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Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling

  • Authors:
    • Shaolin Li
    • Chenge Zhao
    • Shengqin Yu
    • Kaijing Yang
    • Shuying Zhang
    • Sixu Liu
  • View Affiliations / Copyright

    Affiliations: Department of Cardiology, Affiliated Zhongshan Hospital of Dalian University, Dalian, Liaoning 116000, P.R. China, Department of Cardiology, The Fifth Affiliated Hospital of Jinan University, Heyuan, Guangdong 517000, P.R. China
    Copyright: © Li et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 27
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    Published online on: November 21, 2025
       https://doi.org/10.3892/etm.2025.13022
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Abstract

Endothelial dysfunction is a key pathophysiological basis of atherosclerosis (AS). Potential mechanisms by which Hcy causes vascular injury include inhibiting endothelial cell growth, inducing endothelial dysfunction, and promoting vascular remodeling. Suppression of GPX4 synthesis can lead to exacerbated lipid peroxidation, triggering ferroptosis. The objective of the present study was to investigate whether ginsenoside Re attenuates homocysteine (Hcy)‑induced endothelial cell ferroptosis by upregulating glutathione peroxidase 4 (GPX4). After treating EA.hy926 cells with different concentrations of Hcy for 24 h, cell viability was assessed using an MTT assay to determine the appropriate concentrations of Hcy for establishing a cell damage model. EA.hy926 cells were then divided into the following four groups: Control group, Hcy group, low‑dose ginsenoside Re + Hcy group and high‑dose ginsenoside Re + Hcy group. Cell viability was assessed using an MTT assay, whereas the BODIPY™ 581/591 C11 probe was used to measure cellular lipid peroxidation levels. Additionally, a 2',7'‑dichlorodihydrofluorescein diacetate probe was used to measure the intracellular reactive oxygen species (ROS) content, while a microplate reader was used in combination with corresponding assay kits to measure the intracellular glutathione (GSH), malondialdehyde (MDA) and total iron ion levels. Furthermore, western blotting was conducted to determine the expression levels of GPX4, solute carrier family 7 member 11 (SLC7A11) and acyl‑CoA synthetase long‑chain family member 4 in the cells. Results demonstrated that high‑dose and low‑dose ginsenoside Re significantly alleviated the reduction in cell viability induced by Hcy and reduced the increase in ROS and lipid peroxide levels caused by Hcy. High concentrations of ginsenoside Re effectively mitigated the increase in MDA and total iron ion levels and the decrease in GSH levels induced by Hcy. Furthermore, western blotting results revealed that compared with the control group, the Hcy group exhibited lower expression levels of GPX4 and SLC7A11, while ACSL4 expression was elevated. By contrast, both low‑ and high‑concentration ginsenoside Re significantly increased GPX4 and SLC7A11 expression levels and decreased ACSL4 expression levels compared with the Hcy group. In conclusion, ginsenoside Re significantly increased the expression levels of GPX4 in EA.hy926 cells and alleviated Hcy‑induced endothelial cell ferroptosis. Ginsenoside Re may prevent microvascular endothelial dysfunction and subsequent tissue damage by reducing ferroptosis and protecting endothelial cells.
View Figures

Figure 1

GPX4/SLC7A11 is a key pathway in
ferroptosis. Homocysteine inhibition of the GPX4/SLC7A11 pathway
leads to lipid peroxidation and promotes ferroptosis. Ginsenoside
Re can attenuate lipid peroxidation by increasing the expression of
GPX4 and protecting endothelial cells from ferroptosis. The red
symbols indicate inhibition and the green symbol indicates
promotion. GPX4, glutathione peroxidase 4; SLC7A11; solute carrier
family 7 member 11; ROS, reactive oxygen species; GSH,
glutathione.

Figure 2

Hcy induces ferroptosis in EA.hy926
endothelial cells. (A) Effect of 0-5 mM Hcy on the viability of
EA.hy926 cells as detected by MTT assay. (B) Expression levels of
GPX4, SLC7A11 and ACSL4 detected by western blotting after
treatment of EA.hy926 cells with 2 or 5 mM Hcy for 24 h, (C) with
the optical density of the proteins calculated. (D) Cells were
stained with the BODIPY 581/591 C11 probe and lipid peroxidation
levels were detected by inverted fluorescence microscopy (x10
magnification). Data are presented as the mean ± SD.
*P<0.05 vs. Ctrl. GPX4, glutathione peroxidase 4;
SLC7A11; solute carrier family 7 member 11; ACSL4, acyl-CoA
synthetase long-chain family member 4; Hcy, homocysteine; Ctrl,
control.

