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Santa Cruz Biotechnology ribonucleic acid sirna
Alteration of phosphorylation on AKT came with <t>MTDH</t> expression (A and B); extra-/intracellular VEGF expression was reduced according to different concentration of AKT <t>siRNA</t> treatment (C and D); VEGF expression was attenuated in extra-/intracellular after PI3K/Akt inhibitor LY294002 stimulation (E and F); ● Control siRNA had added for vector control. ∗ P
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Images

1) Product Images from "Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck"

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck

Journal: Medicine

doi: 10.1097/MD.0000000000000502

Alteration of phosphorylation on AKT came with MTDH expression (A and B); extra-/intracellular VEGF expression was reduced according to different concentration of AKT siRNA treatment (C and D); VEGF expression was attenuated in extra-/intracellular after PI3K/Akt inhibitor LY294002 stimulation (E and F); ● Control siRNA had added for vector control. ∗ P
Figure Legend Snippet: Alteration of phosphorylation on AKT came with MTDH expression (A and B); extra-/intracellular VEGF expression was reduced according to different concentration of AKT siRNA treatment (C and D); VEGF expression was attenuated in extra-/intracellular after PI3K/Akt inhibitor LY294002 stimulation (E and F); ● Control siRNA had added for vector control. ∗ P

Techniques Used: Expressing, Concentration Assay, Plasmid Preparation

Increasing expression of VEGF in overexpressed MTDH (MTDH cDNA) cells was detected comparing with parental cell (Blank) and vector control (pcDNA) by western blot (A) and ELISA (C and E); reduced VEGF expression was found in MTDH knockdown (shRNA MTDH ) cells compared with parental (Blank) and control cells (shRNA control ) by western blot (B) and ELISA (D and F). ∗ P
Figure Legend Snippet: Increasing expression of VEGF in overexpressed MTDH (MTDH cDNA) cells was detected comparing with parental cell (Blank) and vector control (pcDNA) by western blot (A) and ELISA (C and E); reduced VEGF expression was found in MTDH knockdown (shRNA MTDH ) cells compared with parental (Blank) and control cells (shRNA control ) by western blot (B) and ELISA (D and F). ∗ P

Techniques Used: Expressing, Plasmid Preparation, Western Blot, Enzyme-linked Immunosorbent Assay, shRNA

2) Product Images from "Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated by Rab5"

Article Title: Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated by Rab5

Journal: Shock (Augusta, Ga.)

doi: 10.1097/SHK.0000000000000995

Knockdown of Rab5 inhibits endocytosis of RBC-derived MPs. (A) Cultured endothelial cells were treated with nonsense siRNA (NS) as a negative control, or three separate Rab5 siRNA oligonucleotide sequences (13.3, 13.2, 13.1). After incubation, Rab5 protein was determined by Western blot analysis. Note: both lanes were different exposures of the same gel, as β-actin expression required less exposure as compared with Rab5 . (B) Quantitation of Rab5 to B-actin protein expression. (C) Cultured endothelial cells were transfected with nonsense (NS) or Rab5 siRNA 13.1, then treated with vehicle or CFSE-labelled MPs and analyzed via flow cytometry for the presence of CFSE. (D) Experiments were repeated with trypan blue. (E) Quantification of MFI from data presented in C and D. *p
Figure Legend Snippet: Knockdown of Rab5 inhibits endocytosis of RBC-derived MPs. (A) Cultured endothelial cells were treated with nonsense siRNA (NS) as a negative control, or three separate Rab5 siRNA oligonucleotide sequences (13.3, 13.2, 13.1). After incubation, Rab5 protein was determined by Western blot analysis. Note: both lanes were different exposures of the same gel, as β-actin expression required less exposure as compared with Rab5 . (B) Quantitation of Rab5 to B-actin protein expression. (C) Cultured endothelial cells were transfected with nonsense (NS) or Rab5 siRNA 13.1, then treated with vehicle or CFSE-labelled MPs and analyzed via flow cytometry for the presence of CFSE. (D) Experiments were repeated with trypan blue. (E) Quantification of MFI from data presented in C and D. *p

Techniques Used: Derivative Assay, Cell Culture, Negative Control, Incubation, Western Blot, Expressing, Quantitation Assay, Transfection, Flow Cytometry, Cytometry

Knockdown of Rab5 in endothelial cells attenuates the endothelial activation response after treatment with RBC-derived MPs. Nonsense (NS) siRNA was used as a control for all experiments. (A) Cells treated with NS siRNA, then MPs demonstrate increased ICAM and E-selectin expression. This effect was attenuated in Rab5 -knockdown cells. (B) Quantification of adhesion molecule upregulation data. (C) NS cells treated with MPs also demonstrated increased concentrations of IL-6 as compared to Rab5 -knockdown cells. This effect was inhibited in cells treated with Rab5 siRNA, then MPs. *p
Figure Legend Snippet: Knockdown of Rab5 in endothelial cells attenuates the endothelial activation response after treatment with RBC-derived MPs. Nonsense (NS) siRNA was used as a control for all experiments. (A) Cells treated with NS siRNA, then MPs demonstrate increased ICAM and E-selectin expression. This effect was attenuated in Rab5 -knockdown cells. (B) Quantification of adhesion molecule upregulation data. (C) NS cells treated with MPs also demonstrated increased concentrations of IL-6 as compared to Rab5 -knockdown cells. This effect was inhibited in cells treated with Rab5 siRNA, then MPs. *p

Techniques Used: Activation Assay, Derivative Assay, Expressing

3) Product Images from "Adenosine A1 Receptors Promote Vasa Vasorum Endothelial Cell Barrier Integrity via Gi and Akt-Dependent Actin Cytoskeleton Remodeling"

Article Title: Adenosine A1 Receptors Promote Vasa Vasorum Endothelial Cell Barrier Integrity via Gi and Akt-Dependent Actin Cytoskeleton Remodeling

