Knockdown of circHECTD1 inhibits oxygen-glucose deprivation and reperfusion induced endothelial-mesenchymal
Guo-Hua He1, Zhen Wang2, Wei Xu2
1Department of Neurology, The affiliated Changsha Central Hospital, Hengyang Medical School, University of South China, No.161 Shaoshan Road, Changsha, 410004, Hunan Province, China. heguohh645@163.com.
Insights
Circular RNAs (circRNAs) like circHECTD1 are implicated in ischemic stroke (IS). Silencing circHECTD1 alleviates endothelial-mesenchymal transition (EndoMT) in IS by regulating the miR-335/NOTCH2 pathway, suggesting circHECTD1 as a potential therapeutic target.
Area of Science:
- Biomedical research
- Molecular biology
- Cerebrovascular diseases
Background:
- Ischemic stroke (IS) is a major health concern, particularly for the elderly.
- Circular RNAs (circRNAs) are increasingly recognized for their roles in various diseases, including IS.
- The specific function of circHECTD1 in IS pathogenesis is not well understood.
Purpose of the Study:
- To investigate the role of circHECTD1 in ischemic stroke (IS).
- To elucidate the underlying molecular mechanisms involving circHECTD1, miR-335, and NOTCH2 in IS.
- To evaluate circHECTD1 as a potential therapeutic target for IS.
Main Methods:
- In vitro models using oxygen-glucose deprivation/reperfusion (OGD/R) on human cerebral microvascular endothelial cells (HCMECs).
- In vivo MCAO mouse model to assess circHECTD1 expression in IS.
- Quantitative real-time PCR (qRT-PCR), Western blot, CCK-8 assay, transwell, and tube formation assays.
- RNA immunoprecipitation (RIP) and luciferase assays to determine molecular interactions.
Main Results:
- CircHECTD1 expression was significantly upregulated in IS models.
- OGD/R induced endothelial-mesenchymal transition (EndoMT) in HCMECs, which was reversed by circHECTD1 knockdown.
- Silencing circHECTD1 inhibited OGD/R-induced migration and tube formation of HCMECs.
- CircHECTD1 upregulated NOTCH2 by binding to miR-335, while miR-335 inhibited IS-related EndoMT by targeting NOTCH2.
Conclusions:
- CircHECTD1 knockdown alleviates IS-related EndoMT in HCMECs through the miR-335/NOTCH2 axis.
- CircHECTD1 plays a crucial role in IS pathogenesis by promoting EndoMT.
- CircHECTD1 represents a potential therapeutic target for treating ischemic stroke.
Abstract:
Ischemic stroke (IS) has become a cerebrovascular disease which seriously threatens the elderly people. It has been reported that circRNAs participate in multiple diseases, including IS. However, the role of circHECTD1 in IS remains largely unknown. To mimic IS in vitro, human cerebral microvascular endothelial cells (HCMECs) were treated with oxygen glucose deprivation/reperfusion (OGD/R). Meanwhile, MCAO mouse model was established to detect the expression of circHECTD1 in IS. qRT-PCR and western blot were used to test gene and protein expressions, respectively. CCK-8 assay was used to investigate the cell viability. Moreover, cell migration and tube formation were assessed by transwell and tube formation assays. In addition, RIP and luciferase assay were performed to explore the association among circHECTD1, miR-335 and NOTCH2. CircHECTD1 was significantly upregulated in IS. OGD/R significantly induced EndoMT in HCMECs, while knockdown of circHECTD1 notably reversed this phenomenon. In addition, silencing of circHECTD1 remarkably reversed OGD/R-induced promotion of HCMEC tube formation and migration. Meanwhile, circHECTD1 upregulated the level of NOTCH2 through binding with miR-335. Furthermore, miR-335 inhibited the process of EndoMT in IS via targeting NOTCH2. In summary, circHECTD1 knockdown significantly alleviated EndoMT process in HCMECs via mediation of miR-335/NOTCH2 axis. Thus, circHECTD1 might act as a potential target against IS.
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