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Updated: Apr 17, 2026
Detection of microRNA Expression in Peritoneal Membrane of Rats Using Quantitative Real-time PCR
Published on: June 27, 2017
The abnormal expression level of microRNA in epithelial-mesenchymal transition of peritoneal mesothelial cells
1Department of Nephrology, Shanghai General Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, P. R. China. yuqingsl0618@163.com.
Objective:
To determine the expression level of the microRNA in the process of epithelial-mesenchymal transition (EMT) of the peritoneal mesothelial cells (PMCs) induced by high glucose.
Materials And Methods:
The PMCs were cultured using M199 medium with 10% fetal bovine serum, and the EMT was induced by D-glucose stimulation. Epithelial-mesenchymal transition was determined by changes in cell morphology and the expression levels of the EMT marker genes. Changes in cell morphology were observed by inverted microscope, and the expression levels of the EMT marker genes were determined by real-time PCR. The expression levels of the microRNA were detected by real-time PCR with microRNA-specific stem-loop structure primer.
Results:
The PMCs changed to fusiformis following a high-glucose medium stimulated for 48 hours, and the EMT marker genes changed significantly, such as the decrease of E-cadherin and an increase of Vimentin (p < 0.01). These results proved the EMT had been induced by high-glucose. Applying real-time PCR with microRNA-specific stem-loop structure primer, miR-193a increased notably (p < 0.01), and miR-15a and let-7e decreased (p < 0.01), while miR-16 and miR-21 had no significant changes (p > 0.05). Most importantly, the increase of miR-193a was correlated with stimulus duration.
Conclusions:
MicroRNA with abnormal expression levels have a primary role in regulating the EMT of PMCs induced by high glucose.
Insights
High glucose induces epithelial-mesenchymal transition (EMT) in peritoneal mesothelial cells (PMCs). Specific microRNAs, including miR-193a, are dysregulated during this process, suggesting a regulatory role.
Area of Science:
- Cell Biology
- Molecular Biology
- Renal Physiology
Background:
- Epithelial-mesenchymal transition (EMT) is a critical process in cellular development and disease.
- Peritoneal mesothelial cells (PMCs) play a vital role in maintaining the integrity of the peritoneal membrane.
- High glucose levels are implicated in the progression of peritoneal dialysis-related complications, often involving EMT.
Purpose of the Study:
- To investigate the expression profiles of microRNAs during high glucose-induced EMT in PMCs.
- To identify specific microRNAs involved in the regulation of EMT in peritoneal mesothelial cells under hyperglycemic conditions.
Main Methods:
- Primary culture of peritoneal mesothelial cells (PMCs).
- Induction of EMT using D-glucose stimulation.
- Assessment of EMT by morphological changes and expression of marker genes (e.g., E-cadherin, Vimentin) via real-time PCR.
- Quantification of microRNA expression using real-time PCR with specific primers.
Main Results:
- High glucose (48 hours) induced EMT in PMCs, evidenced by morphological changes and altered expression of EMT markers (decreased E-cadherin, increased Vimentin).
- Significant changes in microRNA expression were observed: miR-193a increased, while miR-15a and let-7e decreased (p < 0.01).
- The upregulation of miR-193a showed a positive correlation with the duration of high glucose exposure.
Conclusions:
- Abnormal expression of specific microRNAs plays a significant role in regulating high glucose-induced EMT in PMCs.
- miR-193a emerges as a key microRNA potentially involved in the pathogenesis of high glucose-mediated peritoneal damage.
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