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KLF4 protects brain microvascular endothelial cells from ischemic stroke induced apoptosis by transcriptionally
Haojie Yang1, Xixiang Xi2, Bin Zhao3
1Institute of TCM Surgery, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of TCM, Shanghai, 200437, China.
Abstract:
In this study, we investigated the expression profile and functional role of Kruppel-like family of transcription factor 4 (KLF4) in mouse cerebral microvascular endothelial cells (MECs) after focal cerebral ischemia and in cultured b.End3 cells after oxygen-glucose deprivation (OGD). Its downstream regulation was also studied. Our data showed that KLF4 was significantly upregulated in the cerebral MECs after transient ischemic insult and in cultured b.End3 cells after 24 h OGD exposure. In in vitro b.End3 cell model, KLF4 shRNA significantly increased OGD-induced activation of caspase-3 and also increased OGD-induced b.End3 cell death. KLF4 shRNA substantially enhanced OGD induced Bim and Bax expression at mRNA and protein levels and also aggravated OGD induced upregulation of E-selectin, MCP-1 and IL-6. Bioinformatic analysis suggested that MALAT1 promoter has a possible KLF4 binding site. In this study, we validated this possible binding site and also demonstrated that enforced KLF4 expression increased MALAT1 transcription. Functionally, knockdown of MALAT1 phenocopied the effect of KLF4 shRNA on enhancing OGD induced cell apoptosis and OGD induced upregulation of pro-apoptotic factors and pro-inflammatory cytokines. Based on these findings, we infer that MALAT1 is a transcriptional target of KLF4 in its protective role in cerebral MECs after ischemic insult.
Insights
Kruppel-like factor 4 (KLF4) protects brain cells after stroke by regulating MALAT1. Upregulated KLF4 reduces cell death and inflammation following ischemic injury.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Focal cerebral ischemia triggers cellular stress in brain microvascular endothelial cells (MECs).
- The role of Kruppel-like factor 4 (KLF4) in protecting MECs during ischemic events requires further elucidation.
Purpose of the Study:
- To investigate the expression and function of KLF4 in mouse MECs post-ischemia.
- To determine KLF4's downstream targets and their role in cerebral ischemia.
Main Methods:
- Analysis of KLF4 expression in MECs after transient ischemic insult and in b.End3 cells after oxygen-glucose deprivation (OGD).
- Utilized KLF4 shRNA and enforced KLF4 expression in vitro.
- Assessed apoptosis, pro-apoptotic factors (Bim, Bax), and inflammatory cytokines (E-selectin, MCP-1, IL-6).
- Bioinformatic analysis and experimental validation of KLF4 binding to MALAT1 promoter.
Main Results:
- KLF4 expression was significantly upregulated in MECs post-ischemia and in b.End3 cells post-OGD.
- KLF4 knockdown exacerbated OGD-induced cell death, caspase-3 activation, and expression of Bim, Bax, E-selectin, MCP-1, and IL-6.
- KLF4 directly binds to and upregulates MALAT1 transcription.
- MALAT1 knockdown mimicked KLF4 knockdown effects, increasing apoptosis and inflammation.
Conclusions:
- KLF4 plays a protective role in cerebral MECs against ischemic injury.
- MALAT1 is a key transcriptional target of KLF4, mediating its protective effects by reducing apoptosis and inflammation.
