part of matrigel-coated transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. E. MPC cells were treated with indicated concentrations of PL for 24 hours. mRNA expression levels of Twist1, Vegfa, Mmp9, and Nanog were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . A graph represents data from three independent experiments as mean +/− SEM. *P
    Figure Legend Snippet: Piperlongumine activates apoptosis and necroptosis, and inhibits cell migration and invasion A. RIP1-IP was performed on cell lysates from MPC cells treated with the indicated concentrations of PL at 21% and 1% O 2 for 24 hours and probed for RIP1 (top lane) and RIP3 (bottom lane). A representative image (n=3) is shown. B. MTT cells were treated with indicated concentrations of PL at 21% and 1% O 2 for 24 hours. Total cell lysates were analyzed by Western blot for cleaved PARP, cleaved caspase 3 and caspase 3. β-tubulin was used as a loading control. The representative image (n=3) is shown. C. 1.5 × 10 5 MPC cells were plated in the upper part of transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. D. 1.5 × 10 5 MPC cells were plated in the upper part of matrigel-coated transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. E. MPC cells were treated with indicated concentrations of PL for 24 hours. mRNA expression levels of Twist1, Vegfa, Mmp9, and Nanog were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . A graph represents data from three independent experiments as mean +/− SEM. *P

    Techniques Used: Migration, MTT Assay, Western Blot, Expressing, Real-time Polymerase Chain Reaction

    Piperlongumine inhibits tumor growth and metastases formation and decreases expression levels of EMT and angiogenesis markers in vivo A. Nude female mice bearing MTT-Luc tumors were treated with 24 mg/kg/day PL (n=9) for 28 days. The control group was treated with vehicle (n=10). Tumor growth was assessed once a week. The graph shows a significant inhibition of tumor growth at all time points of treatment in the treated group (green) compared to control (red) animals. The graph represents data from 9 (treated) and 10 (vehicle) mice assessed at specific time points as mean +/− SEM. Statistical analysis was performed by Mann Whitney, U-test, at each time point. B. Representative tumors from control mice compared to the tumors from animals treated with PL. Scale bar: 1cm. C. Lungs were resected and subjected to bioluminescence imaging. The presence of lung metastases in treated mice was 46% lower than in control group. D. mRNA expression levels of Pou5f1, Twist1, Vegfa, Mmp9, and Nanog in tumors from both treated and control groups were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . The box and whiskers graphs represent data from control (n=10) and treated (n=9) groups. *P
    Figure Legend Snippet: Piperlongumine inhibits tumor growth and metastases formation and decreases expression levels of EMT and angiogenesis markers in vivo A. Nude female mice bearing MTT-Luc tumors were treated with 24 mg/kg/day PL (n=9) for 28 days. The control group was treated with vehicle (n=10). Tumor growth was assessed once a week. The graph shows a significant inhibition of tumor growth at all time points of treatment in the treated group (green) compared to control (red) animals. The graph represents data from 9 (treated) and 10 (vehicle) mice assessed at specific time points as mean +/− SEM. Statistical analysis was performed by Mann Whitney, U-test, at each time point. B. Representative tumors from control mice compared to the tumors from animals treated with PL. Scale bar: 1cm. C. Lungs were resected and subjected to bioluminescence imaging. The presence of lung metastases in treated mice was 46% lower than in control group. D. mRNA expression levels of Pou5f1, Twist1, Vegfa, Mmp9, and Nanog in tumors from both treated and control groups were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . The box and whiskers graphs represent data from control (n=10) and treated (n=9) groups. *P

    Techniques Used: Expressing, In Vivo, Mouse Assay, MTT Assay, Inhibition, MANN-WHITNEY, Imaging, Real-time Polymerase Chain Reaction

    3) Product Images from "HIF1α is Necessary for Exercise-Induced Neuroprotection while HIF2α is Needed for Dopaminergic Neuron Survival in the Substantia Nigra pars compacta"

    Article Title: HIF1α is Necessary for Exercise-Induced Neuroprotection while HIF2α is Needed for Dopaminergic Neuron Survival in the Substantia Nigra pars compacta

    Journal: Neuroscience

    doi: 10.1016/j.neuroscience.2015.03.015

    Epas1 (Hif2α) is increased in the SN with 3 months of wheel running exercise. The expression of hif1α (A), epas1 (B), egln1 (C), and vegfA (D) in the SN from WT mice after 3 months of exercise or SH was measured by qPCR. Epas1 (B) in the SN of exercised mice is significantly elevated (*= p≤0.02). mRNA levels are normalized to 18S ribosomal RNA and compared to expression in SH. Epas1 expression from mice in SH+MPTP and exercise + MPTP showed no changes, neither did hif1α , egln1 , or vegfA expression in any condition.
    Figure Legend Snippet: Epas1 (Hif2α) is increased in the SN with 3 months of wheel running exercise. The expression of hif1α (A), epas1 (B), egln1 (C), and vegfA (D) in the SN from WT mice after 3 months of exercise or SH was measured by qPCR. Epas1 (B) in the SN of exercised mice is significantly elevated (*= p≤0.02). mRNA levels are normalized to 18S ribosomal RNA and compared to expression in SH. Epas1 expression from mice in SH+MPTP and exercise + MPTP showed no changes, neither did hif1α , egln1 , or vegfA expression in any condition.

    Techniques Used: Expressing, Mouse Assay, Real-time Polymerase Chain Reaction

    Expression of hypoxia sensitive molecules in the SN is modulated by wheel running exercise. hif1a (A), epas1 (B), egln1 (C), and vegfA (D) expression from SN mRNA isolated at 1, 3, 5, 7, and 14 days of running show significant reduction at 3 days (A,B,D) and increase at 5 days (A,C) of running compared to controls in SH. *=p≤0.05, #=p≤0.01. N=17–18 for SH, 5–8 for each exercise condition. HIF1α protein from the SN (E, F) is significantly increased at running day 3 and after 21 days of running plus MPTP administration (SN taken at 2 hrs. after the last MPTP injection). *=p≤0.05. SN HIF2α protein (G, H) shows no statistically significant change with running. Error bars =± SEM. Arrows indicate 114kD, expected bands for these HIF1α and HIF2α antibodies are ~120kD. Lysate from COS-7 cells in normoxic and hypoxic conditions are used as negative and positive controls (F, H), and HIF2α human recombinant protein (H) as a positive control (~120kD). Values are expressed as percent of control relative to SH after normalization to β-actin (F, H, ~47kD) for individual samples. N= 4 SN for each condition.
    Figure Legend Snippet: Expression of hypoxia sensitive molecules in the SN is modulated by wheel running exercise. hif1a (A), epas1 (B), egln1 (C), and vegfA (D) expression from SN mRNA isolated at 1, 3, 5, 7, and 14 days of running show significant reduction at 3 days (A,B,D) and increase at 5 days (A,C) of running compared to controls in SH. *=p≤0.05, #=p≤0.01. N=17–18 for SH, 5–8 for each exercise condition. HIF1α protein from the SN (E, F) is significantly increased at running day 3 and after 21 days of running plus MPTP administration (SN taken at 2 hrs. after the last MPTP injection). *=p≤0.05. SN HIF2α protein (G, H) shows no statistically significant change with running. Error bars =± SEM. Arrows indicate 114kD, expected bands for these HIF1α and HIF2α antibodies are ~120kD. Lysate from COS-7 cells in normoxic and hypoxic conditions are used as negative and positive controls (F, H), and HIF2α human recombinant protein (H) as a positive control (~120kD). Values are expressed as percent of control relative to SH after normalization to β-actin (F, H, ~47kD) for individual samples. N= 4 SN for each condition.

    Techniques Used: Expressing, Isolation, Injection, Recombinant, Positive Control

    4) Product Images from "Pressure overload by suprarenal aortic constriction in mice leads to left ventricular hypertrophy without c-Kit expression in cardiomyocytes"

    Article Title: Pressure overload by suprarenal aortic constriction in mice leads to left ventricular hypertrophy without c-Kit expression in cardiomyocytes

    Journal: Scientific Reports

    doi: 10.1038/s41598-020-72273-3

    Pathological LV hypertrophy and perivascular fibrosis post-SAC. Pathological hypertrophy marker mRNAs were evaluated one week post-sham or -SAC surgery in adult male C57BL/6J mice (9-week-old) from ( a ) LV tissue (sham, n = 7; SAC, n = 6) or ( b ) enriched cardiomyocyte fractions (sham, n = 7; SAC, n = 8). ( c ) Representative transverse LV sections demonstrate the development of perivascular fibrosis around the coronaries (indicated by black arrows) after SAC-surgery (right) compared to sham-surgery (left). Sections were stained with Fast Green and Picrosirius Red to identify the cytoplasm and collagen, respectively. ( d ) Hif1a and Vegfa expression in cardiomyocytes one week post-SAC (sham, n = 7; SAC, n = 8). Data are presented as means ± SD, independent comparisons were made by two-tailed Student’s unpaired t-tests; *P
    Figure Legend Snippet: Pathological LV hypertrophy and perivascular fibrosis post-SAC. Pathological hypertrophy marker mRNAs were evaluated one week post-sham or -SAC surgery in adult male C57BL/6J mice (9-week-old) from ( a ) LV tissue (sham, n = 7; SAC, n = 6) or ( b ) enriched cardiomyocyte fractions (sham, n = 7; SAC, n = 8). ( c ) Representative transverse LV sections demonstrate the development of perivascular fibrosis around the coronaries (indicated by black arrows) after SAC-surgery (right) compared to sham-surgery (left). Sections were stained with Fast Green and Picrosirius Red to identify the cytoplasm and collagen, respectively. ( d ) Hif1a and Vegfa expression in cardiomyocytes one week post-SAC (sham, n = 7; SAC, n = 8). Data are presented as means ± SD, independent comparisons were made by two-tailed Student’s unpaired t-tests; *P

    Techniques Used: Marker, Mouse Assay, Staining, Expressing, Two Tailed Test

    5) Product Images from "Antitumor effect of Batf2 through IL-12 p40 up-regulation in tumor-associated macrophages"

    Article Title: Antitumor effect of Batf2 through IL-12 p40 up-regulation in tumor-associated macrophages