Journal: PLoS ONE

doi: 10.1371/journal.pone.0059733

A1R is involved in adenosine-induced VVEC barrier function. Effect of A1R siRNA on CCPA-induced increase in TER in VVEC. ( A, B ) VVEC were incubated with A1R specific siRNA or non-specific siRNA for 48 h and then cells were stimulated with CCPA (1 nM) in TER measurement assay. The depletion of A1R mRNA and protein was confirmed by RT-PCR ( C ) and the Western blot analysis with anti-A1R antibody. ( D ). Results are presented as mean ± SE from three independent experiments.
Figure Legend Snippet: A1R is involved in adenosine-induced VVEC barrier function. Effect of A1R siRNA on CCPA-induced increase in TER in VVEC. ( A, B ) VVEC were incubated with A1R specific siRNA or non-specific siRNA for 48 h and then cells were stimulated with CCPA (1 nM) in TER measurement assay. The depletion of A1R mRNA and protein was confirmed by RT-PCR ( C ) and the Western blot analysis with anti-A1R antibody. ( D ). Results are presented as mean ± SE from three independent experiments.

Techniques Used: Incubation, Reverse Transcription Polymerase Chain Reaction, Western Blot

4) Product Images from "Noxa/Mcl-1 Balance Regulates Susceptibility of Cells to Camptothecin-Induced Apoptosis 1"

Article Title: Noxa/Mcl-1 Balance Regulates Susceptibility of Cells to Camptothecin-Induced Apoptosis 1

Journal: Neoplasia (New York, N.Y.)

doi:

Noxa alone cannot account for the CPT-induced apoptosis. (A) Knockdown of Noxa expression protects cells against CPT-induced apoptosis. HeLa cells were transfected with control or Noxa-specific siRNA (sc-37305; Santa Cruz Biotechnology). At 48 hours posttransfection, cells were treated with CPT (10 µM) for another 12 or 24 hours. The viability of cells was measured by counting apoptotic cells characteristic of aberrant nuclei staining by Hoechst 33342 and was normalized to the number of total cells. Data were presented as means (± SD) of three independent experiments. Cell lysates were also collected and subjected to Western blot analysis using anti-Noxa and anti-caspase-9 antibodies. Endogenous actin was detected by anti-actin antibody as internal control. (B) HeLa cells were treated with CPT (10 µM) alone or in combination with either (a) CHX (20 µg/ml) or (b) LY294002 (40 µM) for indicated periods of time. The viability of cells was measured according to the method described in (A). Noxa protein level and the activation of caspase-9 were assessed by Western blot analysis. (C) CPT-induced Mcl-1 upregulation is inhibited by LY294002. a. HeLa cells were treated with CPT (10 µM) or doxorubicin (2 µg/ml) for indicated periods of time (0, 1, 3, 6, 9, 12, and 24 hours) before cell lysates were prepared. Levels of Mcl-1 protein were assessed by Western blot analysis using anti -Mcl-1 antibody. Actin acted as a loading control. b. HeLa cells were treated with CPT (10 µM) or with CPT (10 µM) plus LY294002 (40 µM) for 12 hours. Cell lysates were then used for Western blot analysis with anti-Mcl-1 antibody and endogenous actin was used as internal control.
Figure Legend Snippet: Noxa alone cannot account for the CPT-induced apoptosis. (A) Knockdown of Noxa expression protects cells against CPT-induced apoptosis. HeLa cells were transfected with control or Noxa-specific siRNA (sc-37305; Santa Cruz Biotechnology). At 48 hours posttransfection, cells were treated with CPT (10 µM) for another 12 or 24 hours. The viability of cells was measured by counting apoptotic cells characteristic of aberrant nuclei staining by Hoechst 33342 and was normalized to the number of total cells. Data were presented as means (± SD) of three independent experiments. Cell lysates were also collected and subjected to Western blot analysis using anti-Noxa and anti-caspase-9 antibodies. Endogenous actin was detected by anti-actin antibody as internal control. (B) HeLa cells were treated with CPT (10 µM) alone or in combination with either (a) CHX (20 µg/ml) or (b) LY294002 (40 µM) for indicated periods of time. The viability of cells was measured according to the method described in (A). Noxa protein level and the activation of caspase-9 were assessed by Western blot analysis. (C) CPT-induced Mcl-1 upregulation is inhibited by LY294002. a. HeLa cells were treated with CPT (10 µM) or doxorubicin (2 µg/ml) for indicated periods of time (0, 1, 3, 6, 9, 12, and 24 hours) before cell lysates were prepared. Levels of Mcl-1 protein were assessed by Western blot analysis using anti -Mcl-1 antibody. Actin acted as a loading control. b. HeLa cells were treated with CPT (10 µM) or with CPT (10 µM) plus LY294002 (40 µM) for 12 hours. Cell lysates were then used for Western blot analysis with anti-Mcl-1 antibody and endogenous actin was used as internal control.

Techniques Used: Cycling Probe Technology, Expressing, Transfection, Staining, Western Blot, Activation Assay

5) Product Images from "Characterization of the human Activin-A receptor type II-like kinase 1 (ACVRL1) promoter and its regulation by Sp1"

Article Title: Characterization of the human Activin-A receptor type II-like kinase 1 (ACVRL1) promoter and its regulation by Sp1

