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TGF-β-mediated Endothelial to Mesenchymal Transition EndMT and the Functional Assessment of EndMT Effectors using CRISPR/Cas9 Gene Editing
Published on: February 26, 2021
The coordinated roles of miR-26a and miR-30c in regulating TGFβ1-induced epithelial-to-mesenchymal transition in
Zongji Zheng1, Meiping Guan1, Yijie Jia1
1Department of Endocrinology and Metabolism, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Abstract:
MicroRNAs (miRNAs) play vital roles in the development of diabetic nephropathy. Here, we compared the protective efficacies of miR-26a and miR-30c in renal tubular epithelial cells (NRK-52E) and determined whether they demonstrated additive effects in the attenuation of renal fibrosis. TGFβ1 suppressed miR-26a and miR-30c expression but up-regulated pro-fibrotic markers in NRK-52E cells, and these changes were also found in the kidney cortex of 40-week-old diabetic Otsuka Long-Evans Tokushima fatty (OLETF) rats. Bioinformatic analyses and luciferase assays further demonstrated that both miR-26a and miR-30c targeted connective tissue growth factor (CTGF); additionally, Snail family zinc finger 1 (Snail1), a potent epithelial-to-mesenchymal transition (EMT) inducer, was targeted by miR-30c. Overexpression of miR-26a and miR-30c coordinately decreased CTGF protein levels and subsequently ameliorated TGFβ1-induced EMT in NRK-52E cells. Co-silencing of miR-26a and miR-30c exhibited the opposite effect. Moreover, miR-26a and miR-30c co-silenced CTGF to decrease ERK1/2 and p38 MAPK activation. Furthermore, miR-26a was up-regulated in urinary extracellular vesicles of diabetic nephropathy patients. Our study provides evidence for the cooperative roles of miR-26a and miR-30c in the pathogenesis of diabetic nephropathy, and the co-targeting of miR-26a and miR-30c could provide a new direction for diabetic nephropathy treatment.
Insights
This study reveals that microRNAs miR-26a and miR-30c work together to reduce kidney fibrosis in diabetic nephropathy. Targeting both may offer a new treatment strategy for this condition.
Area of Science:
- Molecular Biology
- Renal Physiology
- Biochemistry
Background:
- Diabetic nephropathy is a serious complication of diabetes, characterized by progressive kidney damage and fibrosis.
- MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and are implicated in various diseases, including diabetic nephropathy.
- Understanding the specific roles of miRNAs in renal fibrosis is crucial for developing targeted therapies.
Purpose of the Study:
- To compare the protective effects of miR-26a and miR-30c in renal tubular epithelial cells (NRK-52E).
- To investigate the additive effects of miR-26a and miR-30c in attenuating renal fibrosis.
- To elucidate the molecular mechanisms underlying the actions of miR-26a and miR-30c in diabetic nephropathy.
Main Methods:
- Utilized NRK-52E cells and Otsuka Long-Evans Tokushima fatty (OLETF) rat models of diabetic nephropathy.
- Employed bioinformatic analyses and luciferase assays to identify miRNA targets.
- Performed gene silencing and overexpression experiments to assess miRNA functions.
- Analyzed protein levels and signaling pathways, including ERK1/2 and p38 MAPK.
Main Results:
- Transforming growth factor-beta 1 (TGFβ1) suppressed miR-26a and miR-30c expression while up-regulating pro-fibrotic markers in cells and rat kidneys.
- Both miR-26a and miR-30c were identified as direct targets of connective tissue growth factor (CTGF); miR-30c also targets Snail1, an epithelial-to-mesenchymal transition (EMT) inducer.
- Overexpression of miR-26a and miR-30c reduced CTGF levels, ameliorated TGFβ1-induced EMT, and decreased ERK1/2 and p38 MAPK activation.
- miR-26a was found to be upregulated in urinary extracellular vesicles of diabetic nephropathy patients.
Conclusions:
- miR-26a and miR-30c exhibit cooperative roles in the pathogenesis of diabetic nephropathy.
- The co-targeting of CTGF by miR-26a and miR-30c plays a significant role in mitigating renal fibrosis and EMT.
- Combined targeting of miR-26a and miR-30c presents a promising therapeutic avenue for diabetic nephropathy.