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1708598114

    Flow cytometric analyses of macrophages, DCs, and T cells. ( A ) A flow cytometric analysis of CD11b + F4/80 + splenocytes of tumor-bearing mice. Numbers represent the percentage of cells within the total cell population or the actual cell number. Representative plots are shown ( Left ). Data are expressed as mean ± SEM from three independent experiments ( Right , n = 4). ( B – D ) Flow cytometric analyses of DCs within the population of splenocytes from tumor-bearing mice ( B and C ) or within the B16-F1 tumors ( D ). Representative plots are shown ( Left ). Data are expressed as mean ± SEM from two independent experiments ( Right ). Numbers represent either the actual cell number or the percentage of cells within the total cell population ( B , n = 3), CD45 + cell population ( C , n = 3), or CD45 + CD11c + cell population ( D , n = 4). ( E ) Mean MFI of MHC class I, class II, CD80, and CD86 in tumor-infiltrating DCs ( n = 3). Data are expressed as mean ± SEM from two independent experiments. ( F ) The Vegfa mRNA expression in BMDMs that had been stimulated with R848 for 4 h was quantified using qPCR. Data are expressed as mean ± SEM from two independent experiments ( n = 3). ( G ) A flow cytometric analysis of CD45 − CD31 + endothelial cells in the tumor tissues from Batf2 −/− mice and WT littermates. Numbers represent the percentage of cells within the total tumor cell population. Representative plots are shown ( Left ). Data are expressed as mean ± SEM ( Right , n = 2). ( H ) Flow cytometric analyses of the T-cell populations within the spleen, blood, lymph nodes (LN), and BM of naïve or tumor-bearing mice. Numbers represent the percentage of cells within the CD3 + T-cell population. Representative plots are shown ( Left ). Data are expressed as mean ± SEM ( Right , n = 3). All experiments were performed on male littermates. In all figure parts, * P
    Figure Legend Snippet: Flow cytometric analyses of macrophages, DCs, and T cells. ( A ) A flow cytometric analysis of CD11b + F4/80 + splenocytes of tumor-bearing mice. Numbers represent the percentage of cells within the total cell population or the actual cell number. Representative plots are shown ( Left ). Data are expressed as mean ± SEM from three independent experiments ( Right , n = 4). ( B – D ) Flow cytometric analyses of DCs within the population of splenocytes from tumor-bearing mice ( B and C ) or within the B16-F1 tumors ( D ). Representative plots are shown ( Left ). Data are expressed as mean ± SEM from two independent experiments ( Right ). Numbers represent either the actual cell number or the percentage of cells within the total cell population ( B , n = 3), CD45 + cell population ( C , n = 3), or CD45 + CD11c + cell population ( D , n = 4). ( E ) Mean MFI of MHC class I, class II, CD80, and CD86 in tumor-infiltrating DCs ( n = 3). Data are expressed as mean ± SEM from two independent experiments. ( F ) The Vegfa mRNA expression in BMDMs that had been stimulated with R848 for 4 h was quantified using qPCR. Data are expressed as mean ± SEM from two independent experiments ( n = 3). ( G ) A flow cytometric analysis of CD45 − CD31 + endothelial cells in the tumor tissues from Batf2 −/− mice and WT littermates. Numbers represent the percentage of cells within the total tumor cell population. Representative plots are shown ( Left ). Data are expressed as mean ± SEM ( Right , n = 2). ( H ) Flow cytometric analyses of the T-cell populations within the spleen, blood, lymph nodes (LN), and BM of naïve or tumor-bearing mice. Numbers represent the percentage of cells within the CD3 + T-cell population. Representative plots are shown ( Left ). Data are expressed as mean ± SEM ( Right , n = 3). All experiments were performed on male littermates. In all figure parts, * P

    Techniques Used: Flow Cytometry, Mouse Assay, Expressing, Real-time Polymerase Chain Reaction

    6) Product Images from "DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression "

    Article Title: DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression

    Journal: Biology of reproduction

    doi: 10.1095/biolreprod.112.102293

    Expression of VEGFA by uNK cells ( A ) Relative V egfa mRNA expression by CD3E-IL2RB+DBA− and CD3E-IL2RB+DBA+ gd10.5 decidual cells using by qRT-PCR normalized to Hprt. Data are means±SEM from cells prepared in three independent sorting experiments. * P
    Figure Legend Snippet: Expression of VEGFA by uNK cells ( A ) Relative V egfa mRNA expression by CD3E-IL2RB+DBA− and CD3E-IL2RB+DBA+ gd10.5 decidual cells using by qRT-PCR normalized to Hprt. Data are means±SEM from cells prepared in three independent sorting experiments. * P

    Techniques Used: Expressing, Quantitative RT-PCR

    7) Product Images from "Glutathione adducts induced by ischemia and deletion of glutaredoxin-1 stabilize HIF-1α and improve limb revascularization"

    Article Title: Glutathione adducts induced by ischemia and deletion of glutaredoxin-1 stabilize HIF-1α and improve limb revascularization

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1524198113

    Regulation of HIF-1α and angiogenic genes by Glrx. ( A and B ) Glrx overexpression decreased 2-AAPA–dependent HIF-1α stabilization. Ad tet-Glrx was transfected to COS7 cells. After Glrx overexpression was induced by 1 μg doxycycline, these cells were treated with 2-AAPA. ( A ) Representative Western blot of HIF-1α, β-tubulin, and Glrx. ( B ) Densitometry analysis of HIF-1α normalized by β-tubulin ( n = 4 each group). Glrx knockdown increased HIF-1α and angiogenic genes expression. ( C and D ) Glrx knockdown by siGlrx increased HIF-1α and angiogenic genes in C2C12 cells. ( C ) Representative Western blot of HIF-1α, β-tubulin, and Glrx. ( D ) Densitometry analysis of HIF-1α ( n = 8 each group). ( E ) Relative mRNA expression of HIF-1α, ( F ) mRNA of Vegfa, Pdgfa, Pdgfb, and Fgf2 ( n = 6 each group) in siGlrx-treated C2C12 cells. ( G ) Overexpression of human Glrx effect on HIF-1α stabilization by Glrx knockdown. Glrx knockdowned C2C12 cells by siGlrx were transfected with ad human Glrx, which is resistant to siGlrx. Levels of HIF-1α, human, and mouse Glrx were analyzed by Western blotting. ( H ) Effect of HIF-1α knockdown on increment of mRNA levels of Vegfa by siGlrx. HIF-1α was knocked down by CRISPR/Cas9 in C2C12 cells. Then effect of siGlrx on mRNA expression of Vegfa was analyzed by qPCR. † P
    Figure Legend Snippet: Regulation of HIF-1α and angiogenic genes by Glrx. ( A and B ) Glrx overexpression decreased 2-AAPA–dependent HIF-1α stabilization. Ad tet-Glrx was transfected to COS7 cells. After Glrx overexpression was induced by 1 μg doxycycline, these cells were treated with 2-AAPA. ( A ) Representative Western blot of HIF-1α, β-tubulin, and Glrx. ( B ) Densitometry analysis of HIF-1α normalized by β-tubulin ( n = 4 each group). Glrx knockdown increased HIF-1α and angiogenic genes expression. ( C and D ) Glrx knockdown by siGlrx increased HIF-1α and angiogenic genes in C2C12 cells. ( C ) Representative Western blot of HIF-1α, β-tubulin, and Glrx. ( D ) Densitometry analysis of HIF-1α ( n = 8 each group). ( E ) Relative mRNA expression of HIF-1α, ( F ) mRNA of Vegfa, Pdgfa, Pdgfb, and Fgf2 ( n = 6 each group) in siGlrx-treated C2C12 cells. ( G ) Overexpression of human Glrx effect on HIF-1α stabilization by Glrx knockdown. Glrx knockdowned C2C12 cells by siGlrx were transfected with ad human Glrx, which is resistant to siGlrx. Levels of HIF-1α, human, and mouse Glrx were analyzed by Western blotting. ( H ) Effect of HIF-1α knockdown on increment of mRNA levels of Vegfa by siGlrx. HIF-1α was knocked down by CRISPR/Cas9 in C2C12 cells. Then effect of siGlrx on mRNA expression of Vegfa was analyzed by qPCR. † P

    Techniques Used: Over Expression, Transfection, Western Blot, Expressing, CRISPR, Real-time Polymerase Chain Reaction

    8) Product Images from "Different cyclical intermittent hypoxia severities have different effects on hippocampal microvasculature"

    Article Title: Different cyclical intermittent hypoxia severities have different effects on hippocampal microvasculature

    Journal: Journal of Applied Physiology

    doi: 10.1152/japplphysiol.01040.2015

    Fold change in qPCR gene expression for angiogenesis-related genes. The figure illustrates the results of qPCR analyses examining expression of three subsets of genes hypothesized to be related to angiogenesis. Results are shown as fold change in expression relative to the sham condition. Fold change is measured as 2 −ΔΔCT for each gene, where the first Δ is the average CT of the housekeeping genes, and the second Δ is the average ΔCT in the sham condition. Analyses were based on −ΔΔCT values, which are measured on a linear scale. Based on a priori hypothesized pathways, genes were separated into 3 distinct domains/subgroups. A : ligands: VEGFA , ANGPT1 , ANGPT2 , BDNF , FGF , and DLL4 . B : ligand receptors: VEGFR1 , VEGFR2 , TIE-1 , TIE-2 , TRK2 , FGFR-1 , FGFR-2 , NOTCH1 . C : transcription factors: HIF-1α , PGC-1α , and PGC-1β . Statistical significance was based on a domain-specific Bonferroni corrected α-level (equal to 0.05 divided by the number of genes in the domain). Significant or nominally significant differences were observed for VEGFA , ANGPT2 , VEGFR2 , and TIE-2 . The strongest differences among conditions were seen for VEGFA ( P
    Figure Legend Snippet: Fold change in qPCR gene expression for angiogenesis-related genes. The figure illustrates the results of qPCR analyses examining expression of three subsets of genes hypothesized to be related to angiogenesis. Results are shown as fold change in expression relative to the sham condition. Fold change is measured as 2 −ΔΔCT for each gene, where the first Δ is the average CT of the housekeeping genes, and the second Δ is the average ΔCT in the sham condition. Analyses were based on −ΔΔCT values, which are measured on a linear scale. Based on a priori hypothesized pathways, genes were separated into 3 distinct domains/subgroups. A : ligands: VEGFA , ANGPT1 , ANGPT2 , BDNF , FGF , and DLL4 . B : ligand receptors: VEGFR1 , VEGFR2 , TIE-1 , TIE-2 , TRK2 , FGFR-1 , FGFR-2 , NOTCH1 . C : transcription factors: HIF-1α , PGC-1α , and PGC-1β . Statistical significance was based on a domain-specific Bonferroni corrected α-level (equal to 0.05 divided by the number of genes in the domain). Significant or nominally significant differences were observed for VEGFA , ANGPT2 , VEGFR2 , and TIE-2 . The strongest differences among conditions were seen for VEGFA ( P