Journal: BMC Molecular Biology

doi: 10.1186/1471-2199-11-51

Effect of Sp1 expression on ACVRL1 promoter activity . (A) Dose-response effect of Sp1 on the transcriptional activity of ACVRL1 promoter in Schneider S2 and HEK293T cells. S2 (Sp1-less) and HEK293T cells were cotransfected with the pGL2 empty vector or the ACVRL1 promoter construct -1,035/+210 and with increasing amounts of the Sp1 expression vector (pPac-Sp1 and pCIneo-Sp1, respectively). Luciferase activity was corrected with β-galactosidase activity and expressed as fold induction of the transcriptional activity of pALK1 in the absence of exogenous Sp1. (B) Left, scheme showing the distribution of the different Sp1 consensus binding sites along the ACVRL1 promoter (black ovals) in the different constructs. Right, transient transfection of Schneider S2 cells with 25 ng of pPac-Sp1 and the indicated ACVRL1 promoter constructs. Fold-induction values respect to basal activity are indicated on top of each bar. (C) Effect of Sp1-knock down on ACVRL1 transcriptional activity. HEK293T cells were transfected with Sp1 siRNA. Left, Sp1 mRNA and protein levels were measured by semiquantitative RT-PCR and western blot after 48 hr. Right, 24 hr after the siRNA Sp1 transfection, the different ACVRL1 promoter constructs were transfected. The transcriptional activity of all the fragments was measured and normalized by the β-galactosidase activity. Basal pALK1 activity (100%) and the reduction after Sp1 silencing (grey bars) are shown. In every case, Sp1 suppression resulted at least in a decrease of 50% in ACVRL1 transcriptional activity (***p
Figure Legend Snippet: Effect of Sp1 expression on ACVRL1 promoter activity . (A) Dose-response effect of Sp1 on the transcriptional activity of ACVRL1 promoter in Schneider S2 and HEK293T cells. S2 (Sp1-less) and HEK293T cells were cotransfected with the pGL2 empty vector or the ACVRL1 promoter construct -1,035/+210 and with increasing amounts of the Sp1 expression vector (pPac-Sp1 and pCIneo-Sp1, respectively). Luciferase activity was corrected with β-galactosidase activity and expressed as fold induction of the transcriptional activity of pALK1 in the absence of exogenous Sp1. (B) Left, scheme showing the distribution of the different Sp1 consensus binding sites along the ACVRL1 promoter (black ovals) in the different constructs. Right, transient transfection of Schneider S2 cells with 25 ng of pPac-Sp1 and the indicated ACVRL1 promoter constructs. Fold-induction values respect to basal activity are indicated on top of each bar. (C) Effect of Sp1-knock down on ACVRL1 transcriptional activity. HEK293T cells were transfected with Sp1 siRNA. Left, Sp1 mRNA and protein levels were measured by semiquantitative RT-PCR and western blot after 48 hr. Right, 24 hr after the siRNA Sp1 transfection, the different ACVRL1 promoter constructs were transfected. The transcriptional activity of all the fragments was measured and normalized by the β-galactosidase activity. Basal pALK1 activity (100%) and the reduction after Sp1 silencing (grey bars) are shown. In every case, Sp1 suppression resulted at least in a decrease of 50% in ACVRL1 transcriptional activity (***p

Techniques Used: Expressing, Activity Assay, Plasmid Preparation, Construct, Luciferase, Binding Assay, Transfection, Reverse Transcription Polymerase Chain Reaction, Western Blot

Related Articles

Transfection:

Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
Article Snippet: Silencing of RAGE Gene Expression and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) Detection for RAGE mRNA and HMGB1 Release RNA silencing was performed with small interfering RNA (siRNA) targeting mouse RAGE mRNA or negative control siRNA (Santa Cruz, catalog: sc-36375). .. Cells were transfected with siRNA duplexes suspended in lipofectamine reagent (Life Technologies) following the manufacturer's protocol.

Article Title: Diabetic Retinopathy and Signaling Mechanism for Activation of Matrix Metalloproteinase-9
Article Snippet: .. BRECs from 3rd to 4th passage were transfected with small interfering RNA (siRNA) of H-Ras, Raf-1, MEK, or MMP9 using transfection reagents and siRNA duplex from Santa Cruz Biotechnology (Santa Cruz, CA). .. The transfection complex was prepared by adding siRNA duplex (0.25–1 μg) and siRNA transfection reagent.

Article Title: Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated by Rab5
Article Snippet: Confluent monolayers of MLECs were treated with small interfering ribonucleic acid (siRNA) specific for Rab5 GTPase for 48–72 hours per manufacturer protocol (Santa Cruz Biotechnology, Dallas, TX). .. As reported by the manufacturer (Integrated DNA Technologies, Coralville, IA), its genetic sequence is as follows: 5′-CGUUAAUCGCGUAUAAUACGCGUATAUACGCGUAUUAUACGCGAUUAACGAC-3′ Transfection was carried out per manufacturer recommendations.

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: .. Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA). ..

Article Title: Programmed Death 1 and Cytokine Inducible SH2-Containing Protein Dependent Expansion of Regulatory T Cells Upon Stimulation With Mycobacterium tuberculosis
Article Snippet: .. CD4+ CD25− cells were transfected with small interfering RNA (siRNA) for CISH or cAMP response element modulator (CREM) or with scrambled siRNA (Santa Cruz Biotechnology). .. Briefly, 106 CD4+ CD25− cells were transfected with 50 μmol/L siRNA in 200 μL of transfection medium.

Article Title: Involvement of Toll-Like Receptor 2 and Epidermal Growth Factor Receptor Signaling in Epithelial Expression of Airway Remodeling Factors
Article Snippet: .. The small interfering RNA (siRNA) against TLR2 and the no-effect control were from Santa Cruz Biotechnology (Santa Cruz, CA), and HiPerFect transfection reagent was from Qiagen (Valencia, CA). .. Anti–TGF-α antibody and rabbit isotype control were purchased from Abcam (Cambridge, MA).

Article Title: Notch2 regulates matrix metallopeptidase 9 via PI3K/AKT signaling in human gastric carcinoma cell MKN-45
Article Snippet: .. MKN-45 cells were transfected with small interfering RNA (siRNA) against Notch2 and scrambled siRNA (Santa Cruz Biotechnology, CA, United States) constructs using the commercial transfection reagent (Santa Cruz Biotechnology, CA, United States) according to the manufacturer’s instructions. .. Following transfection, cells were incubated at 37 °C in a CO2 incubator for 48 h before being harvested for the assays described below.

