Molecular validation of the precision-cut kidney slice (PCKS) model of renal fibrosis through assessment of TGF-β1-induced Smad and p38/ERK signaling
Copyright © 2016. Published by Elsevier B.V..
BACKGROUND: The precision-cut kidney slice (PCKS) model appears to be a useful ex vivo model of renal fibrosis. However, little in-depth molecular investigation on the PCKS model has been performed. Therefore, the aim of this study will be to investigate and validate the molecular validity of this model.
METHODS: The PCKS model was constructed in male C57BL/6 mice. To induce renal fibrosis, PCKS were incubated in recombinant human TGF-β1 for 48 h. Protein expression of phosphorylated Smad2 (p-Smad2, cytosolic and nuclear), Smad7, phosphorylated ERK1 (p-ERK1), phosphorylated ERK2 (p-ERK2), and phosphorylated p38 MAPK (p-p38 MAPK) was measured using Western blotting. To assess Smad2/3 heteromeric complex formation and phosphorylated Smad3 (p-Smad3) expression, immunoprecipitation was performed with an anti-Smad2 or an anti-Smad3 antibody, respectively, prior to Western blotting.
RESULTS: p-Smad2 and p-Smad3 were significantly upregulated in the PCKS model relative to control (p<0.05). However, we found no significant difference in Smad7 expression between the PCKS model and control (p>0.05). The PCKS model demonstrated significantly greater Smad2/3 complex formation and nuclear translocation relative to control (p<0.05). The PCKS model showed significantly greater expression of p-ERK1, p-ERK2, and p-p38 MAPK relative to control (p<0.05).
CONCLUSIONS: The PCKS model displays several well-established molecular markers of renal fibrosis. However, the PCKS model failed to display Smad7 downregulation and appears to display "over-activation" of p-Smad2 and p-Smad3 as well as "under-activation" of ERK1/2 and p38 MAPK signaling vis-à-vis the well-established in vivo unilateral ureteric obstruction model of renal fibrosis.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2016 |
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| Erschienen: |
2016 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:34 |
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| Enthalten in: |
International immunopharmacology - 34(2016) vom: 18. Mai, Seite 32-36 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Zhang, Shouhua [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 30.12.2016 Date Revised 10.12.2019 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.intimp.2016.01.026 |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM257882030 |
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| 100 | 1 | |a Zhang, Shouhua |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Molecular validation of the precision-cut kidney slice (PCKS) model of renal fibrosis through assessment of TGF-β1-induced Smad and p38/ERK signaling |
| 264 | 1 | |c 2016 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
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| 500 | |a Date Completed 30.12.2016 | ||
| 500 | |a Date Revised 10.12.2019 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a Copyright © 2016. Published by Elsevier B.V. | ||
| 520 | |a BACKGROUND: The precision-cut kidney slice (PCKS) model appears to be a useful ex vivo model of renal fibrosis. However, little in-depth molecular investigation on the PCKS model has been performed. Therefore, the aim of this study will be to investigate and validate the molecular validity of this model | ||
| 520 | |a METHODS: The PCKS model was constructed in male C57BL/6 mice. To induce renal fibrosis, PCKS were incubated in recombinant human TGF-β1 for 48 h. Protein expression of phosphorylated Smad2 (p-Smad2, cytosolic and nuclear), Smad7, phosphorylated ERK1 (p-ERK1), phosphorylated ERK2 (p-ERK2), and phosphorylated p38 MAPK (p-p38 MAPK) was measured using Western blotting. To assess Smad2/3 heteromeric complex formation and phosphorylated Smad3 (p-Smad3) expression, immunoprecipitation was performed with an anti-Smad2 or an anti-Smad3 antibody, respectively, prior to Western blotting | ||
| 520 | |a RESULTS: p-Smad2 and p-Smad3 were significantly upregulated in the PCKS model relative to control (p<0.05). However, we found no significant difference in Smad7 expression between the PCKS model and control (p>0.05). The PCKS model demonstrated significantly greater Smad2/3 complex formation and nuclear translocation relative to control (p<0.05). The PCKS model showed significantly greater expression of p-ERK1, p-ERK2, and p-p38 MAPK relative to control (p<0.05) | ||
| 520 | |a CONCLUSIONS: The PCKS model displays several well-established molecular markers of renal fibrosis. However, the PCKS model failed to display Smad7 downregulation and appears to display "over-activation" of p-Smad2 and p-Smad3 as well as "under-activation" of ERK1/2 and p38 MAPK signaling vis-à-vis the well-established in vivo unilateral ureteric obstruction model of renal fibrosis | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a Validation Study | |
| 650 | 4 | |a ERK | |
| 650 | 4 | |a Precision-cut kidney slice, PCKS | |
| 650 | 4 | |a Renal fibrosis | |
| 650 | 4 | |a Smad, Smad2, Smad3, Smad7 | |
| 650 | 4 | |a p38 | |
| 650 | 7 | |a Smad2 Protein |2 NLM | |
| 650 | 7 | |a Smad2 protein, mouse |2 NLM | |
| 650 | 7 | |a Smad3 Protein |2 NLM | |
| 650 | 7 | |a Smad3 protein, mouse |2 NLM | |
| 650 | 7 | |a Transforming Growth Factor beta1 |2 NLM | |
| 650 | 7 | |a Extracellular Signal-Regulated MAP Kinases |2 NLM | |
| 650 | 7 | |a EC 2.7.11.24 |2 NLM | |
| 650 | 7 | |a p38 Mitogen-Activated Protein Kinases |2 NLM | |
| 650 | 7 | |a EC 2.7.11.24 |2 NLM | |
| 700 | 1 | |a Liu, Queling |e verfasserin |4 aut | |
| 700 | 1 | |a Xiao, Juhua |e verfasserin |4 aut | |
| 700 | 1 | |a Lei, Jun |e verfasserin |4 aut | |
| 700 | 1 | |a Liu, Yi |e verfasserin |4 aut | |
| 700 | 1 | |a Xu, Hongyan |e verfasserin |4 aut | |
| 700 | 1 | |a Hong, Zhengdong |e verfasserin |4 aut | |
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