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Article Open Access

Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway

  • Authors:
    • Cui-Cui Gong
    • Hua-Kang Li
    • Yuan-Zhen Mi
    • Jun-Yang Chen
    • Zeng-Yi Fang
    • Shun-Lian Fu
    • Li Quan
    • Bing Lin
    • Jin-Yi Lang
    • Qiu Chen
    • Ke Xu
    • Mei-Hua Chen
  • View Affiliations / Copyright

    Affiliations: Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan 610032, P.R. China, Department of Radiation Oncology, Radiation Oncology Key Laboratory of Sichuan Province, Sichuan Clinical Research Center for Cancer, Sichuan Cancer Hospital and Institute, Sichuan Cancer Center, Affiliated Cancer Hospital of University of Electronic Science and Technology of China, Chengdu, Sichuan 610041, P.R. China, School of Medicine, University of Electronic Science and Technology of China, Chengdu, Sichuan 611731, P.R. China, Department of Endocrine, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan 610075, P.R. China
    Copyright: © Gong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 69
    |
    Published online on: January 21, 2026
       https://doi.org/10.3892/ijmm.2026.5740
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Abstract

Radiation‑induced lung injury (RILI) remains a dose‑limiting and life‑threatening complication of thoracic radiotherapy. The present study aimed to evaluate the therapeutic efficacy and mechanism of the naturally extracted flavonoid, 5,7,8‑trimethoxyflavone (HY‑N7656), in inhibiting RILI. Lung injury in mice was evaluated using micro‑computed tomography, histopathological analysis, enzyme‑linked immunosorbent assay and western blotting. Network pharmacology was conducted to predict the potential therapeutic targets and signaling pathways of HY‑N7656 in RILI. Cell Counting Kit‑8, wound healing, immunofluorescence, reverse transcription‑quantitative (RT‑q) PCR and protein expression analyses were carried out in vitro using TGF‑β‑stimulated A549 cells to evaluate epithelial‑mesenchymal transition (EMT) and signaling activity. Results of the present study revealed that HY‑N7656 markedly alleviated pulmonary inflammation and fibrosis in irradiated mice, leading to a reduction in α‑smooth muscle actin expression. In addition, EMT was effectively reversed following treatment with HY‑N7656 in A549 alveolar epithelial cells treated with TGF‑β, accompanied by restoration of E‑cadherin expression and downregulation of mesenchymal markers, such as N‑cadherin and vimentin. Network pharmacology analysis and molecular docking validation identified the PI3K/Akt pathway as a central target, which was subsequently confirmed via western blot analysis. Moreover, results of the present study demonstrated that HY‑N7656 inhibited radiation‑induced activation of PI3K and Akt. To the best of the authors' knowledge, the present study was the first to demonstrate that HY‑N7656 modulates the PI3K/Akt signaling pathway to suppress the progression of EMT in RILI, establishing HY‑N7656 as a multi‑target inhibitor of RILI. These findings present a potential strategy to enhance the safety of radiotherapy, warranting further preclinical and clinical evaluation.

View Figures

Figure 1

Effect of HY-N7656 on lung injury in
an IR-induced model. (A) Micro-CT images and H&E-stained lung
tissue sections from mice in the Control, IR and HY-N7656 treatment
groups (7.5, 15 and 30 mg/kg). Magnification, ×100. Scale bar, 200
μm. (B) Quantification of lung density (HU) based on
micro-CT imaging demonstrated a significant decrease in lung
density in the IR group relative to the control group, with
HY-N7656 producing a dose-dependent improvement. (C)
Histopathological scoring (Szapiel score) demonstrated a
significant reduction in tissue damage in HY-N7656-treated mice
compared with the IR group. Data are presented as mean ± SEM (n=5).
*P<0.05, **P<0.01,
***P<0.001, ****P<0.0001; ns, no
significance. HY-N7656, 5,7,8-trimethoxyflavone; IR, ionizing
radiation; HU, Hounsfield unit; micro-CT, micro-computed
tomography; H&E, haematoxylin and eosin; SEM, standard error of
the mean.