Article Title: Superoxide from NADPH oxidase upregulates type 5 phosphodiesterase in human vascular smooth muscle cells: inhibition with iloprost and NONOate
Article Snippet: Paragraph title: Transfection with small interfering RNAs ... Predesigned small interfering RNA (siRNA) targeting human NOX-1 and NOX-4 and scrambled siRNA (as a negative control) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Article Title: Ginkgetin induces apoptosis in 786-O cell line via suppression of JAK2-STAT3 pathway
Article Snippet: .. Small interfering RNA and transient transfection The small interfering RNA (siRNA) targeting human STAT3 and control siRNA were from Santa Cruz. ..

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: .. CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. Membranes were stripped and re‐probed with anti‐ β ‐actin antibody (Santa Cruz Biotechnology) as an internal control.

Luciferase:

Article Title: Novel 1,3,4-Oxadiazole Induces Anticancer Activity by Targeting NF-κB in Hepatocellular Carcinoma Cells
Article Snippet: The small interfering RNA (siRNA) for NF-κB and scrambled control was obtained from Santa Cruz Biotechnology. .. Caspase-Glo 3/7 assay kit and luciferase substrate was purchased from promega (WI, USA).

IA:

Article Title: Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated by Rab5
Article Snippet: Confluent monolayers of MLECs were treated with small interfering ribonucleic acid (siRNA) specific for Rab5 GTPase for 48–72 hours per manufacturer protocol (Santa Cruz Biotechnology, Dallas, TX). .. Three separate Rab5 -specific oligonucleotide sequences (Integrated DNA Technologies, Coralville, IA) were tested: 13.1 (5′-GUAGAAUCAAGUUUCUAAUUCUGAA-3′), 13.2 (5′-UCAAAGGCAAGCAAGUCCUAAUATT-3′), and 13.3 (5′-AAAUUUGGACAUGGCUAAUCGAGGA-3′).

Stable Transfection:

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: .. Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA). ..

Isolation:

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: .. CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. Membranes were stripped and re‐probed with anti‐ β ‐actin antibody (Santa Cruz Biotechnology) as an internal control.

Negative Control:

Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
Article Snippet: .. Silencing of RAGE Gene Expression and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) Detection for RAGE mRNA and HMGB1 Release RNA silencing was performed with small interfering RNA (siRNA) targeting mouse RAGE mRNA or negative control siRNA (Santa Cruz, catalog: sc-36375). .. Cells were transfected with siRNA duplexes suspended in lipofectamine reagent (Life Technologies) following the manufacturer's protocol.

Article Title: Superoxide from NADPH oxidase upregulates type 5 phosphodiesterase in human vascular smooth muscle cells: inhibition with iloprost and NONOate
Article Snippet: .. Predesigned small interfering RNA (siRNA) targeting human NOX-1 and NOX-4 and scrambled siRNA (as a negative control) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). .. Growth-arrested hVSMCs were trypsinized, pelleted and resuspended in nucleofector solution, provided in the Basic nucleofector kit for primary smooth muscle cells (Amaxa biosystems, Cologne, Germany), at a density of 0.8 × 106 cells (100 μL)−1 .

Western Blot:

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: .. CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. Membranes were stripped and re‐probed with anti‐ β ‐actin antibody (Santa Cruz Biotechnology) as an internal control.

Small Interfering RNA:

Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
Article Snippet: .. Silencing of RAGE Gene Expression and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) Detection for RAGE mRNA and HMGB1 Release RNA silencing was performed with small interfering RNA (siRNA) targeting mouse RAGE mRNA or negative control siRNA (Santa Cruz, catalog: sc-36375). .. Cells were transfected with siRNA duplexes suspended in lipofectamine reagent (Life Technologies) following the manufacturer's protocol.

Article Title: Diabetic Retinopathy and Signaling Mechanism for Activation of Matrix Metalloproteinase-9
Article Snippet: .. BRECs from 3rd to 4th passage were transfected with small interfering RNA (siRNA) of H-Ras, Raf-1, MEK, or MMP9 using transfection reagents and siRNA duplex from Santa Cruz Biotechnology (Santa Cruz, CA). .. The transfection complex was prepared by adding siRNA duplex (0.25–1 μg) and siRNA transfection reagent.

Article Title: Programmed Death 1 and Cytokine Inducible SH2-Containing Protein Dependent Expansion of Regulatory T Cells Upon Stimulation With Mycobacterium tuberculosis
Article Snippet: .. CD4+ CD25− cells were transfected with small interfering RNA (siRNA) for CISH or cAMP response element modulator (CREM) or with scrambled siRNA (Santa Cruz Biotechnology). .. Briefly, 106 CD4+ CD25− cells were transfected with 50 μmol/L siRNA in 200 μL of transfection medium.

Article Title: Involvement of Toll-Like Receptor 2 and Epidermal Growth Factor Receptor Signaling in Epithelial Expression of Airway Remodeling Factors
Article Snippet: .. The small interfering RNA (siRNA) against TLR2 and the no-effect control were from Santa Cruz Biotechnology (Santa Cruz, CA), and HiPerFect transfection reagent was from Qiagen (Valencia, CA). .. Anti–TGF-α antibody and rabbit isotype control were purchased from Abcam (Cambridge, MA).

Article Title: Notch2 regulates matrix metallopeptidase 9 via PI3K/AKT signaling in human gastric carcinoma cell MKN-45
Article Snippet: .. MKN-45 cells were transfected with small interfering RNA (siRNA) against Notch2 and scrambled siRNA (Santa Cruz Biotechnology, CA, United States) constructs using the commercial transfection reagent (Santa Cruz Biotechnology, CA, United States) according to the manufacturer’s instructions. .. Following transfection, cells were incubated at 37 °C in a CO2 incubator for 48 h before being harvested for the assays described below.