    Techniques Used: Real-time Polymerase Chain Reaction, Expressing, Pyrolysis Gas Chromatography

    9) Product Images from "IL-10-dependent Tr1 cells attenuate astrocyte activation and ameliorate chronic central nervous system inflammation"

    Article Title: IL-10-dependent Tr1 cells attenuate astrocyte activation and ameliorate chronic central nervous system inflammation

    Journal: Brain

    doi: 10.1093/brain/aww113

    Nasal anti-CD3 suppresses astrocyte activation. ( A–E ) Naïve or EAE NOD mice treated with CD3-specific or isotype control mAbs during the chronic phase as in ( Fig. 1 A). ( A ) Heat map depicting mRNA expression, as detected by Nanostring nCounter analysis, in astrocytes isolated from naïve or EAE NOD mice treated with CD3 specific or isotype control mAbs. Upper panels : Histogram presentation of normalized gene expression in each gene cluster. Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( B ) Quantitative PCR analysis of Ccl2 , Ccl5 , Il6 , Tlr2 , and Aqp4 expression in astrocytes isolated from naïve and EAE NOD mice treated as in A ; expression is presented relative to Gapdh . Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( C ) Immunohistochemistry analysis of spinal cords for aquaporin 4 (AQP4) expression, and the presence of astrocytes (GFAP) on subsequent sections. Scale bar = 50 µm. Data are representative of two independent experiments with n = 6 mice/group. ( D ) Relative expression (to NOD naïve group) of genes associated with the control of demyelination in astrocytes isolated from EAE NOD mice treated with anti-CD3 or isotype control. Representative data of three independent experiments. Statistical analysis by Student’s t- test. ( E ) Quantitative PCR analysis of Mmp3 , Mmp9 and Vegfa expression in astrocytes isolated from naïve and EAE NOD mice treated as in A ; expression is presented relative to Gapdh . Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( F ) Cultured astrocytes were pre-treated for 1 h with IL-10 (100 ng/ml), or vehicle (PBS), followed by activation with IL1β (20 ng/ml) or left untreated. Quantitative PCR analysis of Ccl2 , Ccl5 , Csf2 , Nos2 , Mmp3 , and Mmp9 ; expression is presented as fold change from untreated, relative to Gapdh . Data from five independent experiments (mean and SEM). Statistical analysis by two-way ANOVA, followed by Tukey post hoc analysis. * P
    Figure Legend Snippet: Nasal anti-CD3 suppresses astrocyte activation. ( A–E ) Naïve or EAE NOD mice treated with CD3-specific or isotype control mAbs during the chronic phase as in ( Fig. 1 A). ( A ) Heat map depicting mRNA expression, as detected by Nanostring nCounter analysis, in astrocytes isolated from naïve or EAE NOD mice treated with CD3 specific or isotype control mAbs. Upper panels : Histogram presentation of normalized gene expression in each gene cluster. Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( B ) Quantitative PCR analysis of Ccl2 , Ccl5 , Il6 , Tlr2 , and Aqp4 expression in astrocytes isolated from naïve and EAE NOD mice treated as in A ; expression is presented relative to Gapdh . Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( C ) Immunohistochemistry analysis of spinal cords for aquaporin 4 (AQP4) expression, and the presence of astrocytes (GFAP) on subsequent sections. Scale bar = 50 µm. Data are representative of two independent experiments with n = 6 mice/group. ( D ) Relative expression (to NOD naïve group) of genes associated with the control of demyelination in astrocytes isolated from EAE NOD mice treated with anti-CD3 or isotype control. Representative data of three independent experiments. Statistical analysis by Student’s t- test. ( E ) Quantitative PCR analysis of Mmp3 , Mmp9 and Vegfa expression in astrocytes isolated from naïve and EAE NOD mice treated as in A ; expression is presented relative to Gapdh . Representative data of three independent experiments, statistical analysis by one-way ANOVA, followed by Tukey post hoc analysis. ( F ) Cultured astrocytes were pre-treated for 1 h with IL-10 (100 ng/ml), or vehicle (PBS), followed by activation with IL1β (20 ng/ml) or left untreated. Quantitative PCR analysis of Ccl2 , Ccl5 , Csf2 , Nos2 , Mmp3 , and Mmp9 ; expression is presented as fold change from untreated, relative to Gapdh . Data from five independent experiments (mean and SEM). Statistical analysis by two-way ANOVA, followed by Tukey post hoc analysis. * P

    Techniques Used: Activation Assay, Mouse Assay, Expressing, Isolation, Real-time Polymerase Chain Reaction, Immunohistochemistry, Cell Culture

    10) Product Images from "Angiogenesis is inhibitory for mammalian digit regeneration"

    Article Title: Angiogenesis is inhibitory for mammalian digit regeneration

    Journal: Regeneration

    doi: 10.1002/reg2.24

    VEGF treatment inhibits digit tip regeneration (top is dorsal and proximal is to the left). (A) At 10 DPA when redifferentiation has initiated, transcripts for the anti‐angiogenic factor Pedf are abundant in both the digit stump (arrow) and blastema (arrowhead). (B) During wound healing Vegfa expression is not detected in either the stump or wound bed at 5 DPA. (C) At 9 DPA, transcripts for Vegfa can be detected in the digit stump (arrow) but not the blastema. (D) Immunohistochemical staining for von Willebrand factor (VWF, red) at 7 DPA showing endothelial cells (arrow) in the blastema. (E) VWF staining of a control sample treated with a BSA bead (*) showing endothelial cells (arrow) in the blastema. (F) VWF staining of an experimental digit treated with a VEGF bead (*) showing an enhanced population of endothelial cells (arrow) associated with the bead. Sections shown in (D)−(F) were counterstained with DAPI. (G) Alizarin Red whole mount staining of 14 DPI digits showing that regeneration is not altered by control BSA treated bead implantation. (H) Based on whole mount staining the majority of digits fail to regenerate following treatment with a VEGF microcarrier bead (*). (I) Some VEGF bead treated digits form partial regenerates that include bony spikes that extend distally from the digit stump (arrow). (J) MicroCT rendering of the P3 skeletal element shown in (G). (K) MicroCT rendering of the P3 element shown in (H). (L) Graph showing normalized volume measurements from microCT analyses of control BSA treated (blue), VEGF treated (red) and BMP9 treated (green) digits at 14 DPI.
    Figure Legend Snippet: VEGF treatment inhibits digit tip regeneration (top is dorsal and proximal is to the left). (A) At 10 DPA when redifferentiation has initiated, transcripts for the anti‐angiogenic factor Pedf are abundant in both the digit stump (arrow) and blastema (arrowhead). (B) During wound healing Vegfa expression is not detected in either the stump or wound bed at 5 DPA. (C) At 9 DPA, transcripts for Vegfa can be detected in the digit stump (arrow) but not the blastema. (D) Immunohistochemical staining for von Willebrand factor (VWF, red) at 7 DPA showing endothelial cells (arrow) in the blastema. (E) VWF staining of a control sample treated with a BSA bead (*) showing endothelial cells (arrow) in the blastema. (F) VWF staining of an experimental digit treated with a VEGF bead (*) showing an enhanced population of endothelial cells (arrow) associated with the bead. Sections shown in (D)−(F) were counterstained with DAPI. (G) Alizarin Red whole mount staining of 14 DPI digits showing that regeneration is not altered by control BSA treated bead implantation. (H) Based on whole mount staining the majority of digits fail to regenerate following treatment with a VEGF microcarrier bead (*). (I) Some VEGF bead treated digits form partial regenerates that include bony spikes that extend distally from the digit stump (arrow). (J) MicroCT rendering of the P3 skeletal element shown in (G). (K) MicroCT rendering of the P3 element shown in (H). (L) Graph showing normalized volume measurements from microCT analyses of control BSA treated (blue), VEGF treated (red) and BMP9 treated (green) digits at 14 DPI.

    Techniques Used: Expressing, Immunohistochemistry, Staining

    BMP9 treatment inhibits digit tip regeneration (top is dorsal and proximal is to the left). (A), (B) Alizarin Red whole mount stained digit at 14 DPI treated with a BMP9 microcarrier bead (*). Truncated digit tips formed in approximately half of the BMP9 treated digits (A), whereas the remaining digits formed a shortened but anatomically distinct digit tip suggestive of a remodeling response (B). (C) The majority of BMP9 treated digit tips analyzed at 42 DPI formed shortened digit tips. (D)−(G) Section in situ hybridization studies to localize Vegfa transcripts after treatment with a control BSA or a BMP9 microcarrier bead. (D) Vegfa expression was not detected at 1 DPI in BSA treated control digits. (E) Following BMP9 treatment Vegfa transcripts were abundant in the digit stump (arrowhead) but not directly associated with the microcarrier bead (*). (F) At 7 DPI Vegfa expression was detected in BSA treated control digits at a level similar to that of untreated regenerates at a similar stage (see Fig. 1 C). (G) Vegfa expression remained upregulated (arrowhead) in the digit stump and proximal blastema 7 days after treatment with BMP9.
    Figure Legend Snippet: BMP9 treatment inhibits digit tip regeneration (top is dorsal and proximal is to the left). (A), (B) Alizarin Red whole mount stained digit at 14 DPI treated with a BMP9 microcarrier bead (*). Truncated digit tips formed in approximately half of the BMP9 treated digits (A), whereas the remaining digits formed a shortened but anatomically distinct digit tip suggestive of a remodeling response (B). (C) The majority of BMP9 treated digit tips analyzed at 42 DPI formed shortened digit tips. (D)−(G) Section in situ hybridization studies to localize Vegfa transcripts after treatment with a control BSA or a BMP9 microcarrier bead. (D) Vegfa expression was not detected at 1 DPI in BSA treated control digits. (E) Following BMP9 treatment Vegfa transcripts were abundant in the digit stump (arrowhead) but not directly associated with the microcarrier bead (*). (F) At 7 DPI Vegfa expression was detected in BSA treated control digits at a level similar to that of untreated regenerates at a similar stage (see Fig. 1 C). (G) Vegfa expression remained upregulated (arrowhead) in the digit stump and proximal blastema 7 days after treatment with BMP9.

