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Article

Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas

  • Authors:
    • Ke Hu
    • Shichen Bu
    • Yi Guo
    • Yuxuan Li
    • Shiwen Yu
    • Lulu Wang
    • Chuanqi Cai
    • Yiqing Li
    • Xin Liu
    • Hegui Huang
    • Weici Wang
  • View Affiliations / Copyright

    Affiliations: Department of Vascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, P.R. China, Department of Cardiology, Xiamen Cardiovascular Hospital of Xiamen University, School of Medicine, Xiamen University, Xiamen, Fujian 361006, P.R. China, Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430022, P.R. China, Department of Pharmacy, Wuhan No. 1 Hospital, Wuhan, Hubei 430022, P.R. China
  • Article Number: 127
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    Published online on: June 23, 2025
       https://doi.org/10.3892/ijmm.2025.5568
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Abstract

Arteriovenous fistulas (AVFs) are preferred access points for hemodialysis. The present study aimed to investigate the function of early growth response‑1 (Egr‑1) in the proliferation and migration of smooth muscle cells (SMCs) and assess its potential as a new therapeutic target for AVF treatment. A comprehensive analysis combining public data‑source mining, human tissue collection, animal studies, cell culture experiments and various molecular biology techniques was conducted. The public dataset GSE119296 was used for immunohistochemical analyses of human AVF stenosis samples. SMC‑specific Egr‑1 knockout mice and various in vitro assays on primary rat vascular SMCs were used to evaluate the effect of Egr‑1 on the functional capacity of SMCs. RNA sequencing and chromatin immunoprecipitation sequencing was performed. Egr‑1 was upregulated in human AVF stenosis samples and cultured SMCs. Knockout of Egr‑1 in mice mitigated AVF outflow tract stenosis, improved flow dynamics and diminished neointima formation. In vitro, Egr‑1 ablation reduced SMC proliferation and migration; Egr‑1 transcriptionally activated Egr‑2. Increased Egr‑1 expression facilitated SMC proliferation and migration through Egr‑2 regulation, contributing to AVF stenosis. Consequently, targeting Egr‑1 may offer a novel therapeutic approach for managing AVF intimal hyperplasia and improving AVF patency and function in patients with end‑stage renal disease.
View Figures

Figure 1

Egr-1 is overexpressed in human AVF
stenosis. (A) Principal component analysis plot shows that controls
and disease groups are well clustered. (B) Expression of Egr-1 in
sequencing data. (C) ROC curve was drawn based on Egr-1 expression.
Egr-1 in the tissue of patients with (D) immunohistochemical
staining as well as (E) quantification (n=3 each group). (F)
Representative double-labeled immunofluorescence images of (G)
Egr-1 and (H) Egr-1 and CD31 merged quantification (n=3 each
group). (I) Representative double-labeled immunofluorescence images
of (J) Egr-1 and (K) Egr-1 and α-SMA merged quantification (n=3
each group). Each bar represents the mean ± SD. **P<0.01. NS,
not significant. Egr-1, early growth response-1; AVF, arteriovenous
fistula; ROC, receiver operating characteristic; AUC, area under
the curve.

Figure 2

Egr-1 in SMC affects AVF
hemodynamics. (A) AVF mouse model construction and experimental
procedures. (B) Representative images of luminal blood flow size at
14 and 28 days after surgery. (C and D) Lumen patency rate. (E and
F) Quantification of inner diameters of outflow tract lumen (n=5
each group). (G) Representative images of ultrasound flow spectrum.
(H) Linear regression between peak flow velocity and lumen
diameter. (I-L) Quantification of hemodynamic parameters (n=5 each
group). Each bar represents the mean ± SD. *P<0.05 and
**P<0.01. NS, not significant. Egr-1, early growth response-1;
SMC, smooth muscle cell; AVF, arteriovenous fistula.

Figure 3

Egr-1 in SMC promotes AVF neointima
formation. (A) Representative pictures of outflow vessels at the
time of sampling. (B) Quantification of outflow tract lumen
diameter (n=5 each group). (C) Representative hematoxylin and
eosin-stained sections for outflow vessels. (D and E)
Quantification of outflow vessels lumen area and intimal thickness
(n=5 each group). (F) Representative images of immunofluorescence
double labeling for Egr-1 and α-SMA as well as (G-I) quantification
(n=5 each group). Each bar represents the mean ± SD. *P<0.05 and
**P<0.01. NS, not significant. Egr-1, early growth response-1;
SMC, smooth muscle cell; AVF, arteriovenous fistula; αSMA, α-smooth
muscle actin; L, lumen; KO, knockout.

Figure 4

Egr-1 in SMC promotes collagen
deposition and extracellular matrix formation in AVF. (A and B)
Representative images of Sirius red stained sections for outflow
vessels. (C and D) Representative images of Masson stained sections
for outflow vessels. (E) Representative images of
immunohistochemical staining for MMP2, MMP9, PCNA and Tunel. (F-I)
Quantification of the related detection indicators of MMP2, MMP9,
PCNA and Tunel. (n=5 each group). Each bar represents the mean ±
SD. *P<0.05, **P<0.01 and ***P<0.001. NS, not significant.
Egr-1, early growth response-1; SMC, smooth muscle cell; AVF,
arteriovenous fistula; MMP, matrix metallopeptidase; PCNA,
proliferating cell nuclear antigen.