Article Title: Superoxide from NADPH oxidase upregulates type 5 phosphodiesterase in human vascular smooth muscle cells: inhibition with iloprost and NONOate
Article Snippet: .. Predesigned small interfering RNA (siRNA) targeting human NOX-1 and NOX-4 and scrambled siRNA (as a negative control) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). .. Growth-arrested hVSMCs were trypsinized, pelleted and resuspended in nucleofector solution, provided in the Basic nucleofector kit for primary smooth muscle cells (Amaxa biosystems, Cologne, Germany), at a density of 0.8 × 106 cells (100 μL)−1 .

Article Title: Ginkgetin induces apoptosis in 786-O cell line via suppression of JAK2-STAT3 pathway
Article Snippet: .. Small interfering RNA and transient transfection The small interfering RNA (siRNA) targeting human STAT3 and control siRNA were from Santa Cruz. ..

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: .. CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. Membranes were stripped and re‐probed with anti‐ β ‐actin antibody (Santa Cruz Biotechnology) as an internal control.

Article Title: Novel 1,3,4-Oxadiazole Induces Anticancer Activity by Targeting NF-κB in Hepatocellular Carcinoma Cells
Article Snippet: .. The small interfering RNA (siRNA) for NF-κB and scrambled control was obtained from Santa Cruz Biotechnology. .. Caspase-Glo 3/7 assay kit and luciferase substrate was purchased from promega (WI, USA).

Construct:

Article Title: Notch2 regulates matrix metallopeptidase 9 via PI3K/AKT signaling in human gastric carcinoma cell MKN-45
Article Snippet: .. MKN-45 cells were transfected with small interfering RNA (siRNA) against Notch2 and scrambled siRNA (Santa Cruz Biotechnology, CA, United States) constructs using the commercial transfection reagent (Santa Cruz Biotechnology, CA, United States) according to the manufacturer’s instructions. .. Following transfection, cells were incubated at 37 °C in a CO2 incubator for 48 h before being harvested for the assays described below.

Reverse Transcription Polymerase Chain Reaction:

Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
Article Snippet: .. Silencing of RAGE Gene Expression and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) Detection for RAGE mRNA and HMGB1 Release RNA silencing was performed with small interfering RNA (siRNA) targeting mouse RAGE mRNA or negative control siRNA (Santa Cruz, catalog: sc-36375). .. Cells were transfected with siRNA duplexes suspended in lipofectamine reagent (Life Technologies) following the manufacturer's protocol.

Concentration Assay:

Article Title: Superoxide from NADPH oxidase upregulates type 5 phosphodiesterase in human vascular smooth muscle cells: inhibition with iloprost and NONOate
Article Snippet: Predesigned small interfering RNA (siRNA) targeting human NOX-1 and NOX-4 and scrambled siRNA (as a negative control) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). .. The desired siRNA was then introduced into each reaction mixture at a final concentration of 80 pmoles (1 μg) and transfection was carried out using Nucleofector II device no. 300661 (Amaxa biosystems), according to the manufacturer's instructions.

Incubation:

Article Title: Diabetic Retinopathy and Signaling Mechanism for Activation of Matrix Metalloproteinase-9
Article Snippet: BRECs from 3rd to 4th passage were transfected with small interfering RNA (siRNA) of H-Ras, Raf-1, MEK, or MMP9 using transfection reagents and siRNA duplex from Santa Cruz Biotechnology (Santa Cruz, CA). .. The mixture was incubated for 30 min at room temperature, and the cells were washed with the transfection medium and incubated with the transfection complex for 8 h at 37°C.

Article Title: Notch2 regulates matrix metallopeptidase 9 via PI3K/AKT signaling in human gastric carcinoma cell MKN-45
Article Snippet: MKN-45 cells were transfected with small interfering RNA (siRNA) against Notch2 and scrambled siRNA (Santa Cruz Biotechnology, CA, United States) constructs using the commercial transfection reagent (Santa Cruz Biotechnology, CA, United States) according to the manufacturer’s instructions. .. Following transfection, cells were incubated at 37 °C in a CO2 incubator for 48 h before being harvested for the assays described below.

shRNA:

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: To establish stable cell lines, lentiviral particles encoding MTDH Complementary deoxyribonucleic acid (cDNA) or short hairpin ribonucleic acid (shRNA) and the corresponding control plasmids were transfected into Tu686 and 5-8F cell lines according to the manufacturers’ instructions. .. Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA).

Cell Culture:

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: Paragraph title: Cell Culture and Transfection ... Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA).

Article Title: Programmed Death 1 and Cytokine Inducible SH2-Containing Protein Dependent Expansion of Regulatory T Cells Upon Stimulation With Mycobacterium tuberculosis
Article Snippet: CD4+ CD25− cells were transfected with small interfering RNA (siRNA) for CISH or cAMP response element modulator (CREM) or with scrambled siRNA (Santa Cruz Biotechnology). .. After 6 h, 200 μL of RPMI complete medium was added, and cells were cultured overnight in a 12-well plate.

Expressing:

Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
Article Snippet: .. Silencing of RAGE Gene Expression and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) Detection for RAGE mRNA and HMGB1 Release RNA silencing was performed with small interfering RNA (siRNA) targeting mouse RAGE mRNA or negative control siRNA (Santa Cruz, catalog: sc-36375). .. Cells were transfected with siRNA duplexes suspended in lipofectamine reagent (Life Technologies) following the manufacturer's protocol.

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: .. Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA). ..

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: .. CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. Membranes were stripped and re‐probed with anti‐ β ‐actin antibody (Santa Cruz Biotechnology) as an internal control.