Figure 3

Ginsenoside Re attenuates
erastin-induced endothelial cell ferroptosis. (A) Effects of 0-200
µM ginsenoside Re on the viability of EA.hy926 cells detected by
MTT assay. (B) Erastin (0-20 µM) was added to EA.hy926 cells and
cell viability was detected by MTT assay. (C) EA.hy926 cells were
co-incubated with 5 µM erastin or 5 µM erastin + 12 or 24 µM
ginsenoside Re for 24 h and cell viability was determined using an
MTT assay. (D) Representative bands and (E) quantification analysis
showing protein levels of GPX4 determined by western blotting with
the optical density of the proteins calculated. Data are presented
as the mean ± SD. *P<0.05 vs. Ctrl,
+P<0.05 vs. Erastin, #P<0.05 vs.
Erastin + Re 12. GPX4; glutathione peroxidase 4; Ctrl, control; Re,
ginsenoside Re.

Figure 4

Ginsenoside Re alleviates Hcy-induced
endothelial cell ferroptosis. (A) Hcy (2 mM), Hcy (2 mM) +
ginsenoside Re (12 µM) and Hcy (2 mM) + ginsenoside Re (24 µM) were
added to EA.hy926 cells and the cell viability was determined. (B)
Flow cytometric analysis of intracellular reactive oxygen species
levels in EA.hy926 cells. Detection of intracellular (C) iron ion,
(D) GSH and (E) MDA levels in EA.hy926 cells. Data are presented as
the mean ± SD. *P<0.05 vs. Ctrl;
+P<0.05 vs. Hcy; #P<0.05 vs. Hcy + Re
12. Hcy, homocysteine; Ctrl, control; GSH, glutathione; MDA,
malondialdehyde; Re, ginsenoside Re; DCFH-DA,
2',7'-dichlorodihydrofluorescein diacetate.

Figure 5

Effect of ginsenoside Re on Lip-ROS
levels in EA.hy926 cells. (A) Images (x10 magnification) and (B)
analysis of the detection of Lip-ROS levels through fluorescence
microscopy after cotreatment of EA.hy926 cells with Hcy and
ginsenoside Re for 24 h. (C) Flow cytometry plots and (D) analysis
of the detection of Lip-ROS levels in EA.hy926 cells. Data are
presented as the mean ± SD. *P<0.05 vs. Ctrl;
#P<0.05 vs. Hcy. Lip-ROS, lipid reactive oxygen
species, Hcy, homocysteine; Ctrl, control; Re, ginsenoside Re.

Figure 6

Optimal docking map of ginsenoside Re
and key gene molecules. Ginsenoside Re can bind to (A) GPX4, (B)
ACSL4 and (C) SLC7A11. The yellow dotted line represents hydrogen
bonds, and the number beside the dotted line indicates the length
of the hydrogen bond in Ångströms. GPX4, glutathione peroxidase 4;
SLC7A11; solute carrier family 7 member 11; ACSL4, acyl-CoA
synthetase long-chain family member 4; MET, methionine; LYS,
lysine; ASP, aspartate; ARG, arginine; GLY, glycine; GLN,
glutamine; HIS, histidine.

Figure 7

Effect of ginsenoside Re on the
expression of proteins related to Hcy-induced ferroptosis in
EA.hy926 cells. EA.hy926 cells were co-incubated with 2 mM Hcy, 2
mM Hcy + 12 µM ginsenoside Re or 2 mM Hcy + 24 µM ginsenoside Re
for 24 h. (A) Representative bands and (B) analysis of protein
levels of GPX4, SLC7A11 and ACSL4 determined by western blotting.
The optical density of the proteins was calculated. Data are
presented as the mean ± SD. *P<0.05 vs. Ctrl;
#P<0.05 vs. Hcy; +P<0.05 vs. Hcy + Re
12. GPX4, glutathione peroxidase 4; SLC7A11; solute carrier family
7 member 11; ACSL4, acyl-CoA synthetase long-chain family member 4;
Hcy, homocysteine; Ctrl, control; Re, ginsenoside Re.
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Copy and paste a formatted citation
Spandidos Publications style
Li S, Zhao C, Yu S, Yang K, Zhang S and Liu S: Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling. Exp Ther Med 31: 27, 2026.
APA
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., & Liu, S. (2026). Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling. Experimental and Therapeutic Medicine, 31, 27. https://doi.org/10.3892/etm.2025.13022
MLA
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., Liu, S."Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling". Experimental and Therapeutic Medicine 31.1 (2026): 27.
Chicago
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., Liu, S."Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling". Experimental and Therapeutic Medicine 31, no. 1 (2026): 27. https://doi.org/10.3892/etm.2025.13022
Copy and paste a formatted citation
x
Spandidos Publications style
Li S, Zhao C, Yu S, Yang K, Zhang S and Liu S: Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling. Exp Ther Med 31: 27, 2026.
APA
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., & Liu, S. (2026). Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling. Experimental and Therapeutic Medicine, 31, 27. https://doi.org/10.3892/etm.2025.13022
MLA
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., Liu, S."Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling". Experimental and Therapeutic Medicine 31.1 (2026): 27.
Chicago
Li, S., Zhao, C., Yu, S., Yang, K., Zhang, S., Liu, S."Ginsenoside Re attenuates homocysteine‑induced endothelial cell ferroptosis through upregulation of GPX4/xCT signaling". Experimental and Therapeutic Medicine 31, no. 1 (2026): 27. https://doi.org/10.3892/etm.2025.13022
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