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Updated: Mar 9, 2026
Isolation of Murine Retinal Endothelial Cells for Next-Generation Sequencing
Published on: October 11, 2021
LIN28A blocks the EndMT process of high glucose-induced HRMECs by stabilizing SIRT6 mRNA
Xueying Wang1, Zixu Huang2, Xiaohui Kuang3
1Department of Ophthalmology, Xinxiang Medical University, Henan Provincial People's Hospital, Henan Eye Hospital, Henan Eye Institute, Zhengzhou, People's Republic of China.
Background:
Diabetic retinopathy (DR) is a microvascular condition resulting from microangiopathy, causing gradual retinal damage and potential blindness. Endothelial-mesenchymal transition (EndMT) serves an important function in DR development. Exploring the molecular mechanism of EndMT in DR is needed.
Methods:
Human retinal microvascular endothelial cells (HRMECs) were incubated with high glucose (HG) to induce an in vitro DR model. Lentivirus and small-interfering RNAs were used to construct SIRT6 and LIN28A overexpression or knockdown in HRMECs, respectively. AMPK inhibitor, compound C, was used to block AMPK signaling in HRMECs. α-SMA and PECAM1 levels were identified using immunofluorescence. CCK8 and transwell were used to detect cell viability and migration, respectively. The mRNA stability of SIRT6 was analyzed after HRMECs were exposed to actinomycin D. RNA immunoprecipitation was applied to verify the binding relationship between LIN28A and SIRT6 mRNA in HRMECs.
Results:
In HG-induced HRMECs, the cells undergo the EndMT process, and SIRT6 levels are downregulated. HG treatment reduced the level of p-AMPK in HRMECs, and compound C reversed the inhibition of SIRT6 overexpression on EndMT in HG-induced HRMECs. By binding to SIRT6 mRNA, LIN28A could enhance SIRT6 mRNA stability. Additionally, LIN28A overexpression inhibited EndMT in HG-induced HRMECs, which was reversed by SIRT6 knockdown.
Conclusion:
The stabilization of SIRT6 mRNA by LIN28A activated AMPK signaling, inhibiting the EndMT process in HRMECs caused by HG.
Insights
LIN28A stabilizes SIRT6 mRNA, activating AMPK signaling to inhibit high glucose-induced endothelial-mesenchymal transition (EndMT) in diabetic retinopathy (DR). This finding offers a potential therapeutic target for preventing DR progression.
Area of Science:
- Ophthalmology
- Molecular Biology
- Cell Biology
Background:
- Diabetic retinopathy (DR) is a complication of diabetes causing retinal damage and blindness.
- Endothelial-mesenchymal transition (EndMT) plays a key role in DR pathogenesis.
- Understanding the molecular mechanisms of EndMT in DR is crucial for developing treatments.
Purpose of the Study:
- To investigate the role of LIN28A and SIRT6 in high glucose-induced EndMT in human retinal microvascular endothelial cells (HRMECs).
- To elucidate the molecular pathway involving LIN28A, SIRT6, and AMPK signaling in DR.
Main Methods:
- An in vitro DR model was established using HRMECs exposed to high glucose (HG).
- Lentivirus and small-interfering RNAs were used to manipulate SIRT6 and LIN28A expression.
- Cell viability, migration, and protein/mRNA levels were assessed using CCK8, transwell assays, immunofluorescence, and RNA immunoprecipitation.
Main Results:
- High glucose induced EndMT and downregulated SIRT6 in HRMECs.
- LIN28A overexpression inhibited HG-induced EndMT by enhancing SIRT6 mRNA stability.
- LIN28A-mediated stabilization of SIRT6 activated AMPK signaling, thereby inhibiting EndMT.
Conclusions:
- LIN28A stabilizes SIRT6 mRNA, which in turn activates AMPK signaling.
- This LIN28A-SIRT6-AMPK pathway effectively inhibits high glucose-induced EndMT in retinal endothelial cells.
- Targeting this pathway may offer a novel therapeutic strategy for diabetic retinopathy.