Modification:

Article Title: Metadherin Regulation of Vascular Endothelial Growth Factor Expression Is Dependent Upon the PI3K/Akt Pathway in Squamous Cell Carcinoma of the Head and Neck
Article Snippet: Cell Culture and Transfection Tu686 cells were received from Dr Zhuo Chen of Emory University's Winship Cancer Institute in Atlanta, GA, and maintained as monolayer cultures in Dulbecco's Modified Eagle Media/F12 medium (1:1) supplemented with 10% fetal bovine serum (FBS). .. Tu686 and 5-8F cells stably expressing MTDH were transiently transfected with AKT or control small interfering ribonucleic acid (siRNA) (SC-43609; SC-37007, purchased from Santa Cruz, CA).

Sequencing:

Article Title: Endocytosis of Red Blood Cell Microparticles by Pulmonary Endothelial Cells is Mediated by Rab5
Article Snippet: Confluent monolayers of MLECs were treated with small interfering ribonucleic acid (siRNA) specific for Rab5 GTPase for 48–72 hours per manufacturer protocol (Santa Cruz Biotechnology, Dallas, TX). .. The nonsense (NS) siRNA control duplex sequence was used as control.

Caspase-Glo Assay:

Article Title: Novel 1,3,4-Oxadiazole Induces Anticancer Activity by Targeting NF-κB in Hepatocellular Carcinoma Cells
Article Snippet: The small interfering RNA (siRNA) for NF-κB and scrambled control was obtained from Santa Cruz Biotechnology. .. Caspase-Glo 3/7 assay kit and luciferase substrate was purchased from promega (WI, USA).

Chromogenic In Situ Hybridization:

Article Title: Programmed Death 1 and Cytokine Inducible SH2-Containing Protein Dependent Expansion of Regulatory T Cells Upon Stimulation With Mycobacterium tuberculosis
Article Snippet: .. CD4+ CD25− cells were transfected with small interfering RNA (siRNA) for CISH or cAMP response element modulator (CREM) or with scrambled siRNA (Santa Cruz Biotechnology). .. Briefly, 106 CD4+ CD25− cells were transfected with 50 μmol/L siRNA in 200 μL of transfection medium.

Binding Assay:

Article Title: Novel 1,3,4-Oxadiazole Induces Anticancer Activity by Targeting NF-κB in Hepatocellular Carcinoma Cells
Article Snippet: Nuclear extraction and NF-κB DNA binding kits were purchased from Active motif (USA). .. The small interfering RNA (siRNA) for NF-κB and scrambled control was obtained from Santa Cruz Biotechnology.

Imaging:

Article Title: Decline of miR‐124 in myeloid cells promotes regulatory T‐cell development in hepatitis C virus infection
Article Snippet: CD33+ myeloid cells, isolated from HCV‐infected patients or following small interfering RNA (siRNA) and miRNA transfections, were subjected to Western blot analysis to measure the expression of ERI‐1 and STAT3 using anti‐ERI‐1 (Santa Cruz Biotechnology, Santa Cruz, CA), anti‐STAT3 (Cell Signaling Technology, Danvers, MA) primary antibodies, followed by a horseradish peroxidase‐conjugated secondary antibody (Cell Signaling). .. The proteins were visualized using the Amersham ECL Prime Western Blotting Detection Reagents (GE Healthcare Biosciences, Pittsburgh, PA) and Bio‐Rad chemiDocMP imaging system (Bio‐Rad, Philadelphia, PA).

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    Santa Cruz Biotechnology tsg 6 sirna
    In vitro effect of <t>TSG-6</t> on caerulein- and LPS-stimulated primary pancreatic acinar cells (PACs). a TSG-6 mRNA expression levels in hAT-MSCs transfected with TSG-6 <t>siRNA</t> (TSG-6 siRNA MSCs), hAT-MSCs transfected with control siRNA (scr siRNA MSCs), and naive hAT-MSCs (MSCs). b , c Caerulein- and LPS-stimulated primary PACs were indirectly co-cultured with each of the three hAT-MSC types for 12 h. b Grp78, CHOP, and caspase-12 mRNA expression levels in primary PACs. c Grp78, CHOP, and caspase-12 protein expression levels. d The expression levels of NF-κB p65 activity were analysed by western blotting. e TUNEL staining in PACs in each group observed at ×200 magnification. TUNEL staining revealed the number of cells undergoing apoptosis or pyroptosis. Results are presented as the mean ± SD obtained from three independent experiments * P
    Tsg 6 Sirna, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 97/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology rna sirna
    (A) Knockdown of p65 by small interfering <t>RNA</t> <t>(siRNA)</t> reduces the apoptotic effect of CMO. HepG2 cells were transfected with either control or p65 specific siRNA (50 nM). After 48 h, the cells were treated with CMO (25 or 50 µM) for 24 h, and the enzymatic activity of caspase-3/7 was determined by Caspase-Glo ® 3/7 assay kit. (B) CMO increases the cleavage of PARP and Caspase 3 in HCCLM3 cells. HCCLM3 cells were treated with 50 µM CMO for 12, 24, 36, and 48 h, after which, the whole-cell extracts were prepared, and 30 µg of protein was resolved on 12% SDS-PAGE gel, electrotransferred onto nitrocellulose membranes, and probed for cleaved PARP and cleaved caspase 3 antibodies. The data are expressed as mean ± SD, compared with the untreated control (* p
    Rna Sirna, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology control sirna
    <t>TRP53-regulated</t> EndMT modulates the M1 and M2 populations of increased TAMs after radiotherapy. a RNA-seq data analysis of the samples represented in Fig. 4a . Venn diagram depicting differentially expressed genes in irradiated HUVECs. Reverse-regulated genes ( > 1.2-fold) between TGFβR2 knockdown+IR and Trp53 -knockdown (with and without TGFβR2 knockdown)+IR conditions (compared with IR alone) were selected (subgroup A). Only matrisome- and surface-associated genes in subset A are shown in the heatmap generated by k-means clustering (subgroup B). b GeneMANIA network analysis of subset B showing significant enrichment for the GO term ‘leukocyte migration’. Red-filled circles represent genes upregulated in the Trp53 <t>siRNA+IR</t> group vs. the IR-alone group. Blue-filled circles represent genes upregulated in the TGFβR2 siRNA+IR group vs. the IR-alone group. The names of leukocyte migration-related genes are indicated in red. c Immunofluorescence detection of F4/80, CD31, and Arg1, or iNOS, CD31, and F4/80 in KP tumours from WT and EC-p53KO mice, with or without irradiation (23 days after irradiation). Scale bar = 20 μm. d Quantification of F4/80 + Arg1 + and F4/80 + iNOS + cells/field (magnification, ×200) using immunofluorescence images of KP tumours from WT, EC-p53KO, and TGFβR2KD mice, with or without irradiation (23 days after irradiation). e Ratio of CD206 + in F4/80 + cells from WT and EC-p53KO tumours 7 days after irradiation, as determined by flow cytometry. f Immunofluorescence staining of F4/80 and Arg1 in WT bone marrow-derived macrophages after coculture with KP tumour-derived ECs from WT and EC-p53KO mice for 48 h after irradiation. Scale bar = 50 μm (crop, 20 μm). Flow-cytometric analysis of CD206 + , iNOS + , and EdU + cells among F4/80 + cells (magnification, ×200) is shown. For ( d – f ), error bars indicate SEM; * p
    Control Sirna, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 99/100, based on 1323 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    In vitro effect of TSG-6 on caerulein- and LPS-stimulated primary pancreatic acinar cells (PACs). a TSG-6 mRNA expression levels in hAT-MSCs transfected with TSG-6 siRNA (TSG-6 siRNA MSCs), hAT-MSCs transfected with control siRNA (scr siRNA MSCs), and naive hAT-MSCs (MSCs). b , c Caerulein- and LPS-stimulated primary PACs were indirectly co-cultured with each of the three hAT-MSC types for 12 h. b Grp78, CHOP, and caspase-12 mRNA expression levels in primary PACs. c Grp78, CHOP, and caspase-12 protein expression levels. d The expression levels of NF-κB p65 activity were analysed by western blotting. e TUNEL staining in PACs in each group observed at ×200 magnification. TUNEL staining revealed the number of cells undergoing apoptosis or pyroptosis. Results are presented as the mean ± SD obtained from three independent experiments * P