    Techniques Used: Staining, In Situ Hybridization, Expressing

    11) Product Images from "Brivanib Attenuates Hepatic Fibrosis In Vivo and Stellate Cell Activation In Vitro by Inhibition of FGF, VEGF and PDGF Signaling"

    Article Title: Brivanib Attenuates Hepatic Fibrosis In Vivo and Stellate Cell Activation In Vitro by Inhibition of FGF, VEGF and PDGF Signaling

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0092273

    Brivanib does not affect the transcription of growth factors and their receptors in thioacetamide treated mice. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and TAA mice treated with no brivanib or with brivanib 25/kg, 50 mg/kg, or 100 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. mRNA levels of PDGF , PDGFBR , TGFB1 , TGFBR2 , FGF2 , FGFR2 , VEGFA , and VEGFR2 are not affected by brivanib treatment of TAA mice.
    Figure Legend Snippet: Brivanib does not affect the transcription of growth factors and their receptors in thioacetamide treated mice. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and TAA mice treated with no brivanib or with brivanib 25/kg, 50 mg/kg, or 100 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. mRNA levels of PDGF , PDGFBR , TGFB1 , TGFBR2 , FGF2 , FGFR2 , VEGFA , and VEGFR2 are not affected by brivanib treatment of TAA mice.

    Techniques Used: Mouse Assay, Real-time Polymerase Chain Reaction

    Brivanib stimulates the transcription of growth factors and their receptors in bile duct ligated mice. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and BDL mice treated with no brivanib or with brivanib 25/kg or 50 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. (A, B) BDL mice treated with brivanib show a dose-dependent increase in mRNA levels of PDGFB and PDGFRB . (C, D) mRNA levels of TGFB1 increases as the concentration of brivanib increases in BDL mice; mRNA levels of TGFBR2 are higher in BDL mice treated with 25 mg/kg and 50 mg/kg of brivanib, compared to the no brivanib group. (E, F) mRNA levels of FGF2 and FGFR2 are slightly higher with brivanib compared to no brivanib in BDL mice. (G, H) the mRNA levels of VEGFA and VEGFR2 are not affected by brivanib treatment in BDL mice.
    Figure Legend Snippet: Brivanib stimulates the transcription of growth factors and their receptors in bile duct ligated mice. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and BDL mice treated with no brivanib or with brivanib 25/kg or 50 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. (A, B) BDL mice treated with brivanib show a dose-dependent increase in mRNA levels of PDGFB and PDGFRB . (C, D) mRNA levels of TGFB1 increases as the concentration of brivanib increases in BDL mice; mRNA levels of TGFBR2 are higher in BDL mice treated with 25 mg/kg and 50 mg/kg of brivanib, compared to the no brivanib group. (E, F) mRNA levels of FGF2 and FGFR2 are slightly higher with brivanib compared to no brivanib in BDL mice. (G, H) the mRNA levels of VEGFA and VEGFR2 are not affected by brivanib treatment in BDL mice.

    Techniques Used: Mouse Assay, Real-time Polymerase Chain Reaction, Concentration Assay

    Brivanib decreases the transcription of growth factors and their receptors in carbon tetrachloride. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and CCl 4 mice treated with no brivanib or with brivanib 25 mg/kg, 50 mg/kg, or 100 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. (A) mRNA levels of PDGFB are not affected by brivanib treatment in CCl 4 . (B) mRNA levels of PDGFRB decrease as the concentration of brivanib increases in CCl 4 . (C) mRNA levels of TGFB1 decrease as the concentration of brivanib increases in CCl 4 . (D) mRNA levels of TGFBR2 are lower in brivanib-treated CCl 4 mice compared to no brivanib. (E) mRNA levels of FGF2 are not affected by brivanib treatment in CCl 4 . (F) mRNA levels of FGFR2 are lower in brivanib-treated CCl 4 mice compared to no brivanib. (G, H) mRNA levels of VEGFA and VEGFR2 decrease as the concentration of brivanib increases in CCl 4 mice.
    Figure Legend Snippet: Brivanib decreases the transcription of growth factors and their receptors in carbon tetrachloride. Hepatic levels of growth factors and growth factor receptor mRNA were measured by real time PCR in sham and CCl 4 mice treated with no brivanib or with brivanib 25 mg/kg, 50 mg/kg, or 100 mg/kg. In all of the sham experiments, higher concentrations of brivanib decreased the mRNA levels of the growth factors and their receptors. (A) mRNA levels of PDGFB are not affected by brivanib treatment in CCl 4 . (B) mRNA levels of PDGFRB decrease as the concentration of brivanib increases in CCl 4 . (C) mRNA levels of TGFB1 decrease as the concentration of brivanib increases in CCl 4 . (D) mRNA levels of TGFBR2 are lower in brivanib-treated CCl 4 mice compared to no brivanib. (E) mRNA levels of FGF2 are not affected by brivanib treatment in CCl 4 . (F) mRNA levels of FGFR2 are lower in brivanib-treated CCl 4 mice compared to no brivanib. (G, H) mRNA levels of VEGFA and VEGFR2 decrease as the concentration of brivanib increases in CCl 4 mice.

    Techniques Used: Real-time Polymerase Chain Reaction, Mouse Assay, Concentration Assay

    12) Product Images from "Obesity‐induced reduction of adipose eosinophils is reversed with low‐calorie dietary intervention . Obesity‐induced reduction of adipose eosinophils is reversed with low‐calorie dietary intervention"

    Article Title: Obesity‐induced reduction of adipose eosinophils is reversed with low‐calorie dietary intervention . Obesity‐induced reduction of adipose eosinophils is reversed with low‐calorie dietary intervention

    Journal: Physiological Reports

    doi: 10.14814/phy2.13919

    Inflammatory and tissue remodeling gene expression profile of adipose tissue during diet‐induced weight loss. (A–C) Eosinophil‐associated genes during dietary weight loss, including (A) Siglecf , (B) Prg2 , (C) Ccr3 . Macrophage marker (D) Emr1 , proinflammatory macrophage markers (E) Tnfa and (F) Itgax and anti‐inflammatory macrophage marker (G) Arg1 . Tissue remodeling factors (H) Mmp9 , (I) Vegfa , and (J) Fgf2 . Data are presented as mean ± SEM of 4–6 mice/group. * P
    Figure Legend Snippet: Inflammatory and tissue remodeling gene expression profile of adipose tissue during diet‐induced weight loss. (A–C) Eosinophil‐associated genes during dietary weight loss, including (A) Siglecf , (B) Prg2 , (C) Ccr3 . Macrophage marker (D) Emr1 , proinflammatory macrophage markers (E) Tnfa and (F) Itgax and anti‐inflammatory macrophage marker (G) Arg1 . Tissue remodeling factors (H) Mmp9 , (I) Vegfa , and (J) Fgf2 . Data are presented as mean ± SEM of 4–6 mice/group. * P

    Techniques Used: Expressing, Marker, Mouse Assay

    13) Product Images from "Attenuation of periostin in retinal Müller glia by TNF-α and IFN-γ"

    Article Title: Attenuation of periostin in retinal Müller glia by TNF-α and IFN-γ

    Journal: International Journal of Ophthalmology

    doi: 10.18240/ijo.2019.02.05

    Periostin-knockdown attenuated VEGFA mRNA expression levels in MIO-M1 cells qRT-PCR results showed in bar graph revealed downregulation in MIO-M1 cells of periostin (A) and VEGFA (B) at mRNA expression levels after periostin knockdown. Results were normalized to human GAPDH mRNA expression levels correspondingly, and data were represented the mean±SEM ( n =4). b P
    Figure Legend Snippet: Periostin-knockdown attenuated VEGFA mRNA expression levels in MIO-M1 cells qRT-PCR results showed in bar graph revealed downregulation in MIO-M1 cells of periostin (A) and VEGFA (B) at mRNA expression levels after periostin knockdown. Results were normalized to human GAPDH mRNA expression levels correspondingly, and data were represented the mean±SEM ( n =4). b P

    Techniques Used: Expressing, Quantitative RT-PCR

    The effect of IFN-γ and TNF-α stimulation to the mRNA levels of VEGFA in MIO-M1 cells Fold change showed in bar plots in the expression for VEGFA in Müller cells after stimulation by IFN-γ and TNF-α by different concentration, 2, 20, 200 ng/mL for IFN-γ (A) and 0.1, 1, 10 ng/mL for TNF-α (B) respectively after 8h and 24h stimulation. GAPDH was used as normalization in all qRT-PCR results. Data were represented the mean±SEM ( n =4). b P
    Figure Legend Snippet: The effect of IFN-γ and TNF-α stimulation to the mRNA levels of VEGFA in MIO-M1 cells Fold change showed in bar plots in the expression for VEGFA in Müller cells after stimulation by IFN-γ and TNF-α by different concentration, 2, 20, 200 ng/mL for IFN-γ (A) and 0.1, 1, 10 ng/mL for TNF-α (B) respectively after 8h and 24h stimulation. GAPDH was used as normalization in all qRT-PCR results. Data were represented the mean±SEM ( n =4). b P

