Overexpression of Annexin A2 Receptor Inhibits Neovascularization via the Promotion of Krüppel-Like Transcription Factor 2

Abstract

Background/Aims: Annexin A2 receptor (AX2R) can mediate annexin A2 signalling and induce apoptosis in a variety of cells, but its role in neovascularization (NV) remains unclear. Krüppel-like transcription factor 2 (KLF2) is known to be expressed in a range of cell types and to participate in a number of processes during development and disease, such as endothelial homeostasis, vasoregulation and vascular growth/remodelling. The aim of our study was to investigate the role of AX2R in NV and the plausible molecular mechanism. Methods: We constructed a eukaryotic overexpression plasmid for AX2R (Lenti-AX2R) by using polymerase chain reaction (PCR). The full-length human AX2R gene was transfected into human retinal endothelial cells (HRECs) and human umbilical vein endothelial cells (HUVECs) using lentivirus vectors to overexpress AX2R. All experiments were divided into three groups: control, negative control (Lenti-EGFP), and Lenti-AX2R.Cell proliferation, cell migration, tube formation, mouse aortic ring assays and mouse matrigel plug assay were applied to analyse the effect of AX2R in NV. Furthermore, we conducted flow cytometry to evaluate whether AX2R could influence the cell cycle. A series of cell cycle-related proteins including cyclin A1, cyclin B1, cyclin D1, cyclin E1, CDK1, and p-CDC2 were detected by WB. The mRNA and protein levels of KLF2, vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptor 2 (VEGFR2) were further quantified by RT-PCR and WB to reveal the possible mechanism. Results: Overexpression of AX2R significantly inhibited cell proliferation, migration and tube formation in both types of endothelial cells (ECs), HRECs and HUVECs. It also suppressed vessel sprouting in the mouse aortic ring assay and NV in mouse matrigel plug assay. Furthermore, infection with Lenti-AX2R lentivirus arrested the cell cycle in S/G2 and influenced the expression of a series of cell cycle-related proteins. We also found that the overexpression of AX2R increased the expression of KLF2, mediating VEGF and VEGFR2. Conclusions: Overexpression of AX2R contributes to the inhibition of NV via suppressing KLF2 ubiquitin-dependent protein degradation, which might therefore be a therapeutic option for NV. It could be considered more broadly as an anti-angiogenic agent in the treatment of neovascular-related diseases in the future.

Multiomics Data Reveal the Important Role of ANXA2R in T Cell–mediated Rejection After Renal Transplantation

“…Beyond that, ANXA2R was also found to distribute in the cytoplasm and involved in the regulation of apoptosis and inhibition of angiogenesis in an Annexin II-independent manner. [50][51][52] In our study, ANXA2R was found to be highly expressed in patients with TCMR in both RNA and protein levels. Meanwhile, we found ANXA2R has strong positive correlations with multiple immune cells, especially the CD8 T cell.…”

Annexin A2 upregulation protects human retinal endothelial cells from oxygen‑glucose deprivation injury by activating autophagy

“…Accumulating evidence highlighted that the role of A2 in retinal neovascularization, which results in a number of severe eye diseases and even potentially blindness (3,11,20). Ischemic and hypoxic stimulation are common pathological states in many eye diseases associated with retinal neovascularization.…”

Cyclin‑dependent kinase 1 disruption inhibits angiogenesis by inducing cell cycle arrest and apoptosis

“…The cells that were authenticated by cell short tandem repeat genotyping were cultured at 37°C, in a 5% CO 2 cell incubator. Cells were identified via cell short tandem repeats (STRs) (16). When the cells were grown to ~80% confluence, they were used for the different assays.…”