    Journal: Stem Cell Research & Therapy

    Article Title: TSG-6 secreted by human adipose tissue-derived mesenchymal stem cells ameliorates severe acute pancreatitis via ER stress downregulation in mice

    doi: 10.1186/s13287-018-1009-8

    Figure Lengend Snippet: In vitro effect of TSG-6 on caerulein- and LPS-stimulated primary pancreatic acinar cells (PACs). a TSG-6 mRNA expression levels in hAT-MSCs transfected with TSG-6 siRNA (TSG-6 siRNA MSCs), hAT-MSCs transfected with control siRNA (scr siRNA MSCs), and naive hAT-MSCs (MSCs). b , c Caerulein- and LPS-stimulated primary PACs were indirectly co-cultured with each of the three hAT-MSC types for 12 h. b Grp78, CHOP, and caspase-12 mRNA expression levels in primary PACs. c Grp78, CHOP, and caspase-12 protein expression levels. d The expression levels of NF-κB p65 activity were analysed by western blotting. e TUNEL staining in PACs in each group observed at ×200 magnification. TUNEL staining revealed the number of cells undergoing apoptosis or pyroptosis. Results are presented as the mean ± SD obtained from three independent experiments * P

    Article Snippet: The results showed that treatment with naive hAT-MSCs or hAT-MSCs transduced with scr siRNA may significantly reduce the apoptotic ratio, compared to the case in the positive control group; however, we failed to observe similar effects with hAT-MSCs transduced with TSG-6 siRNA.

    Techniques: In Vitro, Expressing, HAT Assay, Transfection, Cell Culture, Activity Assay, Western Blot, TUNEL Assay, Staining

    (A) Knockdown of p65 by small interfering RNA (siRNA) reduces the apoptotic effect of CMO. HepG2 cells were transfected with either control or p65 specific siRNA (50 nM). After 48 h, the cells were treated with CMO (25 or 50 µM) for 24 h, and the enzymatic activity of caspase-3/7 was determined by Caspase-Glo ® 3/7 assay kit. (B) CMO increases the cleavage of PARP and Caspase 3 in HCCLM3 cells. HCCLM3 cells were treated with 50 µM CMO for 12, 24, 36, and 48 h, after which, the whole-cell extracts were prepared, and 30 µg of protein was resolved on 12% SDS-PAGE gel, electrotransferred onto nitrocellulose membranes, and probed for cleaved PARP and cleaved caspase 3 antibodies. The data are expressed as mean ± SD, compared with the untreated control (* p

    Journal: Frontiers in Oncology

    Article Title: Novel 1,3,4-Oxadiazole Induces Anticancer Activity by Targeting NF-κB in Hepatocellular Carcinoma Cells

    doi: 10.3389/fonc.2018.00042

    Figure Lengend Snippet: (A) Knockdown of p65 by small interfering RNA (siRNA) reduces the apoptotic effect of CMO. HepG2 cells were transfected with either control or p65 specific siRNA (50 nM). After 48 h, the cells were treated with CMO (25 or 50 µM) for 24 h, and the enzymatic activity of caspase-3/7 was determined by Caspase-Glo ® 3/7 assay kit. (B) CMO increases the cleavage of PARP and Caspase 3 in HCCLM3 cells. HCCLM3 cells were treated with 50 µM CMO for 12, 24, 36, and 48 h, after which, the whole-cell extracts were prepared, and 30 µg of protein was resolved on 12% SDS-PAGE gel, electrotransferred onto nitrocellulose membranes, and probed for cleaved PARP and cleaved caspase 3 antibodies. The data are expressed as mean ± SD, compared with the untreated control (* p

    Article Snippet: The small interfering RNA (siRNA) for NF-κB and scrambled control was obtained from Santa Cruz Biotechnology.