    Techniques Used: Expressing, Concentration Assay, Quantitative RT-PCR

    mRNA expression of VEGFA had no relevance with deficiency of periostin (periostin KO mice) in OIR retinas qRT-PCR results showed in bar plots for mRNA expression levels of periostin was remarkably downregulated in KO mice compared to WT control mice (A). No change was detected for VEGFA expression both in periostin KO mice and WT control mice (B). Results were normalized to mouse GAPDH mRNA expression levels correspondingly, and data were represented the mean±SEM ( n =4). b P
    Figure Legend Snippet: mRNA expression of VEGFA had no relevance with deficiency of periostin (periostin KO mice) in OIR retinas qRT-PCR results showed in bar plots for mRNA expression levels of periostin was remarkably downregulated in KO mice compared to WT control mice (A). No change was detected for VEGFA expression both in periostin KO mice and WT control mice (B). Results were normalized to mouse GAPDH mRNA expression levels correspondingly, and data were represented the mean±SEM ( n =4). b P

    Techniques Used: Expressing, Mouse Assay, Quantitative RT-PCR

    14) Product Images from "Integrative Analysis of DNA Methylation and Gene Expression Data Identifies EPAS1 as a Key Regulator of COPD"

    Article Title: Integrative Analysis of DNA Methylation and Gene Expression Data Identifies EPAS1 as a Key Regulator of COPD

    Journal: PLoS Genetics

    doi: 10.1371/journal.pgen.1004898

    Gene expression levels of Epas1 and Vegfa were lower in chronic smoking mice than non-smoking age-matched mice at the time when COPD develops in different mouse models. A ) Gene expression levels of Epas1 and Vegfa in C57BL/6J mice that develop COPD after 6 months chronic exposure to cigarette smoke. B ) Gene expression levels of Epas1 and Vegfa in A/J mice that develop COPD after 2 months chronic exposure to cigarette smoke. The t-test was used to compare Epas1 or Vegfa expression levels in mice with or without chronic smoke exposure.
    Figure Legend Snippet: Gene expression levels of Epas1 and Vegfa were lower in chronic smoking mice than non-smoking age-matched mice at the time when COPD develops in different mouse models. A ) Gene expression levels of Epas1 and Vegfa in C57BL/6J mice that develop COPD after 6 months chronic exposure to cigarette smoke. B ) Gene expression levels of Epas1 and Vegfa in A/J mice that develop COPD after 2 months chronic exposure to cigarette smoke. The t-test was used to compare Epas1 or Vegfa expression levels in mice with or without chronic smoke exposure.

    Techniques Used: Expressing, Mouse Assay

    15) Product Images from "Analysis of the Role of Igf2 in Adrenal Tumour Development in Transgenic Mouse Models"

    Article Title: Analysis of the Role of Igf2 in Adrenal Tumour Development in Transgenic Mouse Models

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0044171

    Analysis of the cooperation between β-catenin activation and Igf2 overexpression in 10 month-old mice. A- Igf2 overexpression in 10 month-old ΔCat mice does not significantly alter the adrenal phenotype. AdIgf2 mice were mated with ΔCat mice to generate ΔCat;AdIgf2 compound animals. The adrenal phenotype in 10 month-old mice was analysed by haematoxylin eosin staining (a–c) and immunohistochemistry for Dab2. Black arrows show infiltrating subcapsular mesenchymal cells. B- Analysis of Ki67 expression in 10 month-old ΔCat;AdIgf2 adrenals. Ki67 expression was analysed by immunohistochemistry in wild-type (B, a), ΔCat (B, b) and ΔCat;Igf2 (B, c) adrenals. C- Igf2 overexpression in ΔCat mice does not significantly increase proliferation. a-Counting of Ki67-positive cells. Numbers of Ki67-positive cells in a whole adrenal section were counted separately in the cortex (Co) and medulla (M). For each zone, the number of cells was corrected for surface and is expressed as a percentage of Ki67-positive cells in one of the control individuals. Bars represent the mean of at least 7 individual counts in wild-type, ΔCat and ΔCat;AdIgf2 adrenals ± standard deviation. P -value was calculated using Student's t -test. b- Analysis of Cyclin D1 expression. Cyclin D1 expression was analysed by RTqPCR with cDNAs from wild-type, ΔCat and ΔCat;AdIgf2 adrenals. Bars represent the mean relative quantification (Rq Tg/WT) of gene expression in at least 7 adrenals per genotype ± standard deviation. P -value was calculated using Student's t -test. D- Analysis of malignancy markers expression. Expression of VegfA, Connexina43 (cnx43) and Nov/Ccn3 was analysed by RTqPCR with cDNAs from wild-type (WT), ΔCat and ΔCat;AdIgf2 adrenals. Bars represent the mean relative quantification (Rq Tg/WT) of gene expression for each marker in at least 7 adrenals per genotype ± standard deviation. P -value was calculated using Student's t -test. M, medulla; F, fasciculata; G, glomerulosa. Scale bar is 80 µm.
    Figure Legend Snippet: Analysis of the cooperation between β-catenin activation and Igf2 overexpression in 10 month-old mice. A- Igf2 overexpression in 10 month-old ΔCat mice does not significantly alter the adrenal phenotype. AdIgf2 mice were mated with ΔCat mice to generate ΔCat;AdIgf2 compound animals. The adrenal phenotype in 10 month-old mice was analysed by haematoxylin eosin staining (a–c) and immunohistochemistry for Dab2. Black arrows show infiltrating subcapsular mesenchymal cells. B- Analysis of Ki67 expression in 10 month-old ΔCat;AdIgf2 adrenals. Ki67 expression was analysed by immunohistochemistry in wild-type (B, a), ΔCat (B, b) and ΔCat;Igf2 (B, c) adrenals. C- Igf2 overexpression in ΔCat mice does not significantly increase proliferation. a-Counting of Ki67-positive cells. Numbers of Ki67-positive cells in a whole adrenal section were counted separately in the cortex (Co) and medulla (M). For each zone, the number of cells was corrected for surface and is expressed as a percentage of Ki67-positive cells in one of the control individuals. Bars represent the mean of at least 7 individual counts in wild-type, ΔCat and ΔCat;AdIgf2 adrenals ± standard deviation. P -value was calculated using Student's t -test. b- Analysis of Cyclin D1 expression. Cyclin D1 expression was analysed by RTqPCR with cDNAs from wild-type, ΔCat and ΔCat;AdIgf2 adrenals. Bars represent the mean relative quantification (Rq Tg/WT) of gene expression in at least 7 adrenals per genotype ± standard deviation. P -value was calculated using Student's t -test. D- Analysis of malignancy markers expression. Expression of VegfA, Connexina43 (cnx43) and Nov/Ccn3 was analysed by RTqPCR with cDNAs from wild-type (WT), ΔCat and ΔCat;AdIgf2 adrenals. Bars represent the mean relative quantification (Rq Tg/WT) of gene expression for each marker in at least 7 adrenals per genotype ± standard deviation. P -value was calculated using Student's t -test. M, medulla; F, fasciculata; G, glomerulosa. Scale bar is 80 µm.

    Techniques Used: Activation Assay, Over Expression, Mouse Assay, Staining, Immunohistochemistry, Expressing, Standard Deviation, Marker

    16) Product Images from "Interleukin-12 inhibits pathological neovascularization in mouse model of oxygen-induced retinopathy"

    Article Title: Interleukin-12 inhibits pathological neovascularization in mouse model of oxygen-induced retinopathy

    Journal: Scientific Reports

    doi: 10.1038/srep28140

    Effect of intravitreal IL-12 on the expressions of cytokines and the infiltration of CD4+ and CD8+ T cells in OIR retinas. Expressions of IFN-γ ( A ), IP-10 ( B ), and MIG ( C ) were significantly increased in the eyes that were injected with rIL-12. However, the expressions of VEGFA ( D ) and FGF2 ( E ) are not significantly changed in the rIL-12 injected group. Double staining for CD31 (green) and CD4 or CD8 (red) in cryostat sections of OIR eyes that were injected with PBS or rIL-12. Some CD4+ ( F ) and CD8+ cells ( G ) infiltrated in the retina (arrowheads) of the rIL-12 injected group. ** P
    Figure Legend Snippet: Effect of intravitreal IL-12 on the expressions of cytokines and the infiltration of CD4+ and CD8+ T cells in OIR retinas. Expressions of IFN-γ ( A ), IP-10 ( B ), and MIG ( C ) were significantly increased in the eyes that were injected with rIL-12. However, the expressions of VEGFA ( D ) and FGF2 ( E ) are not significantly changed in the rIL-12 injected group. Double staining for CD31 (green) and CD4 or CD8 (red) in cryostat sections of OIR eyes that were injected with PBS or rIL-12. Some CD4+ ( F ) and CD8+ cells ( G ) infiltrated in the retina (arrowheads) of the rIL-12 injected group. ** P

    Techniques Used: Injection, Double Staining

    IL-12 has no direct effect on the growth of human microvascular retinal epithelial cells (HRECs) in vitro . Photographs of tube formation were taken after cultured alone or stimulated by rIL-12 ( A ). The lengths of the tubes were quantitative assessed in each group. The total lengths were not significantly different among the groups ( B , n = 12/group). The mRNA expressions of VEGFA ( C ) and FGF2 ( D ) were not changed after exposure to rIL-12 (n = 4/group). P > 0.05 compared to the controls without rIL-12.
    Figure Legend Snippet: IL-12 has no direct effect on the growth of human microvascular retinal epithelial cells (HRECs) in vitro . Photographs of tube formation were taken after cultured alone or stimulated by rIL-12 ( A ). The lengths of the tubes were quantitative assessed in each group. The total lengths were not significantly different among the groups ( B , n = 12/group). The mRNA expressions of VEGFA ( C ) and FGF2 ( D ) were not changed after exposure to rIL-12 (n = 4/group). P > 0.05 compared to the controls without rIL-12.