Figure 2

Effects of HY-N7656 on lung
morphology and histopathological changes in irradiated mice. (A)
Experimental design showing the timeline of radiation therapy,
HY-N7656 administration and evaluation at Weeks 4, 8, 12 and 16
through micro-CT scan, H&E staining, Masson staining and IHC
analysis. (B) Representative micro-CT images of lung morphology at
4, 8, 12 and 16 weeks post-treatment. Radiation-induced lung injury
is indicated by arrows. (C) Representative histological analysis of
lung tissue at 16 weeks. H&E, Masson (magnification, ×100;
scale bar, 200 μm) and α-SMA staining (magnification, ×200;
scale bar, 100 μm) revealed progressive fibrosis in the IR
group, while HY-N7656 treatment reduced fibrosis and α-SMA
expression, indicating a protective effect. (D) Quantification of
lung density (HU) based on micro-CT imaging at each time point
showed significant improvement in lung density in HY-N7656-treated
groups compared with the IR group. (E) Szapiel score for H&E
staining in lung tissue sections. (F) Ashcroft score for Masson
staining in lung tissue sections. (G) Quantification of
α-SMA-positive area (%) in lung tissue. Data are presented as mean
± SEM (n=5), *P<0.05, **P<0.01,
***P<0.001, ****P<0.0001, ns, no
significance. HY-N7656, 5,7,8-trimethoxyflavone; IR, ionizing
radiation; HU, Hounsfield unit; micro-CT, micro-computed
tomography; H&E, hematoxylin and eosin; IHC,
immunohistochemistry; α-SMA, α-smooth muscle actin; SEM, standard
error of the mean.

Figure 3

HY-N7656-mediated inhibition of RILI
and associated mechanisms in irradiated mice. (A) Detection of
serum TGF-β levels using ELISA at 16 weeks post-irradiation in each
group of mice (n=5). (B) Statistical analysis of relative lung
coefficient in each group of mice (n=5). (C) Quantification of
hydroxyproline content in lung tissues at 16 weeks post-radiation
(n=5). (D) Western blot analysis of EMT markers in RILI mice at 16
weeks post-irradiation. Quantification of (E) E-cadherin and (F)
vimentin protein expression levels. Data are presented as mean ±
SEM, *P<0.05, **P<0.01,
***P<0.001, ****P<0.0001, ns: no
significance. HY-N7656, 5,7,8-trimethoxyflavone; RILI,
radiation-induced lung injury; TGF-β, transforming growth factor-β;
ELISA, enzyme-linked immunosorbent assay; EMT,
epithelial-mesenchymal transition; SEM, standard error of the mean;
ns, not significant.

Figure 4

The effect of HY-N7656 on cell
migration and EMT markers. (A) Representative images of wound
healing assays highlighting cell migration at 0 and 48 h after
treatment in control, HY-N7656, TGF-β and TGF-β + HY-N7656 groups.
The red lines indicate the wound area (scale bar, 500 μm).
(B) Quantification of cell migration represented as fold change
relative to control. (C) Immunofluorescence staining of E-cadherin
(red) and DAPI (blue) in A549 cells (scale bar, 20 μm). (D)
Immunofluorescence staining of vimentin (green) and DAPI (blue) in
A549 cells (scale bar, 20 μm). (E) Western blot analysis of
N-cadherin, E-cadherin and vimentin protein expression levels in
cells. GAPDH was used as a loading control. Quantification of (F)
E-cadherin, (G) N-cadherin and (H) vimentin protein expression.
Data are presented as mean ± SEM (n=3), *P<0.05,
**P<0.01, ***P<0.001,
****P<0.0001, ns, not significant. HY-N7656,
5,7,8-trimethoxyflavone; EMT, epithelial-mesenchymal transition;
TGF-β, transforming growth factor-β; DAPI,
4',6-diamidino-2-phenylindole; GAPDH, glyceraldehyde 3-phosphate
dehydrogenase; SEM, standard error of the mean.