    Techniques: Small Interfering RNA, Transfection, Activity Assay, Caspase-Glo Assay, SDS Page

    Knockdown of HSP72 abolishes the protection of FGF21 against hypoxia in CMECs. (A,B) The effects of siRNA targeting to HSP72 on the mRNA (A) and protein (B) expression of HSP72 in CMECs. * p

    Journal: Frontiers in Pharmacology

    Article Title: FGF21 Protects Against Hypoxia Injury Through Inducing HSP72 in Cerebral Microvascular Endothelial Cells

    doi: 10.3389/fphar.2019.00101

    Figure Lengend Snippet: Knockdown of HSP72 abolishes the protection of FGF21 against hypoxia in CMECs. (A,B) The effects of siRNA targeting to HSP72 on the mRNA (A) and protein (B) expression of HSP72 in CMECs. * p

    Article Snippet: siRNA-Mediated Knockdown The CMECs at 60% confluence in a 24-well plate were transfected with siRNA targeting HSP72 (Santa-Cruz Biotechnology, Santa-Cruz, CA, USA) or siRNA-scramble (siRNA-control) with Lipofectamine LTX Reagent (Invitrogen) and Opti-MEM TM (Invitrogen).

    Techniques: Expressing

    TRP53-regulated EndMT modulates the M1 and M2 populations of increased TAMs after radiotherapy. a RNA-seq data analysis of the samples represented in Fig. 4a . Venn diagram depicting differentially expressed genes in irradiated HUVECs. Reverse-regulated genes ( > 1.2-fold) between TGFβR2 knockdown+IR and Trp53 -knockdown (with and without TGFβR2 knockdown)+IR conditions (compared with IR alone) were selected (subgroup A). Only matrisome- and surface-associated genes in subset A are shown in the heatmap generated by k-means clustering (subgroup B). b GeneMANIA network analysis of subset B showing significant enrichment for the GO term ‘leukocyte migration’. Red-filled circles represent genes upregulated in the Trp53 siRNA+IR group vs. the IR-alone group. Blue-filled circles represent genes upregulated in the TGFβR2 siRNA+IR group vs. the IR-alone group. The names of leukocyte migration-related genes are indicated in red. c Immunofluorescence detection of F4/80, CD31, and Arg1, or iNOS, CD31, and F4/80 in KP tumours from WT and EC-p53KO mice, with or without irradiation (23 days after irradiation). Scale bar = 20 μm. d Quantification of F4/80 + Arg1 + and F4/80 + iNOS + cells/field (magnification, ×200) using immunofluorescence images of KP tumours from WT, EC-p53KO, and TGFβR2KD mice, with or without irradiation (23 days after irradiation). e Ratio of CD206 + in F4/80 + cells from WT and EC-p53KO tumours 7 days after irradiation, as determined by flow cytometry. f Immunofluorescence staining of F4/80 and Arg1 in WT bone marrow-derived macrophages after coculture with KP tumour-derived ECs from WT and EC-p53KO mice for 48 h after irradiation. Scale bar = 50 μm (crop, 20 μm). Flow-cytometric analysis of CD206 + , iNOS + , and EdU + cells among F4/80 + cells (magnification, ×200) is shown. For ( d – f ), error bars indicate SEM; * p

    Journal: Nature Communications

    Article Title: Tumour-vasculature development via endothelial-to-mesenchymal transition after radiotherapy controls CD44v6+ cancer cell and macrophage polarization

    doi: 10.1038/s41467-018-07470-w

    Figure Lengend Snippet: TRP53-regulated EndMT modulates the M1 and M2 populations of increased TAMs after radiotherapy. a RNA-seq data analysis of the samples represented in Fig. 4a . Venn diagram depicting differentially expressed genes in irradiated HUVECs. Reverse-regulated genes ( > 1.2-fold) between TGFβR2 knockdown+IR and Trp53 -knockdown (with and without TGFβR2 knockdown)+IR conditions (compared with IR alone) were selected (subgroup A). Only matrisome- and surface-associated genes in subset A are shown in the heatmap generated by k-means clustering (subgroup B). b GeneMANIA network analysis of subset B showing significant enrichment for the GO term ‘leukocyte migration’. Red-filled circles represent genes upregulated in the Trp53 siRNA+IR group vs. the IR-alone group. Blue-filled circles represent genes upregulated in the TGFβR2 siRNA+IR group vs. the IR-alone group. The names of leukocyte migration-related genes are indicated in red. c Immunofluorescence detection of F4/80, CD31, and Arg1, or iNOS, CD31, and F4/80 in KP tumours from WT and EC-p53KO mice, with or without irradiation (23 days after irradiation). Scale bar = 20 μm. d Quantification of F4/80 + Arg1 + and F4/80 + iNOS + cells/field (magnification, ×200) using immunofluorescence images of KP tumours from WT, EC-p53KO, and TGFβR2KD mice, with or without irradiation (23 days after irradiation). e Ratio of CD206 + in F4/80 + cells from WT and EC-p53KO tumours 7 days after irradiation, as determined by flow cytometry. f Immunofluorescence staining of F4/80 and Arg1 in WT bone marrow-derived macrophages after coculture with KP tumour-derived ECs from WT and EC-p53KO mice for 48 h after irradiation. Scale bar = 50 μm (crop, 20 μm). Flow-cytometric analysis of CD206 + , iNOS + , and EdU + cells among F4/80 + cells (magnification, ×200) is shown. For ( d – f ), error bars indicate SEM; * p

    Article Snippet: For silencing experiments, cells were transfected with siRNAs targeting Trp53 and Tgfbr2 as well as control siRNA (Santa Cruz Biotechnology) using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s recommendations.

    Techniques: RNA Sequencing Assay, Irradiation, Generated, Migration, Immunofluorescence, Mouse Assay, Flow Cytometry, Cytometry, Staining, Derivative Assay