Glomerular Endothelial Cell-Derived miR-200c Impairs Glomerular Homeostasis by Targeting Podocyte VEGF-A
Abstract
:1. Introduction
2. Results
2.1. miR-200c Is Upregulated in Urine from Patients with MCD, MGN and FSGS
2.2. miR-200c Overexpression Causes Proteinuria, Edema, Podocyte Effacement and Glomerular Endotheliosis in Zebrafish
2.3. miR-200c Is Induced due to Cell Stress in Glomerular Endothelial Cells
2.4. miR-200c Overexpression Decreases VEGF-A in Podocytes and Downregulates ZEBs in Podocytes and Glomerular Endothelial Cells
2.5. miR-200c Overexpression Decreases ZEB1 in Glomerular Endothelial Cells and ZEB1 and VEGF in Podocytes on Protein Level and Induces Cytoskeleton Rearrangement
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.1.1. Transient Transfection of miRNA Mimics
4.1.2. RNA Isolation, cDNA Transcription, and qPCR
- hGAPDH fw: 5′-GTCAGCCGCATCTTCTTTTG-3′
- hGAPDH rev: 5′-GCGCCCAATACGACCAAATC-3′
- hHPRT fw: 5′-GACCAGTCAACAGGGGACAT-3′
- hHPRT rev: 5′-AACACTTCGTGGGGTCCTTTTC-3′
- hVEGF-A fw: 5′-CAACAAATGTGAATGCAGACCAAA-3′
- hVEGF-A rev: 5′-CCCTTTCCCTTTCCTCGAACT-3′
- hZEB1 fw: 5′-CCAGCCAAATGGAAATCAGGATG-3′
- hZEB1 rev: 5′-TGGGCGGTGTAGAATCAGAG-3′
- hZEB2 fw: 5′-CCTCTGTAGATGGTCCAGAAGA-3′
- hZEB2 rev: 5′-AATTGCGGTCTGGATCGTGG-3′
- zvegf-aa fw: 5′-CAAGTGTGAATGCAGGCCAAA-3′
- zvegf-aa rev: 5′-AGCATTTACAGGTGAGGGGG-3′
- zvegf-ab fw: 5′- GAATGCCAGTGTCGGATGAAA-3′
- zvegf-ab rev: 5′-TAGCTGAAGTTAGGCAGGATGG-3′
- zhprt fw: 5′-ACCAAAACACTATGCGGCTG-3′
- zhprt re: 5′-GTGTCCACCCATGTCCTTCA-3′
4.1.3. MiR Isolation, cDNA Transcription and qPCR
4.1.4. Immunofluorescence Microscopy
4.2. ELISA
4.3. Zebrafish Experiments
Zebrafish Assay for Proteinuria
4.4. Transmission Electron Microscopy
4.5. Statistics
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Conflicts of Interest
References
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Ursu, R.; Sopel, N.; Ohs, A.; Tati, R.; Buvall, L.; Nyström, J.; Schiffer, M.; Müller-Deile, J. Glomerular Endothelial Cell-Derived miR-200c Impairs Glomerular Homeostasis by Targeting Podocyte VEGF-A. Int. J. Mol. Sci. 2022, 23, 15070. https://doi.org/10.3390/ijms232315070
Ursu R, Sopel N, Ohs A, Tati R, Buvall L, Nyström J, Schiffer M, Müller-Deile J. Glomerular Endothelial Cell-Derived miR-200c Impairs Glomerular Homeostasis by Targeting Podocyte VEGF-A. International Journal of Molecular Sciences. 2022; 23(23):15070. https://doi.org/10.3390/ijms232315070
Chicago/Turabian StyleUrsu, Raluca, Nina Sopel, Alexandra Ohs, Ramesh Tati, Lisa Buvall, Jenny Nyström, Mario Schiffer, and Janina Müller-Deile. 2022. "Glomerular Endothelial Cell-Derived miR-200c Impairs Glomerular Homeostasis by Targeting Podocyte VEGF-A" International Journal of Molecular Sciences 23, no. 23: 15070. https://doi.org/10.3390/ijms232315070