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Puerariae radix flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate

  • Authors:
    • Jing Dong
    • Yupeng Guo
    • Tao Ji
    • Hongjun Guan
    • Yanbin Ma
    • Shengzhong Rong
  • View Affiliations / Copyright

    Affiliations: College of Public Health, Mudanjiang Medical University, Mudanjiang, Heilongjiang 157011, P.R. China, College of Science, Harbin University of Science and Technology, Harbin, Heilongjiang 150080, P.R. China
    Copyright: © Dong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 170
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    Published online on: August 27, 2025
       https://doi.org/10.3892/br.2025.2048
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Abstract

Puerariae radix flavones (PRFs), bioactive components derived from the Pueraria lobata plant, exhibit anti‑inflammatory and anti‑tumor properties. However, their therapeutic potential for bladder cancer remains poorly understood. The present study aimed to investigate the anti‑tumor effects and molecular mechanisms underlying the effects of PRF on human bladder cancer T24 cells. In vitro experiments were performed out in PRF‑treated T24 cells, and the effects of PRF were assessed through MTT assay, acridine orange/ethidium bromide staining, agarose gel electrophoresis, reverse transcription‑quantitative PCR and ELISA. PRF significantly inhibited T24 cell proliferation via inducing apoptosis through the extrinsic apoptosis pathway. Mechanistically, PRF induced FAS receptor/tumor necrosis factor‑α receptor, enhanced the activity of cysteinyl aspartate‑specific protease‑3 and downregulated NF‑κB. To the best of our knowledge, the present study is the first to demonstrate that PRF could suppress the proliferation of human bladder cancer T24 cells and to elucidate its underlying molecular mechanism. These findings may provide novel insights into PRF as a promising therapeutic agent for bladder cancer treatment.
View Figures

Figure 1

Inhibitory effect of PRF on bladder
cancer T24 cells. (A) Inhibitory rate of each PRF treatment group
on cell viability at each time point. *P<0.05. (B) At
each time point, the cell viability inhibition rate exhibited a
concentration-dependent increase with PRF. PRF, Puerariae
radix flavone.

Figure 2

Acridine orange/ethidium bromide
staining of T24 cells exposed PRF. (A) Control for 24 h, the yellow
arrow indicates viable T24 cells exhibiting green fluorescence with
intact plasma membranes. (B) 50 µg/ml and 24 h, yellow arrow
indicates early apoptotic cells demonstrating green fluorescence
with punctate orange-red fluorescence, along with characteristic
morphological changes including cell shrinkage, nuclear
condensation, and apoptotic body formation. (C) 100 µg/ml and 24 h,
yellow arrow indicates apoptotic cells in early and late stages,
exhibiting green fluorescence accompanied by orange-red
fluorescence. Early-stage cells show apoptotic body formation,
while late-stage cells display plasma membrane disintegration,
chromatin fragmentation, and diffuse nuclear orange-red
fluorescence. (D) 200 µg/ml and 24 h, the yellow arrow indicates
late apoptotic cells exhibiting orange-red fluorescence,
characterized by chromatin fragmentation and nuclear
disintegration. (E) Control and 48 h, the yellow arrow indicates
viable T24 cells exhibiting green fluorescence with intact plasma
membranes. (F) 50 µg/ml and 48 h, yellow arrow indicates early
apoptotic cells demonstrating green fluorescence with punctate
orange-red fluorescence, along with characteristic morphological
changes including cell shrinkage, nuclear condensation, and
apoptotic body formation. (G) 100 µg/ml and 24 h, yellow arrow
indicates apoptotic cells in early and late stages, exhibiting
green fluorescence accompanied by orange-red fluorescence.
Early-stage cells show apoptotic body formation, while late-stage
cells display plasma membrane disintegration, chromatin
fragmentation and diffuse nuclear orange-red fluorescence. (H) 200
µg/ml and 48 h, the yellow arrow indicates late apoptotic cells
exhibiting orange-red fluorescence, characterized by chromatin
fragmentation and nuclear disintegration. Magnification, x200. PRF,
Puerariae radix flavone.

Figure 3

Fragmentation of genomic DNA in T24
cells treated with PRF at concentrations of 0, 50, 100, and 200
µg/ml for 24 or 48 h. Compared with the 0 µg/ml PRF group, no
significant changes were observed in the other treatment groups,
and no characteristic DNA ladder was detected. PRF, Puerariae
radix flavone.

Figure 4

Effects of PRF on the expression of
apoptosis-related genes in T24 cells. T24 cells were treated with
PRF (0, 50, 100 and 200 µg/ml) for 24 or 48 h. The mRNA levels of
(A) FAS, (B) FASL, (C) TNFα, (D) TNFR1, (E) CASP3, (F) NF-κB, (G)
BCL2 and (H) BAX were assessed by quantitative PCR (normalized to
GAPDH). *P<0.05. PRF, Puerariae radix
flavones; CASP3, cysteinyl aspartate-specific protease-3.

Figure 5

PRF regulates intracellular and
secreted levels of FAS and TNF-α in T24 cells. T24 cells were
exposed to PRF (0, 50, 100 or 200 µg/ml) for 24 or 48 h. (A) FAS
levels in cell lysate (intracellular proteins). (B) Secreted FAS
levels in conditioned medium. (C) TNF-α levels in cell lysates
(intracellular proteins). (D) Secreted TNF-α levels in conditioned
medium. Protein quantification was performed by ELISA.
*P<0.05. PRF, Puerariae radix flavone.

Figure 6

Mechanism of apoptosis. Apoptotic
signaling primarily occurs via two pathways: the extrinsic death
receptor pathway and the intrinsic mitochondrial pathway. In T24
cells, PRF induces apoptosis predominantly by activating FAS and
TNFR1, thereby initiating the death receptor pathway. This
ultimately leads to the activation of CASPASE3, resulting in
characteristic biochemical and morphological changes characteristic
of programmed cell death. Concurrently, NF-κB participates in the
regulation of this apoptotic pathway. TNFR, tumor necrosis factor
receptor; TRAF, TNFR-Associated Factor; TRADD, TNFR-associated
death domain; RIP, receptor-interacting protein; IKK, IκB kinase;
tBid, truncated Bid; PRF, Puerariae radix flavone.
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Copy and paste a formatted citation
Spandidos Publications style
Dong J, Guo Y, Ji T, Guan H, Ma Y and Rong S: <em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate. Biomed Rep 23: 170, 2025.
APA
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., & Rong, S. (2025). <em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate. Biomedical Reports, 23, 170. https://doi.org/10.3892/br.2025.2048
MLA
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., Rong, S."<em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate". Biomedical Reports 23.5 (2025): 170.
Chicago
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., Rong, S."<em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate". Biomedical Reports 23, no. 5 (2025): 170. https://doi.org/10.3892/br.2025.2048
Copy and paste a formatted citation
x
Spandidos Publications style
Dong J, Guo Y, Ji T, Guan H, Ma Y and Rong S: <em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate. Biomed Rep 23: 170, 2025.
APA
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., & Rong, S. (2025). <em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate. Biomedical Reports, 23, 170. https://doi.org/10.3892/br.2025.2048
MLA
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., Rong, S."<em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate". Biomedical Reports 23.5 (2025): 170.
Chicago
Dong, J., Guo, Y., Ji, T., Guan, H., Ma, Y., Rong, S."<em>Puerariae radix</em> flavones induce apoptosis in bladder cancer T24 cells via the FAS/TNFR1 pathway: A potential therapeutic candidate". Biomedical Reports 23, no. 5 (2025): 170. https://doi.org/10.3892/br.2025.2048
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