Functional Analysis of Dual-Specificity Protein Phosphatases in Angiogenesis
Therapeutic perspectives targeting angiogenesis in cancer stimulated an intense investigation of the mechanisms triggering and governing angiogenic processes. Several publications have highlighted the importance of typical dual-specificity phosphatases (DSPs) or MKPs in endothelial cells and their role in controlling different biological functions implicated in angiogenesis such as migration, proliferation, apoptosis, tubulogenesis, and cell adhesion. However, among atypical DSPs, the only one investigated in angiogenesis was DUSP3. We recently identified this DSP as a new key player in endothelial cells and angiogenesis. In this chapter we provide with detailed protocols and models used to investigate the role of DUSP3 in endothelial cells and angiogenesis. We start the chapter with an overview of the role of several DSPs in angiogenesis. We continue with providing a full description of a highly efficient transfection protocol to deplete DUSP3 using small interfering RNA (siRNA) in the primary human umbilical vein endothelial cells (HUVEC). We next describe the major assays used to investigate different processes involved in angiogenesis such as tube formation assay, proliferation assay and spheroids sprouting assay. We finish the chapter by validating our results in DUSP3-knockout mice using in vivo angiogenesis assays such as Matrigel plug and Lewis lung carcinoma cell subcutaneous xenograft model followed by anti-CD31 immunofluorescence and ex vivo aortic ring assay. All methods described can be adapted to other phosphatases and signaling molecules.
| Medienart: |
E-Artikel |
|---|
| Erscheinungsjahr: |
2016 |
|---|---|
| Erschienen: |
2016 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:1447 |
|---|---|
| Enthalten in: |
Methods in molecular biology (Clifton, N.J.) - 1447(2016) vom: 12., Seite 331-49 |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Amand, Mathieu [VerfasserIn] |
|---|
| Links: |
|---|
| Anmerkungen: |
Date Completed 02.01.2018 Date Revised 03.04.2018 published: Print Citation Status MEDLINE |
|---|
| doi: |
10.1007/978-1-4939-3746-2_18 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM263342026 |
|---|
| LEADER | 01000naa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM263342026 | ||
| 003 | DE-627 | ||
| 005 | 20231224203707.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231224s2016 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.1007/978-1-4939-3746-2_18 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n0877.xml |
| 035 | |a (DE-627)NLM263342026 | ||
| 035 | |a (NLM)27514814 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Amand, Mathieu |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Functional Analysis of Dual-Specificity Protein Phosphatases in Angiogenesis |
| 264 | 1 | |c 2016 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Completed 02.01.2018 | ||
| 500 | |a Date Revised 03.04.2018 | ||
| 500 | |a published: Print | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a Therapeutic perspectives targeting angiogenesis in cancer stimulated an intense investigation of the mechanisms triggering and governing angiogenic processes. Several publications have highlighted the importance of typical dual-specificity phosphatases (DSPs) or MKPs in endothelial cells and their role in controlling different biological functions implicated in angiogenesis such as migration, proliferation, apoptosis, tubulogenesis, and cell adhesion. However, among atypical DSPs, the only one investigated in angiogenesis was DUSP3. We recently identified this DSP as a new key player in endothelial cells and angiogenesis. In this chapter we provide with detailed protocols and models used to investigate the role of DUSP3 in endothelial cells and angiogenesis. We start the chapter with an overview of the role of several DSPs in angiogenesis. We continue with providing a full description of a highly efficient transfection protocol to deplete DUSP3 using small interfering RNA (siRNA) in the primary human umbilical vein endothelial cells (HUVEC). We next describe the major assays used to investigate different processes involved in angiogenesis such as tube formation assay, proliferation assay and spheroids sprouting assay. We finish the chapter by validating our results in DUSP3-knockout mice using in vivo angiogenesis assays such as Matrigel plug and Lewis lung carcinoma cell subcutaneous xenograft model followed by anti-CD31 immunofluorescence and ex vivo aortic ring assay. All methods described can be adapted to other phosphatases and signaling molecules | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a Angiogenesis | |
| 650 | 4 | |a Aortic ring assay | |
| 650 | 4 | |a DSPs | |
| 650 | 4 | |a DUSP3 | |
| 650 | 4 | |a Endothelial cells | |
| 650 | 4 | |a HUVEC | |
| 650 | 4 | |a Immunofluorescence | |
| 650 | 4 | |a LLC | |
| 650 | 4 | |a Matrigel plug assay | |
| 650 | 4 | |a Proliferation | |
| 650 | 4 | |a Spheroid assay | |
| 650 | 4 | |a Sprouting | |
| 650 | 7 | |a RNA, Small Interfering |2 NLM | |
| 650 | 7 | |a DUSP3 protein, human |2 NLM | |
| 650 | 7 | |a EC 3.1.3.48 |2 NLM | |
| 650 | 7 | |a Dual Specificity Phosphatase 3 |2 NLM | |
| 650 | 7 | |a EC 3.1.3.48 |2 NLM | |
| 650 | 7 | |a Dusp3 protein, mouse |2 NLM | |
| 650 | 7 | |a EC 3.1.3.48 |2 NLM | |
| 700 | 1 | |a Erpicum, Charlotte |e verfasserin |4 aut | |
| 700 | 1 | |a Gilles, Christine |e verfasserin |4 aut | |
| 700 | 1 | |a Noël, Agnès |e verfasserin |4 aut | |
| 700 | 1 | |a Rahmouni, Souad |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t Methods in molecular biology (Clifton, N.J.) |d 1984 |g 1447(2016) vom: 12., Seite 331-49 |w (DE-627)NLM074849794 |x 1940-6029 |7 nnns |
| 773 | 1 | 8 | |g volume:1447 |g year:2016 |g day:12 |g pages:331-49 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1007/978-1-4939-3746-2_18 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |d 1447 |j 2016 |b 12 |h 331-49 | ||