PLA2G4A promotes right-sided colorectal cancer progression by inducing CD39+γδ Treg polarization
Yang Zhan, Lei Zheng, Jia Liu, Dongzhi Hu, Junfeng Wang, Kai Liu, Jiansheng Guo, Ti Zhang, Dalu Kong
Yang Zhan, Lei Zheng, Jia Liu, Dongzhi Hu, Junfeng Wang, Kai Liu, Jiansheng Guo, Ti Zhang, Dalu Kong
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Research Article Oncology

PLA2G4A promotes right-sided colorectal cancer progression by inducing CD39+γδ Treg polarization

Abstract

The γδ T cell is a promising candidate cell in tumor immunotherapy. However, γδ T cells polarize to CD39+γδ Tregs upon colorectal cancer (CRC) induction, and the underlying mechanism remains unclear. Here, we show that the frequency of CD39+γδ Tregs, which positively correlated with poor prognosis, was significantly higher in right-sided CRC (RSCRC) than in the left-sided CRC (LSCRC). Interestingly, CD39+γδ Tregs from RSCRC showed stronger immunosuppressive phenotype and function than LSCRC. Furthermore, the quantitative mass spectrometry data show that CD39+γδ Treg polarization was related to the abnormal activation of the Phospholipase a2-IVa/Arachidonic acid (PLA2G4A/AA) metabolic pathway in RSCRC. Using an in vitro coculture system and an orthotopic murine model of CRC, we show that the overexpression of Pla2g4a in CT26 cells induced CD39+γδ Tregs, inhibiting the antitumor immune response. Finally, we found that the overall survival of the PLA2G4Ahi group was significantly shortened compared with PLA2G4Alo RSCRC, while the survival of LSCRC showed the opposite. Collectively, RSCRC with abnormal PLA2G4A expression educates γδ T cells into CD39+γδ Tregs to promote tumor progression and metastasis. Our work highlights the interaction between cancer cells and immune cells by distinguishing the primary tumor site and deepens the understanding of the tumor microenvironment and immunosuppression.

Authors

Yang Zhan, Lei Zheng, Jia Liu, Dongzhi Hu, Junfeng Wang, Kai Liu, Jiansheng Guo, Ti Zhang, Dalu Kong

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Figure 5

Overexpression of Pla2g4a in CT26-induced CD39+γδ Tregs in vitro.

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Overexpression of Pla2g4a in CT26-induced CD39+γδ Tregs in vitro. CT26-P...
CT26-Pla2g4a and CT26-Vec cells were cocultured with intraepithelial lymphocytes derived from the ileum (IL-IELs), colon (CO-IELs), or spleen of BALB/c mice. (A and C) Representative pictures showed the number of CT26 cells in the coculture system after 12 (A) and 36 hours (C). Scale bars: 200 μm. (B and D) Bar diagrams summarizes the cell numbers of CT26-Pla2g4a and CT26-Vec cells cocultured with IL-IELs after 12 (B) and 36 (D) hours. Data are shown as mean ± SEM, using 2-way ANOVA followed by Sidak’s multiple-comparisons test. (E–G) Representative histogram plot and bar diagram summarizes the expression of CD39 on γδ T cells in IL-IELs (E), CO-IELs (F) and splenocytes (G) group. (H) Representative flow cytometric analysis of CD39+ Tregs in the total CD3+ T cells from splenocytes. Bar diagram summarizes the percentages of CD39+γδ Tregs. Unpaired 2-tailed t test was used to compare variables. (I) Silencing Pla2g4a in CT26-Pla2g4a rescued the antitumor response mediated by γδ T cells. CT26-Pla2g4a were transfected with Pla2g4a siRNA or control siRNA. After 24 hours, CT26-Vec, CT26-Pla2g4a, and CT26-Pla2g4a–knock-down cells were cocultured with γδ T cells sorted from BALB/c mice with or without anti-PLA2G4A antibody (5 mg/mL). Scale bars: 200 μm. (J) Bar diagram summarizes the cell number of CT26. (K) Representative histogram plot of CD39 expression on γδ T cells in each group by flow cytometry. Bar diagram summarizes the frequency of CD39+ γδ Tregs/γδ T cells. One-way ANOVA followed by Tukey’s multiple-comparison test was used for multiple-group analysis. All of the experiments were repeated 3 times; n = 3/group. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.