    Techniques Used: In Vitro, Cell Culture

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    Quantitative RT-PCR:

    Article Title: DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression
    Article Snippet: In the 3 remaining experiments, sorted cells were lysed and RNA was isolated using a PicoPure isolation kit (Molecular Devices; Toronto, ON) following manufacturer’s instructions. .. RNA was reverse transcribed and amplified using the Ovation Pico WTA System (NuGEN,San Carlos, CA) to PCR template cDNA. qRT-PCR used the ABI Prism 7500 system with TaqMan® Gene Expression Master Mix (Applied Biosystems, Foster City, CA) TaqMan® Gene Expression Assays containing primers and TaqMan probes were purchased from ABI as follow; Mm01168134_m1( Ifng ), Mm00444241_m1( Il22 ), (Mm01192943_m1)( Il22ra1 ), Mm00435613_m1( Pgf ), Mm01281449_m1( Vegfa ), Mm02342889_g1( Ren1 ), Mm00802048_m1( Ace ), Mm00487638_m1( Cma1 ), Mm0059962_m1( Agt ), Mm00616371_m1( Agtr1a ), Mm01341373_m1( Agtr2 ), Mm01255747_g1( Nppa ), and Mm01545399_m1(hypoxanthine guanine phosphoribosyl transferase; Hprt ). .. PCR conditions were initial incubation (2 min; 50°C), enzyme heat activation (10 min; 95°C), 40 cycles (15s at 95°C; 1 min at 60°).

    Expressing:

    Article Title: DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression
    Article Snippet: In the 3 remaining experiments, sorted cells were lysed and RNA was isolated using a PicoPure isolation kit (Molecular Devices; Toronto, ON) following manufacturer’s instructions. .. RNA was reverse transcribed and amplified using the Ovation Pico WTA System (NuGEN,San Carlos, CA) to PCR template cDNA. qRT-PCR used the ABI Prism 7500 system with TaqMan® Gene Expression Master Mix (Applied Biosystems, Foster City, CA) TaqMan® Gene Expression Assays containing primers and TaqMan probes were purchased from ABI as follow; Mm01168134_m1( Ifng ), Mm00444241_m1( Il22 ), (Mm01192943_m1)( Il22ra1 ), Mm00435613_m1( Pgf ), Mm01281449_m1( Vegfa ), Mm02342889_g1( Ren1 ), Mm00802048_m1( Ace ), Mm00487638_m1( Cma1 ), Mm0059962_m1( Agt ), Mm00616371_m1( Agtr1a ), Mm01341373_m1( Agtr2 ), Mm01255747_g1( Nppa ), and Mm01545399_m1(hypoxanthine guanine phosphoribosyl transferase; Hprt ). .. PCR conditions were initial incubation (2 min; 50°C), enzyme heat activation (10 min; 95°C), 40 cycles (15s at 95°C; 1 min at 60°).

    Article Title: VEGFD Protects Retinal Ganglion Cells and, consequently, Capillaries against Excitotoxic Injury
    Article Snippet: Extracted RNA was reverse transcribed into first-strand cDNA using a high-capacity cDNA reverse transcription kit (Applied Biosystems, Foster City, CA, USA). qRT-PCR was performed on a StepOne Plus real-time PCR system using TaqMan gene expression assays for the indicated genes (Applied Biosystems). .. The following TaqMan gene expression assays were used in this study: Thy1 (Mm01174153_m1), nefl (Mm01315666_m1), ocld (Mm01282968_m1), cld5 (Mm00727012_m1), tjp1 (Mm00493699_s1), vegfd (Mm00438963_m1), vegfc (Mm01202432_m1), vegf (Mm01281449_m1), flt4 (Mm01292618_m1), flt1 (Mm00438980_m1), and kdr (Mm00440099_m1). .. Expression of target genes was normalized against the expression of Gapdh (Mm99999915_g1) or, in the case of bEnd.3 cells, Gusb (Mm00446953_m1), which were used as endogenous control genes.

    Article Title: HIF1α is Necessary for Exercise-Induced Neuroprotection while HIF2α is Needed for Dopaminergic Neuron Survival in the Substantia Nigra pars compacta
    Article Snippet: mRNA was isolated from individual SN using a MELT™ kit (Invitrogen, Life Technologies, Carlsbad, CA), following manufacturers suggested protocol. .. Taqman Gene Expression assays (Life Technologies, Carlsbad, CA ) following manufacturers suggested protocol were used for Hif1a (#Mm00468875_m1), Epas1 (#Mm01236112_m1), VegfA (#Mm01281449_m1), Egln1 (#Mm00459770_m1), and RPS18S (#Mm02601778_g1) (n=5–8 for each exercise condition, n=18 for SH). .. Samples were processed on an Eppendorf RealPlex2 Mastercycler and analyzed using included Realplex software (Fisher Scientific, Pittsburgh, PA).

    Binding Assay:

    Article Title: Different cyclical intermittent hypoxia severities have different effects on hippocampal microvasculature
    Article Snippet: Gene expression was measured in an ABI Prism 7900HT Real Time PCR system using TaqMan FAM-labeled specific probes (Applied Biosystems). .. The following TaqMan probes were used: for housekeeping genes to normalize data, TATA box binding protein ( TBP ; Mm00446971_m1) and hypoxanthine-guanine phosphoribosyl transferase ( HPRT ; Mm01545399_m1); for genes of ligands involved in the angiogenesis process, vascular endothelial growth factor A ( VEGFA ; Mm01281449_m1), angiopoietin-1 ( ANGPT1 ; Mm00456503_m1), angiopoietin-2 ( ANGPT2 ; Mm00545822_m1), brain-derived neurotrophic factor ( BDNF ; Mm04230607_s1), fibroblast growth factor 2 ( FGF-2 ; Mm00433287_m1), and delta-like ligand 4 ( DLL4 ; Mm00444619_m1), a negative regulator of angiogenesis; for genes of receptors of the above ligands, FMS-like tyrosine kinase 1 ( FLT-1 , alias VEGFR1 ; Mm00438980_m1), kinase insert domain protein receptor ( KDR , alias VEGFR2 ; Mm01222421_m1), tyrosine kinase with immunoglobulin-like and EGF-like domains 1 ( TIE-1 ; Mm00441786_m1), which acts as a cell-surface receptor for ANGPT1, endothelial-specific receptor tyrosine kinase ( TEK , alias TIE-2 ; Mm00443243_m1), which acts as a cell-surface receptor for ANGPT2, neurotrophic tyrosine kinase receptor type 2 ( NTRK2 , alias TRK2 ; Mm00435422_m1), which acts as a cell-surface receptor for BDNF, FGF receptor 1 ( FGFR-1 ; Mm00438930_m1), FGF receptor 2 ( FGFR-2 ; Mm01269930_m1), and Notch1 ( NOTCH1 ; Mm00435249_m1), which acts as a cell-surface receptor for DLL4; for genes of transcription factors, hypoxia inducible factor 1, α-subunit ( HIF-1α ; Mm00468869_m1), peroxisome proliferative activated receptor-γ coactivator 1α ( PPARGC-1α , alias PGC-1α ; Mm00447181_m1), peroxisome proliferative activated receptor-γ coactivator 1β ( PPARGC-1β , alias PGC-1β ; Mm00504720_m1); for genes of glucose metabolism, solute carrier family 2 (facilitated glucose transporter) member 1 ( SLC2A1 , alias GLUT1 ; Mm00441480_m1), mannoside acetylglucosaminyltransferase 5 ( MGAT5 ; Mm00455036_m1), insulin-like growth factor I ( IGF-I ; Mm00439560_m1), insulin receptor ( INSR ; Mm01211875_m1). .. Gene expression levels were normalized to the geometric mean of TBP and HPRT ; expression of these housekeeping genes did not change with CIH (data not shown).

    Cell Surface Receptor Assay:

    Article Title: Different cyclical intermittent hypoxia severities have different effects on hippocampal microvasculature
    Article Snippet: Gene expression was measured in an ABI Prism 7900HT Real Time PCR system using TaqMan FAM-labeled specific probes (Applied Biosystems). .. The following TaqMan probes were used: for housekeeping genes to normalize data, TATA box binding protein ( TBP ; Mm00446971_m1) and hypoxanthine-guanine phosphoribosyl transferase ( HPRT ; Mm01545399_m1); for genes of ligands involved in the angiogenesis process, vascular endothelial growth factor A ( VEGFA ; Mm01281449_m1), angiopoietin-1 ( ANGPT1 ; Mm00456503_m1), angiopoietin-2 ( ANGPT2 ; Mm00545822_m1), brain-derived neurotrophic factor ( BDNF ; Mm04230607_s1), fibroblast growth factor 2 ( FGF-2 ; Mm00433287_m1), and delta-like ligand 4 ( DLL4 ; Mm00444619_m1), a negative regulator of angiogenesis; for genes of receptors of the above ligands, FMS-like tyrosine kinase 1 ( FLT-1 , alias VEGFR1 ; Mm00438980_m1), kinase insert domain protein receptor ( KDR , alias VEGFR2 ; Mm01222421_m1), tyrosine kinase with immunoglobulin-like and EGF-like domains 1 ( TIE-1 ; Mm00441786_m1), which acts as a cell-surface receptor for ANGPT1, endothelial-specific receptor tyrosine kinase ( TEK , alias TIE-2 ; Mm00443243_m1), which acts as a cell-surface receptor for ANGPT2, neurotrophic tyrosine kinase receptor type 2 ( NTRK2 , alias TRK2 ; Mm00435422_m1), which acts as a cell-surface receptor for BDNF, FGF receptor 1 ( FGFR-1 ; Mm00438930_m1), FGF receptor 2 ( FGFR-2 ; Mm01269930_m1), and Notch1 ( NOTCH1 ; Mm00435249_m1), which acts as a cell-surface receptor for DLL4; for genes of transcription factors, hypoxia inducible factor 1, α-subunit ( HIF-1α ; Mm00468869_m1), peroxisome proliferative activated receptor-γ coactivator 1α ( PPARGC-1α , alias PGC-1α ; Mm00447181_m1), peroxisome proliferative activated receptor-γ coactivator 1β ( PPARGC-1β , alias PGC-1β ; Mm00504720_m1); for genes of glucose metabolism, solute carrier family 2 (facilitated glucose transporter) member 1 ( SLC2A1 , alias GLUT1 ; Mm00441480_m1), mannoside acetylglucosaminyltransferase 5 ( MGAT5 ; Mm00455036_m1), insulin-like growth factor I ( IGF-I ; Mm00439560_m1), insulin receptor ( INSR ; Mm01211875_m1). .. Gene expression levels were normalized to the geometric mean of TBP and HPRT ; expression of these housekeeping genes did not change with CIH (data not shown).

