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Molecular Analysis of Endothelial-mesenchymal Transition Induced by Transforming Growth Factor-β Signaling
Published on: August 3, 2018
TβRII Regulates the Proliferation of Metanephric Mesenchyme Cells through Six2 In Vitro
Zhaomin Mao1, Zhongshi Lyu2, Liyuan Huang3
1The M.O.E. Key Laboratory of Laboratory Medical Diagnostics, the College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, China. mao1204086118@163.com.
Abstract:
The transforming growth factor-β (TGFβ) family signaling pathways play an important role in regulatory cellular networks and exert specific effects on developmental programs during embryo development. However, the function of TGFβ signaling pathways on the early kidney development remains unclear. In this work, we aim to detect the underlying role of TGFβ type II receptor (TβRII) in vitro, which has a similar expression pattern as the crucial regulator Six2 during early kidney development. Firstly, the 5-ethynyl-2'-deoxyuridine (EdU) assay showed knock down of TβRII significantly decreased the proliferation ratio of metanephric mesenchyme (MM) cells. Additionally, real-time Polymerase Chain Reaction (PCR) and Western blot together with immunofluorescence determined that the mRNA and protein levels of Six2 declined after TβRII knock down. Also, Six2 was observed to be able to partially rescue the proliferation phenotype caused by the depletion of TβRII. Moreover, bioinformatics analysis and luciferase assay indicated Smad3 could transcriptionally target Six2. Further, the EdU assay showed that Smad3 could also rescue the inhibition of proliferation caused by the knock down of TβRII. Taken together, these findings delineate the important function of the TGFβ signaling pathway in the early development of kidney and TβRII was shown to be able to promote the expression of Six2 through Smad3 mediating transcriptional regulation and in turn activate the proliferation of MM cells.
Insights
Transforming growth factor-β type II receptor (TβRII) is crucial for early kidney development. TβRII promotes metanephric mesenchyme cell proliferation by upregulating Six2 expression via Smad3 signaling.
Area of Science:
- Developmental Biology
- Cell Signaling
- Renal Development
Background:
- Transforming growth factor-β (TGFβ) signaling is vital for cellular regulation and embryonic development.
- The specific role of TGFβ signaling in early kidney development is not fully understood.
- The TGFβ type II receptor (TβRII) shows a similar expression pattern to Six2, a key regulator in kidney development.
Purpose of the Study:
- To investigate the role of TβRII in early kidney development in vitro.
- To elucidate the molecular mechanisms by which TβRII influences metanephric mesenchyme (MM) cell proliferation and Six2 expression.
Main Methods:
- 5-ethynyl-2'-deoxyuridine (EdU) assay to assess cell proliferation.
- Real-time Polymerase Chain Reaction (PCR) and Western blot to analyze gene and protein expression.
- Immunofluorescence to confirm protein localization and levels.
- Bioinformatics analysis and luciferase assay to identify transcriptional targets.
- Gene knockdown experiments to evaluate the function of TβRII and Smad3.
Main Results:
- Knockdown of TβRII significantly reduced MM cell proliferation.
- TβRII knockdown led to decreased mRNA and protein levels of Six2.
- Six2 partially rescued the proliferation defect caused by TβRII depletion.
- Smad3 was identified as a transcriptional target of Six2.
- Smad3 activation rescued the proliferation inhibition resulting from TβRII knockdown.
Conclusions:
- TβRII plays a critical role in promoting early kidney development by regulating MM cell proliferation.
- TβRII enhances Six2 expression through Smad3-mediated transcriptional regulation.
- This pathway is essential for activating MM cell proliferation during kidney development.
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