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Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion

  • Authors:
    • Hongli Chen
    • Jiao Chen
    • Bomiao Cui
    • Die Lv
    • Wenwen Han
    • Yun Feng
    • Ping Zhang
  • View Affiliations / Copyright

    Affiliations: State Key Laboratory of Oral Diseases and National Center for Stomatology and National Clinical Research Center for Oral Diseases and West China Hospital of Stomatology, Sichuan University, Chengdu, Sichuan 610041, P.R. China
    Copyright: © Chen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 145
    |
    Published online on: August 26, 2025
       https://doi.org/10.3892/or.2025.8978
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Abstract

Oral squamous cell carcinoma (OSCC) ranks as the sixth most prevalent malignancy worldwide, and is characterized by high morbidity and mortality rates. Elucidating the molecular and cellular mechanisms of tumor‑directed immune escape through ecological niche remodeling is crucial for advancing tumor biotherapy. The serum of patients with cancer contains not only tumor biomarkers but also immune regulators secreted by immune cells and/or cancer cells. Notably, the interstitial fluid within the cancer ecological niche is derived from serum. The cross‑talk between serum and cancer cells determines the future of cancer cells, either cell survival or death. The present study revealed that serum from patients with OSCC could remodel the cancer immune escape ecological niche by promoting antigen‑induced regulatory T‑cell (Treg) differentiation and T‑cell exhaustion. When serum from patients with OSCC was added to a phytohemagglutinin‑stimulated peripheral blood mononuclear cell (PBMC) culture system, the Treg subset was significantly increased compared with that in the culture system treated with fetal bovine serum. Moreover, when the serum of patients with OSCC was added to a PBMC culture system stimulated with tumor antigens, the activation of the CD3+ subset was significantly inhibited, and high levels of IL‑4, IL‑10 and TGF‑β were detected in the supernatant; moreover, CD3+ T cells expressed high levels of T‑cell immunoglobulin and mucin‑domain containing‑3 and programmed death ligand 1, which is known to induce T‑cell apoptosis and exhaustion. Finally, the antitumor effect of T cells were significantly decreased. These results indicated that the serum of patients with cancer can promote the inhibition and exhaustion of antitumor T cells, thereby remodeling the tumor immune escape ecological niche.
View Figures

Figure 1

Effects of HTS on T-lymphocyte
differentiation and proliferation. Peripheral blood mononuclear
cells were pre-labeled with CFSE and cultured in medium containing
20% FBS, HTS or HS, with or without 30 µg/ml PHA for 72 h. Cells
were then collected, stained with CD3-APC and analyzed by flow
cytometry using Kaluza 2.1 software. (A) Differentiation of
CD3+ T cell subsets after stimulation with: (Aa) FBS,
HTS or HS without PHA; (Ab) FBS, HTS or HS with PHA; (Ac) FBS +
HTS, FBS + HS or HTS+ HS with PHA. (B) Proliferation of
CD3+ T cells after stimulation with: (Ba) FBS, HTS or HS
without PHA; (Bb) FBS, HTS or HS with PHA; (Bc) FBS + HTS, FBS + HS
or HTS+ HS severally with PHA. (C) Percentage of CD3+
T-cell subsets in each group. (D) Percentage of CFSElow
CD3+ T-cell subsets in each group. The figures are
representative of three independent experiments. Data are presented
as the mean ± SD of each group. *P<0.05. CFSE,
5,6-carboxyfluorescein diacetate succinimidyl ester; FBS, fetal
bovine serum; HS, human healthy serum; HTS, human tumor serum; PHA,
phytohemagglutinin.

Figure 2

Effect of HTS on the proliferation
and differentiation of CD4+ and CD8+ T-cell
subsets. Peripheral blood mononuclear cells were pre-labeled with
CFSE and cultured in medium containing 20% FBS or HTS, and 30 µg/ml
phytohemagglutinin for 72 h. Proliferation and differentiation of
the T-cell subsets were analyzed by flow cytometry using Kaluza 2.1
software. (A) Proliferation of CD4+ T-cell subsets after
stimulation with: (Aa) FBS and (Ab) HTS. (B) Proliferation of
CD8+ T-cell subsets after stimulation with: (Ba) FBS and
(Bb) HTS. (C) Proliferation of CD4+ and CD8+
T-cell subsets. (D) Differentiation of
CD3+CD4+ T-cell subsets after stimulation
with: (Da) FBS and (Db) HTS. (E) Differentiation of
CD3+CD8+ T-cell subsets after stimulation
with: (Ea) FBS and (Eb) HTS. (F) Differentiation of
CD3+CD4+ and CD3+CD8+
T-cell subsets. The figures are representative of three independent
experiments. Data are presented as the mean ± SD for each group.
*P<0.05;. CFSE, 5,6-carboxyfluorescein diacetate succinimidyl
ester; FBS, fetal bovine serum; HTS, human tumor serum.