    Pyrolysis Gas Chromatography:

    Article Title: Different cyclical intermittent hypoxia severities have different effects on hippocampal microvasculature
    Article Snippet: Gene expression was measured in an ABI Prism 7900HT Real Time PCR system using TaqMan FAM-labeled specific probes (Applied Biosystems). .. The following TaqMan probes were used: for housekeeping genes to normalize data, TATA box binding protein ( TBP ; Mm00446971_m1) and hypoxanthine-guanine phosphoribosyl transferase ( HPRT ; Mm01545399_m1); for genes of ligands involved in the angiogenesis process, vascular endothelial growth factor A ( VEGFA ; Mm01281449_m1), angiopoietin-1 ( ANGPT1 ; Mm00456503_m1), angiopoietin-2 ( ANGPT2 ; Mm00545822_m1), brain-derived neurotrophic factor ( BDNF ; Mm04230607_s1), fibroblast growth factor 2 ( FGF-2 ; Mm00433287_m1), and delta-like ligand 4 ( DLL4 ; Mm00444619_m1), a negative regulator of angiogenesis; for genes of receptors of the above ligands, FMS-like tyrosine kinase 1 ( FLT-1 , alias VEGFR1 ; Mm00438980_m1), kinase insert domain protein receptor ( KDR , alias VEGFR2 ; Mm01222421_m1), tyrosine kinase with immunoglobulin-like and EGF-like domains 1 ( TIE-1 ; Mm00441786_m1), which acts as a cell-surface receptor for ANGPT1, endothelial-specific receptor tyrosine kinase ( TEK , alias TIE-2 ; Mm00443243_m1), which acts as a cell-surface receptor for ANGPT2, neurotrophic tyrosine kinase receptor type 2 ( NTRK2 , alias TRK2 ; Mm00435422_m1), which acts as a cell-surface receptor for BDNF, FGF receptor 1 ( FGFR-1 ; Mm00438930_m1), FGF receptor 2 ( FGFR-2 ; Mm01269930_m1), and Notch1 ( NOTCH1 ; Mm00435249_m1), which acts as a cell-surface receptor for DLL4; for genes of transcription factors, hypoxia inducible factor 1, α-subunit ( HIF-1α ; Mm00468869_m1), peroxisome proliferative activated receptor-γ coactivator 1α ( PPARGC-1α , alias PGC-1α ; Mm00447181_m1), peroxisome proliferative activated receptor-γ coactivator 1β ( PPARGC-1β , alias PGC-1β ; Mm00504720_m1); for genes of glucose metabolism, solute carrier family 2 (facilitated glucose transporter) member 1 ( SLC2A1 , alias GLUT1 ; Mm00441480_m1), mannoside acetylglucosaminyltransferase 5 ( MGAT5 ; Mm00455036_m1), insulin-like growth factor I ( IGF-I ; Mm00439560_m1), insulin receptor ( INSR ; Mm01211875_m1). .. Gene expression levels were normalized to the geometric mean of TBP and HPRT ; expression of these housekeeping genes did not change with CIH (data not shown).

    Real-time Polymerase Chain Reaction:

    Article Title: Glutathione adducts induced by ischemia and deletion of glutaredoxin-1 stabilize HIF-1α and improve limb revascularization
    Article Snippet: Then reverse transcription was performed to generate cDNA by using the High-Capacity RNA to cDNA kit (4387406; Thermo Fischer Scientific). .. Quantitative PCR was performed using gene-specific TaqMan primers (Invitrogen): 18S (Mm02601777_g1), Hif1α (Mm00468878_m1), Vegfa (Mm01281449_m1), Pdgfa (Mm01205760_m1), and Pdgfb (Mm00440677_m1), Fgf2(Mm00433207_m1). .. Expression levels were analyzed by comparative Ct (ΔΔCT) with StepOne real-time PCR software (Applied Biosystems).

    Article Title: Antitumor effect of Batf2 through IL-12 p40 up-regulation in tumor-associated macrophages
    Article Snippet: The mRNA expression level of each gene was normalized to the 18S rRNA expression level (18S rRNA; Applied Biosystems). .. TaqMan probes for quantitative PCR were purchased from Life Technologies with the following assay ID numbers: Il12b (Mm00434174_m1), Batf2 (Mm01231591_m1), Il6 (Mm00446190_m1), Ifnb1 (Mm00439552_s1), Tnf (Mm00443258_m1), Nos2 (Mm00440502_m1), Vegfa (Mm01281449_m1), and Tbx21 (Mm00450960_m1). ..

    other:

    Article Title: Hypoxia potentiates the cytotoxic effect of piperlongumine in pheochromocytoma models
    Article Snippet: The probes included Mm00441533_g1, Mm03053917_g1, Mm01247357_m1, Mm01281449_m1, Mm04208233_g1, Mm00441531_m1, Mm02384862_g1.

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    Thermo Fisher gene exp vegfa mm01281449 m1
    NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , <t>vegfa</t> , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p
    Gene Exp Vegfa Mm01281449 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , vegfa , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p

    Journal: Molecular Therapy. Methods & Clinical Development

    Article Title: VEGFD Protects Retinal Ganglion Cells and, consequently, Capillaries against Excitotoxic Injury

    doi: 10.1016/j.omtm.2019.12.009

    Figure Lengend Snippet: NMDA-Triggered Excitotoxicity Reduces VEGFD Expression in RGCs (A) qRT-PCR analysis of vegfd , vegfc , vegfa , flt4 , flt1 , and kdr expression in retinal homogenates after intravitreal injection of NMDA. Unpaired t test. n = 6. vegfd , p = 0.0085; vegfc , p = 0.2052; vegfa , p = 0.8524; flt4 , p = 0.9414; flt1 , p = 0.6074; kdr , p = 0.2016. (B) Representative images of human retinal whole mounts. Retinas were immunolabeled with antibodies against Brn3a (red) and VEGFD (green). Scale bars, 10 μm. Negative controls of human retinas were immunolabeled only with secondary antibodies. (C) Representative bands of VEGFD, VEGFA, and β-actin cDNA products following qRT-PCR analysis of human retinal extracts obtained from two donors (subject 1, subject 2). (D) Quantification of VEGFD protein levels in cells in the ganglion cell layer of retinas of mice injected as indicated. Unpaired t test. n = 3. PBS versus NMDA, p = 0.0192. (E) Representative images of mouse sagittal retina sections at d7 after intravitreal injections of NMDA or PBS. Retinas were immunolabeled for VEGFD (green). Nuclei were labeled with Hoechst (blue). GCL, ganglion cell layer. Scale bar, 20 μm. (F) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in brain ECs (b.END3) 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.1744; vegfc , p = 0.7598; vegfa , p = 0.8998; flt4 , p = 0.4457. (G) qRT-PCR analysis of vegfd , vegfc , vegfa , and flt4 expression in primary astrocytes 24 h after 20 μM NMDA treatment. Unpaired t test. n = 3. vegfd , p = 0.8716; vegfc , p = 0.4918; vegfa , p = 0.9902; flt4 , p = 0.9331. Bars represent mean ± SEM. Dots represent single values. **p

    Article Snippet: The following TaqMan gene expression assays were used in this study: Thy1 (Mm01174153_m1), nefl (Mm01315666_m1), ocld (Mm01282968_m1), cld5 (Mm00727012_m1), tjp1 (Mm00493699_s1), vegfd (Mm00438963_m1), vegfc (Mm01202432_m1), vegf (Mm01281449_m1), flt4 (Mm01292618_m1), flt1 (Mm00438980_m1), and kdr (Mm00440099_m1).

    Techniques: Expressing, Quantitative RT-PCR, Injection, Immunolabeling, Mouse Assay, Labeling

    Piperlongumine activates apoptosis and necroptosis, and inhibits cell migration and invasion A. RIP1-IP was performed on cell lysates from MPC cells treated with the indicated concentrations of PL at 21% and 1% O 2 for 24 hours and probed for RIP1 (top lane) and RIP3 (bottom lane). A representative image (n=3) is shown. B. MTT cells were treated with indicated concentrations of PL at 21% and 1% O 2 for 24 hours. Total cell lysates were analyzed by Western blot for cleaved PARP, cleaved caspase 3 and caspase 3. β-tubulin was used as a loading control. The representative image (n=3) is shown. C. 1.5 × 10 5 MPC cells were plated in the upper part of transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. D. 1.5 × 10 5 MPC cells were plated in the upper part of matrigel-coated transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. E. MPC cells were treated with indicated concentrations of PL for 24 hours. mRNA expression levels of Twist1, Vegfa, Mmp9, and Nanog were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . A graph represents data from three independent experiments as mean +/− SEM. *P

    Journal: Oncotarget

    Article Title: Hypoxia potentiates the cytotoxic effect of piperlongumine in pheochromocytoma models

    doi: 10.18632/oncotarget.9643

    Figure Lengend Snippet: Piperlongumine activates apoptosis and necroptosis, and inhibits cell migration and invasion A. RIP1-IP was performed on cell lysates from MPC cells treated with the indicated concentrations of PL at 21% and 1% O 2 for 24 hours and probed for RIP1 (top lane) and RIP3 (bottom lane). A representative image (n=3) is shown. B. MTT cells were treated with indicated concentrations of PL at 21% and 1% O 2 for 24 hours. Total cell lysates were analyzed by Western blot for cleaved PARP, cleaved caspase 3 and caspase 3. β-tubulin was used as a loading control. The representative image (n=3) is shown. C. 1.5 × 10 5 MPC cells were plated in the upper part of transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. D. 1.5 × 10 5 MPC cells were plated in the upper part of matrigel-coated transwell chambers and allowed to migrate for 24 hours in the presence of 0, 1, and 5μM PL. The box and whiskers graph represents data from three independent experiments. E. MPC cells were treated with indicated concentrations of PL for 24 hours. mRNA expression levels of Twist1, Vegfa, Mmp9, and Nanog were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . A graph represents data from three independent experiments as mean +/− SEM. *P

    Article Snippet: The probes included Mm00441533_g1, Mm03053917_g1, Mm01247357_m1, Mm01281449_m1, Mm04208233_g1, Mm00441531_m1, Mm02384862_g1.