Figure 5

Network pharmacology analysis. (A)
After intersecting 118 targets of HY-N7656 with 1,712 RILI-related
targets, 58 common targets of HY-N7656 acting on RILI were
identified. (B) PPI network diagram of the 58 intersecting targets,
with node importance decreasing from large to small and color
transitioning from yellow to green. (C) GO enrichment of HY-N7656
targets for the treatment of RILI, highlighting the top 10 enriched
BPs, CCs and MFs. (D) Top 20 KEGG pathways enriched for HY-N7656 in
the treatment of RILI. (E) Visualization of molecular docking
between HY-N7656 and PI3K. (F) Visualization of molecular docking
between HY-N7656 and Akt. HY-N7656, 5,7,8-trimethoxyflavone; RILI,
radiation-induced lung injury; PPI, protein-protein interaction;
GO, Gene Ontology; BP, biological process; CC, cellular component;
MF, molecular function; KEGG, Kyoto Encyclopedia of Genes and
Genomes; PI3K, phosphoinositide 3-kinase.

Figure 6

HY-N7656 inhibits activation of the
PI3K/Akt signaling pathway in A549 cells. (A) Western blot analysis
of PI3K, AKT, PI3K phosphorylation and AKT phosphorylation in A549
cells. Quantitative analysis of (B) p-Akt and (C) p-PI3K protein
levels. GAPDH was used as the loading control. Data are presented
as mean ± SEM. *P<0.05, ***P<0.001. (D)
Mechanism of action for HY-N7656 in alleviating RILI via inhibition
of PI3K/Akt mediated partial EMT. HY-N7656,
5,7,8-trimethoxyflavone; PI3K, phosphoinositide 3-kinase; AKT,
protein kinase B; RILI, radiation-induced lung injury; EMT,
epithelial-mesenchymal transition; GAPDH, glyceraldehyde
3-phosphate dehydrogenase; SEM, standard error of the mean; p-,
phosphorylated.
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Copy and paste a formatted citation
Spandidos Publications style
Gong C, Li H, Mi Y, Chen J, Fang Z, Fu S, Quan L, Lin B, Lang J, Chen Q, Chen Q, et al: <p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>. Int J Mol Med 57: 69, 2026.
APA
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S. ... Chen, M. (2026). <p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>. International Journal of Molecular Medicine, 57, 69. https://doi.org/10.3892/ijmm.2026.5740
MLA
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S., Quan, L., Lin, B., Lang, J., Chen, Q., Xu, K., Chen, M."<p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>". International Journal of Molecular Medicine 57.3 (2026): 69.
Chicago
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S., Quan, L., Lin, B., Lang, J., Chen, Q., Xu, K., Chen, M."<p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>". International Journal of Molecular Medicine 57, no. 3 (2026): 69. https://doi.org/10.3892/ijmm.2026.5740
Copy and paste a formatted citation
x
Spandidos Publications style
Gong C, Li H, Mi Y, Chen J, Fang Z, Fu S, Quan L, Lin B, Lang J, Chen Q, Chen Q, et al: <p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>. Int J Mol Med 57: 69, 2026.
APA
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S. ... Chen, M. (2026). <p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>. International Journal of Molecular Medicine, 57, 69. https://doi.org/10.3892/ijmm.2026.5740
MLA
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S., Quan, L., Lin, B., Lang, J., Chen, Q., Xu, K., Chen, M."<p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>". International Journal of Molecular Medicine 57.3 (2026): 69.
Chicago
Gong, C., Li, H., Mi, Y., Chen, J., Fang, Z., Fu, S., Quan, L., Lin, B., Lang, J., Chen, Q., Xu, K., Chen, M."<p>Natural compound 5,7,8‑trimethoxyflavone mitigates radiation‑induced lung injury by suppressing EMT and PI3K/Akt pathway</p>". International Journal of Molecular Medicine 57, no. 3 (2026): 69. https://doi.org/10.3892/ijmm.2026.5740
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