Figure 3

Effect of HTS on Treg
differentiation. Peripheral blood mononuclear cells were stimulated
with 30 µg/ml PHA and cultured in medium supplemented with FBS or
HTS for 72 h. The CD4+CD25+FOXP3+
Treg subset was analyzed by flow cytometry using Kaluza 2.1
software. (A) FSC and SSC gating to exclude debris, cell aggregates
and dead cells, focusing on single, viable cells. (B) Treg cell
subset in PBMCs stimulated with PHA and FBS. (D) Treg cell subset
in PBMCs stimulated with PHA and HTS. (C) Percentage of Tregs in
PBMCs after stimulation with PHA and different serums. The figures
are representative of three independent experiments. Data are
presented as the mean ± SD for each group. *P<0.05 vs. FBS. Cy5,
Cyanine5; FBS, fetal bovine serum; FOXP3, Forkhead box P3; FSC,
forward scatter; HTS, human tumor serum; PHA, phytohemagglutinin;
SSC, side scatter; Treg, regulatory T cell.

Figure 4

Effects of HTS on T-cell expression
and secretion of immune molecules. Peripheral blood mononuclear
cells were stimulated with tumor cell total protein antigen and
cultured in medium containing FBS or HTS for 14 days. Supernatants
were collected at day 14 to measure the concentrations of IL-4,
IL-10 and TGF-β using ELISA kits. Cells were harvested at day 14,
stained with anti-human CD3-FITC, PD-1-PE, TIM3-PE and CD69-APC,
and the surface markers were analyzed by flow cytometry using
Kaluza 2.1 software. Expression of (A) PD-1, (B) TIM3 and (C) CD69
on T cells. (D) Secretion of immune inhibitory molecules by
HTS-induced T cells. Figures are representative of three
independent experiments. Data are presented as the mean ± SD of
each group. *P<0.05 vs. FBS. FBS, fetal bovine serum; HTS, human
tumor serum; PD1, programmed death ligand 1; TIM3, T-cell
immunoglobulin and mucin-domain containing-3.

Figure 5

Effects of tumor antigen and HTS on T
cell-mediated cancer cell death. PBMCs were stimulated with Cal-27
total protein antigen and cultured in medium containing 20% FBS or
HTS for 14 days. PBMCs were collected and labeled with CFSE and
co-cultured with Cal-27 cells at a ratio of 2:1 in the IncuCyte Rs3
system for 18 h. Cells were then collected and labeled with Annexin
V-APC/PI kits. The Annexin
V+PI+CFSE− cell subset was
analyzed by flow cytometry using Kaluza 2.1 software. (A) Apoptosis
of cancer cells after co-culture with PBMCs. (B) Percentage of
apoptotic cancer cells. (C) T cells were co-cultured with Cal-27
cells in medium containing 20% FBS for 1 and 6 h (yellow arrows
indicate the adherence of T cells to cancer cells). (D) T cells
were co-cultured with Cal-27 cells in medium containing 20% HTS for
1 and 6 h. Figures are representative of three independent
experiments. Data are presented as the mean ± SD of each group.
*P<0.05 vs. FBS. CFSE, 5,6-carboxyfluorescein diacetate
succinimidyl ester; FBS, fetal bovine serum; HTS, human tumor
serum; PBMC, peripheral blood mononuclear cell.
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Copy and paste a formatted citation
Spandidos Publications style
Chen H, Chen J, Cui B, Lv D, Han W, Feng Y and Zhang P: Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion. Oncol Rep 54: 145, 2025.
APA
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., & Zhang, P. (2025). Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion. Oncology Reports, 54, 145. https://doi.org/10.3892/or.2025.8978
MLA
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., Zhang, P."Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion". Oncology Reports 54.5 (2025): 145.
Chicago
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., Zhang, P."Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion". Oncology Reports 54, no. 5 (2025): 145. https://doi.org/10.3892/or.2025.8978
Copy and paste a formatted citation
x
Spandidos Publications style
Chen H, Chen J, Cui B, Lv D, Han W, Feng Y and Zhang P: Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion. Oncol Rep 54: 145, 2025.
APA
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., & Zhang, P. (2025). Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion. Oncology Reports, 54, 145. https://doi.org/10.3892/or.2025.8978
MLA
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., Zhang, P."Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion". Oncology Reports 54.5 (2025): 145.
Chicago
Chen, H., Chen, J., Cui, B., Lv, D., Han, W., Feng, Y., Zhang, P."Serum from patients with oral squamous cell carcinoma remodels the tumor immune escape ecological niche by promoting regulatory T‑cell differentiation and T‑cell exhaustion". Oncology Reports 54, no. 5 (2025): 145. https://doi.org/10.3892/or.2025.8978
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