    Techniques: Migration, MTT Assay, Western Blot, Expressing, Real-time Polymerase Chain Reaction

    Piperlongumine inhibits tumor growth and metastases formation and decreases expression levels of EMT and angiogenesis markers in vivo A. Nude female mice bearing MTT-Luc tumors were treated with 24 mg/kg/day PL (n=9) for 28 days. The control group was treated with vehicle (n=10). Tumor growth was assessed once a week. The graph shows a significant inhibition of tumor growth at all time points of treatment in the treated group (green) compared to control (red) animals. The graph represents data from 9 (treated) and 10 (vehicle) mice assessed at specific time points as mean +/− SEM. Statistical analysis was performed by Mann Whitney, U-test, at each time point. B. Representative tumors from control mice compared to the tumors from animals treated with PL. Scale bar: 1cm. C. Lungs were resected and subjected to bioluminescence imaging. The presence of lung metastases in treated mice was 46% lower than in control group. D. mRNA expression levels of Pou5f1, Twist1, Vegfa, Mmp9, and Nanog in tumors from both treated and control groups were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . The box and whiskers graphs represent data from control (n=10) and treated (n=9) groups. *P

    Journal: Oncotarget

    Article Title: Hypoxia potentiates the cytotoxic effect of piperlongumine in pheochromocytoma models

    doi: 10.18632/oncotarget.9643

    Figure Lengend Snippet: Piperlongumine inhibits tumor growth and metastases formation and decreases expression levels of EMT and angiogenesis markers in vivo A. Nude female mice bearing MTT-Luc tumors were treated with 24 mg/kg/day PL (n=9) for 28 days. The control group was treated with vehicle (n=10). Tumor growth was assessed once a week. The graph shows a significant inhibition of tumor growth at all time points of treatment in the treated group (green) compared to control (red) animals. The graph represents data from 9 (treated) and 10 (vehicle) mice assessed at specific time points as mean +/− SEM. Statistical analysis was performed by Mann Whitney, U-test, at each time point. B. Representative tumors from control mice compared to the tumors from animals treated with PL. Scale bar: 1cm. C. Lungs were resected and subjected to bioluminescence imaging. The presence of lung metastases in treated mice was 46% lower than in control group. D. mRNA expression levels of Pou5f1, Twist1, Vegfa, Mmp9, and Nanog in tumors from both treated and control groups were assessed by quantitative real-time PCR. The target gene transcript levels were normalized to Actb . The box and whiskers graphs represent data from control (n=10) and treated (n=9) groups. *P

    Article Snippet: The probes included Mm00441533_g1, Mm03053917_g1, Mm01247357_m1, Mm01281449_m1, Mm04208233_g1, Mm00441531_m1, Mm02384862_g1.

    Techniques: Expressing, In Vivo, Mouse Assay, MTT Assay, Inhibition, MANN-WHITNEY, Imaging, Real-time Polymerase Chain Reaction

    Epas1 (Hif2α) is increased in the SN with 3 months of wheel running exercise. The expression of hif1α (A), epas1 (B), egln1 (C), and vegfA (D) in the SN from WT mice after 3 months of exercise or SH was measured by qPCR. Epas1 (B) in the SN of exercised mice is significantly elevated (*= p≤0.02). mRNA levels are normalized to 18S ribosomal RNA and compared to expression in SH. Epas1 expression from mice in SH+MPTP and exercise + MPTP showed no changes, neither did hif1α , egln1 , or vegfA expression in any condition.

    Journal: Neuroscience

    Article Title: HIF1α is Necessary for Exercise-Induced Neuroprotection while HIF2α is Needed for Dopaminergic Neuron Survival in the Substantia Nigra pars compacta

    doi: 10.1016/j.neuroscience.2015.03.015

    Figure Lengend Snippet: Epas1 (Hif2α) is increased in the SN with 3 months of wheel running exercise. The expression of hif1α (A), epas1 (B), egln1 (C), and vegfA (D) in the SN from WT mice after 3 months of exercise or SH was measured by qPCR. Epas1 (B) in the SN of exercised mice is significantly elevated (*= p≤0.02). mRNA levels are normalized to 18S ribosomal RNA and compared to expression in SH. Epas1 expression from mice in SH+MPTP and exercise + MPTP showed no changes, neither did hif1α , egln1 , or vegfA expression in any condition.

    Article Snippet: Taqman Gene Expression assays (Life Technologies, Carlsbad, CA ) following manufacturers suggested protocol were used for Hif1a (#Mm00468875_m1), Epas1 (#Mm01236112_m1), VegfA (#Mm01281449_m1), Egln1 (#Mm00459770_m1), and RPS18S (#Mm02601778_g1) (n=5–8 for each exercise condition, n=18 for SH).

    Techniques: Expressing, Mouse Assay, Real-time Polymerase Chain Reaction

    Expression of hypoxia sensitive molecules in the SN is modulated by wheel running exercise. hif1a (A), epas1 (B), egln1 (C), and vegfA (D) expression from SN mRNA isolated at 1, 3, 5, 7, and 14 days of running show significant reduction at 3 days (A,B,D) and increase at 5 days (A,C) of running compared to controls in SH. *=p≤0.05, #=p≤0.01. N=17–18 for SH, 5–8 for each exercise condition. HIF1α protein from the SN (E, F) is significantly increased at running day 3 and after 21 days of running plus MPTP administration (SN taken at 2 hrs. after the last MPTP injection). *=p≤0.05. SN HIF2α protein (G, H) shows no statistically significant change with running. Error bars =± SEM. Arrows indicate 114kD, expected bands for these HIF1α and HIF2α antibodies are ~120kD. Lysate from COS-7 cells in normoxic and hypoxic conditions are used as negative and positive controls (F, H), and HIF2α human recombinant protein (H) as a positive control (~120kD). Values are expressed as percent of control relative to SH after normalization to β-actin (F, H, ~47kD) for individual samples. N= 4 SN for each condition.

    Journal: Neuroscience

    Article Title: HIF1α is Necessary for Exercise-Induced Neuroprotection while HIF2α is Needed for Dopaminergic Neuron Survival in the Substantia Nigra pars compacta

    doi: 10.1016/j.neuroscience.2015.03.015

    Figure Lengend Snippet: Expression of hypoxia sensitive molecules in the SN is modulated by wheel running exercise. hif1a (A), epas1 (B), egln1 (C), and vegfA (D) expression from SN mRNA isolated at 1, 3, 5, 7, and 14 days of running show significant reduction at 3 days (A,B,D) and increase at 5 days (A,C) of running compared to controls in SH. *=p≤0.05, #=p≤0.01. N=17–18 for SH, 5–8 for each exercise condition. HIF1α protein from the SN (E, F) is significantly increased at running day 3 and after 21 days of running plus MPTP administration (SN taken at 2 hrs. after the last MPTP injection). *=p≤0.05. SN HIF2α protein (G, H) shows no statistically significant change with running. Error bars =± SEM. Arrows indicate 114kD, expected bands for these HIF1α and HIF2α antibodies are ~120kD. Lysate from COS-7 cells in normoxic and hypoxic conditions are used as negative and positive controls (F, H), and HIF2α human recombinant protein (H) as a positive control (~120kD). Values are expressed as percent of control relative to SH after normalization to β-actin (F, H, ~47kD) for individual samples. N= 4 SN for each condition.

    Article Snippet: Taqman Gene Expression assays (Life Technologies, Carlsbad, CA ) following manufacturers suggested protocol were used for Hif1a (#Mm00468875_m1), Epas1 (#Mm01236112_m1), VegfA (#Mm01281449_m1), Egln1 (#Mm00459770_m1), and RPS18S (#Mm02601778_g1) (n=5–8 for each exercise condition, n=18 for SH).

    Techniques: Expressing, Isolation, Injection, Recombinant, Positive Control

    Pathological LV hypertrophy and perivascular fibrosis post-SAC. Pathological hypertrophy marker mRNAs were evaluated one week post-sham or -SAC surgery in adult male C57BL/6J mice (9-week-old) from ( a ) LV tissue (sham, n = 7; SAC, n = 6) or ( b ) enriched cardiomyocyte fractions (sham, n = 7; SAC, n = 8). ( c ) Representative transverse LV sections demonstrate the development of perivascular fibrosis around the coronaries (indicated by black arrows) after SAC-surgery (right) compared to sham-surgery (left). Sections were stained with Fast Green and Picrosirius Red to identify the cytoplasm and collagen, respectively. ( d ) Hif1a and Vegfa expression in cardiomyocytes one week post-SAC (sham, n = 7; SAC, n = 8). Data are presented as means ± SD, independent comparisons were made by two-tailed Student’s unpaired t-tests; *P

    Journal: Scientific Reports

    Article Title: Pressure overload by suprarenal aortic constriction in mice leads to left ventricular hypertrophy without c-Kit expression in cardiomyocytes

    doi: 10.1038/s41598-020-72273-3

    Figure Lengend Snippet: Pathological LV hypertrophy and perivascular fibrosis post-SAC. Pathological hypertrophy marker mRNAs were evaluated one week post-sham or -SAC surgery in adult male C57BL/6J mice (9-week-old) from ( a ) LV tissue (sham, n = 7; SAC, n = 6) or ( b ) enriched cardiomyocyte fractions (sham, n = 7; SAC, n = 8). ( c ) Representative transverse LV sections demonstrate the development of perivascular fibrosis around the coronaries (indicated by black arrows) after SAC-surgery (right) compared to sham-surgery (left). Sections were stained with Fast Green and Picrosirius Red to identify the cytoplasm and collagen, respectively. ( d ) Hif1a and Vegfa expression in cardiomyocytes one week post-SAC (sham, n = 7; SAC, n = 8). Data are presented as means ± SD, independent comparisons were made by two-tailed Student’s unpaired t-tests; *P

    Article Snippet: The following TaqMan probes (Thermo Fisher) were used: Nppa (Mm01255747_g1), Nppb (Mm01255770_g1), Myh7 (Mm00600555_m1), Acta1 (Mm00808218_g1), Hprt (Mm01545399_m1), Hif1a (Mm00468869_m1), Vegfa (Mm01281449_m1), Kit (Mm00445212_m1).

    Techniques: Marker, Mouse Assay, Staining, Expressing, Two Tailed Test