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MiR548ai antagonism attenuates exosome-induced endothelial cell dysfunction

ABSTRACT: Endothelial cell (EC) and smooth muscle cell (SMC) are major cell types adjacent in the vascular wall. Recent progress indicates that their communication is crucial for vascular homeostasis and pathogenesis. In particular, dysfunctional (proliferative) SMCs through exosomes can induce EC dysfunction (impaired growth). The current study suggests that miR548ai, a rarely known microRNA, may provide a molecular target for protection against SMC/exosome-induced EC dysfunction. We performed microarray profiling of microRNAs of dysfunctional human primary aortic SMCs induced by different cytokines (PDGF-BB, TGFβ1, TNFα, IL1β). Among the microRNAs commonly upregulated by these cytokines, miR548ai showed the most robust changes, as also validated through quantitative PCR. This cytokine-induced miR548ai upregulation was recapitulated in the qPCR determination of SMC-derived exosomal microRNAs. Consistent with SMC-to-EC communication, the exosomes extracted from cytokine-stimulated SMCs impaired human EC proliferation and migration. Of particular interest, this SMC exosomal impingement on ECs was countered by transfection of miR548ai inhibitor microRNA into ECs. Furthermore, the miR548ai inhibitor transfected into SMCs attenuated SMC dysfunction/proliferation. Thus, these results identify miR548ai as a novel target; namely, miR548ai inhibitor mitigates EC dysfunction induced by exosomes derived from dysfunctional SMCs. This new knowledge may aid the future development of microRNA-based treatment of vascular disorders.

SUBMITTER: Xie X

PROVIDER: S-EPMC8553949 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background: Endothelial dysfunction is commonly accompanied by a reduced capacity for nitric oxide (NO) production and decreased NO sensitivity, playing a central role in numerous vascular diseases. Saturated free fatty acids are known to reduce NO production and then induce endothelial dysfunction. Alternative splicing participates in the regulation of cellular and tissular homeostasis and is highly regulated by serine-arginine protein kinase (SRPK1). The role of SRPK1 in the biology of endothelial cells remains elusive. Icariside Ⅱ (ICA Ⅱ) has been reported to have protective effects on endothelial function. However, the specific molecular mechanisms are still unknown. The purpose of this study is to explore the role of SRPK1 in the biology of endothelial cells and the underlying mechanism of ICA Ⅱ on palmitic acid (PA) induced endothelial dysfunction. Methods: Endothelial dysfunction was induced using PA in human umbilical vein endothelial cells (HUVECs). The expression and phosphorylation of related proteins in the SRPK1-Akt-eNOS signaling pathway were detected by Western Blot. Cell Counting Kit-8 assay and Ki-67 immunofluorescence were used to estimate cell viability. Endothelial cell function was assessed by detecting NO production using DAF-FM DA. Interaction between ICA Ⅱ and SRPK1 was demonstrated by a biotinylated protein interaction pull-down assay. Results: The expressions of eNOS, Akt, and SRPK1 were down-regulated in the endothelial dysfunction stimulated by PA. SRPK1 inhibitor SPHINX31 restrained endothelial cell viability in a dose-dependent manner. Moreover, inhibition of SRPK1 using SPHINX31 and knockdown of SRPK1 by shRNA also showed a down-regulation of the proteins associated with the SRPK1-Akt-eNOS signaling pathway. Biotinylated protein interaction pull-down assay revealed that ICA Ⅱ could be directly bound with SRPK1. On the other hand, ICA Ⅱ could attenuate the PA-induced endothelial dysfunction and restore cell viability through the SRPK1-Akt-eNOS pathway. Conclusions: ICA Ⅱ, bound with SRPK1, could attenuate the endothelial dysfunction induced by the PA in HUVECs via the SRPK1-Akt-eNOS signaling pathway.

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MiR548ai antagonism attenuates exosome-induced endothelial cell dysfunction

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