Apoptosis induction in human lung and colon cancer cells via impeding VEGF signaling pathways.

Maryam Farzaneh Behelgardi,S. Mohsen Asghari,Zahra Gholami Shahvir

doi:10.1007/s11033-022-07203-9

Abstract

There is ample evidence to suggest that vascular endothelial growth factor (VEGF) is a potent mitogen factor in vasculogenesis and angiogenesis and that blockade of VEGF-mediated signals can also prevent tumor growth via enforcing cell apoptosis. In the current study, we assessed the suppressing effect of VGB4, a VEGF antagonist peptide with the binding ability to both VEGF receptor1 and VEGF receptor2, on VEGF-induced proliferation and migration of the human lung adenocarcinoma cell line A549 and the human colon adenocarcinoma cell line HT29 using MTT assay, colony formation assay, and Scratch-wound assay. To evaluate the apoptotic inductive effect of VGB4 on A549 and HT29 cells, apoptosis analysis was carried out by flow cytometry and TUNEL assay. Likewise, p53 and PTEN expression level was examined by immunofluorescence microscopy. In addition, the level of proteins involved in VEGF signaling pathways related to apoptosis was investigated using western blot analysis. Our results indicated that VGB4 markedly inhibited VEGF-induced proliferation and migration, and induced apoptosis of A549 and HT29 cells dose dependently. Encouragingly, significant downregulation of B-cell lymphoma 2 (Bcl2), X-linked inhibitor of apoptosis, Procaspase9, and procaspase3, as well as upregulation of PTEN and P53 tumor suppressors, BCL2 associated X, Cytochrome c, cleaved caspase9, and cleaved caspase3 in VGB4-treated A549 and HT29 cells, further confirmed the profound inductive influence of VGB4 on apoptotic pathways. These findings along with the results from our previous studies show that VGB4 may be considerable for cancer therapy.

Highlights

The phenomenon of tumorigenesis is caused due to an imbalance between cell proliferation and cell death
Our results show that a significant reduction in AKT phosphorylation, followed by VGB4 binding to VEGFR1 and VEGFR2 and inactivation of downstream signaling pathways, down-regulates the expression of the apoptotic-related factors in VGB4-treated A549 and HT29 cells
The results of MTT detection showed that the cell viability of A549 and HT29 cells was attenuated by VGB4 dose-dependently with IC50 value of 0.74 μM, whereas the scrambled (Scr) peptide had no inhibitory effect on VEGFA-induced cell proliferation even at 0.93 μM (Fig. 1A)

Summary

Introduction

The phenomenon of tumorigenesis is caused due to an imbalance between cell proliferation and cell death. Neovascularization, as a result of the constant activation of the VEGF/VEGFR signaling pathways, led to the uncontrolled cell proliferation in many types of human cancers by providing nutrients [1]. Some of the mechanisms engaged by cancer cells to avoid apoptosis initiation include upregulation of survival proteins (Bcl and IAPs), downregulation of pro-apoptotic effectors (BAX and Cytochrome c), loss of PTEN and p53 tumor suppressors, and inactivation of the final effectors of apoptosis (Caspases) [4,5,6]. Due to the important role of VEGF as one of the families implied in the regulation of vascular angiogenesis [7] and upregulation of VEGF and VEGF receptors in many types of cancers, therapeutic strategies against VEGF/VEGFR signaling pathways have been widely studied [8]. VEGF upregulates Bcl and suppresses apoptosis in human cancer cells and withdrawal of VEGF induces cell apoptosis both in vitro and in vivo [9,10,11]

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Results

Conclusion