Figure 5

Egr-1 facilitates SMC proliferation
and migration in vitro. (A) Cell monolayers were scratched
and treated with vehicle or shEgr-1 in the presence of PDGF-BB for
24 h. (B) Quantification of the area of wound closure (n=5 each
group). (C) Migrated VSMCs were imaged after being stimulated with
PDGF-BB for 12 h with vehicle or shEgr-1. (D) Quantification of
migrated cells (n=5 each group). (E and F) Representative images of
Edu treated with vehicle or shEgr-1 in the presence of PDGF-BB for
24 h. (F) Quantification of proliferation cells (n=5 each group).
(G) Levels of proteins integral to the proliferation and migration
capacity of Egr-1 knockdown in VSMCs. Each bar represents the mean
± SD. *P<0.05 and **P<0.01. NS, not significant. Egr-1, early
growth response-1; SMC, smooth muscle cell; sh, short hairpin;
PDGF-BB, platelet-derived growth factor-BB; sh, short hairpin;
VSMCs, vascular smooth muscle cells; αSMA, α-smooth muscle
actin.

Figure 6

Joint identification of downstream
targets of Egr-1 by RNA-seq and ChIP-seq. (A) GO pathway enrichment
analyses of the differentially expressed genes. (B) KEGG pathway
enrichment analyses of the differentially expressed genes. (C)
Distribution of different peaks in different regions of the gene.
(D) Distribution diagram and heatmap of the peak near the
transcription start site. (E) To derive the intersecting genes from
RNA-seq and ChIP-seq differential gene sets and construct a Venn
diagram. Egr-1, early growth response-1; RNA-seq, RNA Sequencing;
ChIP-seq, chromatin immunoprecipitation sequencing; GO, Gene
Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Figure 7

Egr-1 regulates smooth muscle
proliferation and migration by transcriptionally activating Egr-2.
(A) Heatmap showing the enrichment of Egr-1 at the TSS of the Egr-2
gene. (B) Quantification of the luciferase reporter assay. (C)
Expression level of Egr-2 in a clinical patient dataset. (D) ROC
curve generated by Egr-2. (E and F) Fluorescence intensity of Egr-2
is markedly upregulated in AVF patient samples. (G) Overexpression
of Egr-2 can mitigate the effects of Egr-1 knockdown in VSMCs. Each
bar represents the mean ± SD. *P<0.05 and **P<0.01. NS, not
significant. Egr-1, early growth response-1; TSS, transcription
start site; VSMCs, vascular smooth muscle cells; AVF, arteriovenous
fistula; ROC, receiver operating characteristic; AUC, area under
the curve; αSMA, α-smooth muscle actin; sh, short hairpin.

Figure 8

Pathophysiological mechanism of Egr-1
in neointimal hyperplasia and lumen stenosis in arteriovenous
fistula. The stimulation of PDGF-BB induces the upregulation of
EGR-1, which subsequently activates the transcription of EGR-2.
This process regulates phenotype switch as well as markers of
proliferation and migration of the vascular smooth muscle cell,
ultimately leading to neointimal hyperplasia and lumen stenosis in
AVF. Egr, early growth response; PDGF-BB, platelet-derived growth
factor-BB; AVF, arteriovenous fistula; αSMA, α-smooth muscle actin;
VSMCs, vascular smooth muscle cells; PDGF-BB, platelet-derived
growth factor-BB.
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Copy and paste a formatted citation
Spandidos Publications style
Hu K, Bu S, Guo Y, Li Y, Yu S, Wang L, Cai C, Li Y, Liu X, Huang H, Huang H, et al: Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas. Int J Mol Med 56: 127, 2025.
APA
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L. ... Wang, W. (2025). Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas. International Journal of Molecular Medicine, 56, 127. https://doi.org/10.3892/ijmm.2025.5568
MLA
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L., Cai, C., Li, Y., Liu, X., Huang, H., Wang, W."Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas". International Journal of Molecular Medicine 56.3 (2025): 127.
Chicago
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L., Cai, C., Li, Y., Liu, X., Huang, H., Wang, W."Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas". International Journal of Molecular Medicine 56, no. 3 (2025): 127. https://doi.org/10.3892/ijmm.2025.5568
Copy and paste a formatted citation
x
Spandidos Publications style
Hu K, Bu S, Guo Y, Li Y, Yu S, Wang L, Cai C, Li Y, Liu X, Huang H, Huang H, et al: Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas. Int J Mol Med 56: 127, 2025.
APA
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L. ... Wang, W. (2025). Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas. International Journal of Molecular Medicine, 56, 127. https://doi.org/10.3892/ijmm.2025.5568
MLA
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L., Cai, C., Li, Y., Liu, X., Huang, H., Wang, W."Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas". International Journal of Molecular Medicine 56.3 (2025): 127.
Chicago
Hu, K., Bu, S., Guo, Y., Li, Y., Yu, S., Wang, L., Cai, C., Li, Y., Liu, X., Huang, H., Wang, W."Egr‑1 promotes the proliferation and migration of vascular smooth muscle cells by transcriptionally activating Egr‑2 in arteriovenous fistulas". International Journal of Molecular Medicine 56, no. 3 (2025): 127. https://doi.org/10.3892/ijmm.2025.5568
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