Vessels encapsulating tumor clusters promote noninvasive metastasis of hepatocellular carcinoma by shaping an immunosuppressive microenvironment
Bi-Yu Huang, Zheng-Qi Mi, Xiao-Yu Zhang, Yu-Chen Ji, Meng-Zhi Wu, Zi-Feng Cheng, Chen Xie, Shuai He, Jing Zhu, Jian-Hong Fang, Chong Wu, Bin-Kui Li, Yun-Fei Yuan, Limin Zheng, Shi-Mei Zhuang
Bi-Yu Huang, Zheng-Qi Mi, Xiao-Yu Zhang, Yu-Chen Ji, Meng-Zhi Wu, Zi-Feng Cheng, Chen Xie, Shuai He, Jing Zhu, Jian-Hong Fang, Chong Wu, Bin-Kui Li, Yun-Fei Yuan, Limin Zheng, Shi-Mei Zhuang
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Research Article Hepatology Immunology Vascular biology

Vessels encapsulating tumor clusters promote noninvasive metastasis of hepatocellular carcinoma by shaping an immunosuppressive microenvironment

Abstract

Vessels encapsulating tumor clusters (VETC), a distinct vascular pattern in hepatocellular carcinoma (HCC), facilitates noninvasive metastasis in whole clusters. The interaction between VETC and the tumor microenvironment requires exploration. Here, we found that, compared with human non-VETC-HCCs, VETC-tumors exhibited more PD1+CD8+ T cells and Tregs, especially TNFRSF4+ Tregs and Ki67+ Tregs, which showed increased immunosuppressive and proliferative activity. Such immunosuppressive status was also detected in tumor emboli of VETC-HCCs, and Treg density in emboli was positively associated with metastatic cell proliferation. VETC-HCCs revealed abundance correlation, closer spatial proximity, and stronger immunosuppressive ligand-receptor interactions between TNFRSF4+ Tregs/Ki67+ Tregs and PD1+CD8+ T cells. Depleting Tregs in mice reduced PD1+CD8+ T cells in primary lesions, tumor emboli, and metastatic foci of VETC-allografts, and attenuated allograft metastasis. TGF-β1 levels were upregulated in endothelial cells of VETC-HCCs and associated with TNFRSF4+ Tregs/Ki67+ Tregs enrichment. Disrupting VETC formation decreased endothelial TGF-β1 expression and reduced TNFRSF4+ Tregs, Ki67+ Tregs, PD1+CD8+ T cells, and Treg/CD8+ T cell ratios. Collectively, VETC may enhance Treg activity via TGF-β1, while Tregs promote and sustain CD8+ T cell exhaustion through immune inhibitory ligand-receptor interaction, thereby shaping the immunosuppressive microenvironment and enabling tumor clusters to retain such niche to disseminate. These findings disclose mechanisms of tumor immune microenvironment formation and provide rationales for precision medicine.

Authors

Bi-Yu Huang, Zheng-Qi Mi, Xiao-Yu Zhang, Yu-Chen Ji, Meng-Zhi Wu, Zi-Feng Cheng, Chen Xie, Shuai He, Jing Zhu, Jian-Hong Fang, Chong Wu, Bin-Kui Li, Yun-Fei Yuan, Limin Zheng, Shi-Mei Zhuang

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Figure 6

High TGF-β1 expression in TECs is associated with enhanced Treg enrichment and immunosuppressive/proliferative activities in human VETC HCCs.

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High TGF-β1 expression in TECs is associated with enhanced Treg enrichme...
(A) Positive correlation between the VETC index and the ratio of Treg/CD8+ T cells in human VETC HCCs (n = 65). (B–D) Spatial proximity between Treg clusters and TECs. The distances from Tregs (B), TNFRSF4+ Tregs (C) or Ki67+ Tregs (D) to TECs in HCC samples are presented. Scale bars: 50 μm. (E) The score of immunosuppressive genes in TECs. The genes are listed in Supplemental Table 10. The central line of the violin plot denotes median value (50th percentile), the upper and lower lines denote the 75th and 25th percentile, respectively. (F) The relative levels of TGF-β1 in TECs. (G) Positive correlation between the proportion of TNFRSF4+ Tregs (left panel) or Ki67+ Tregs (right panel) and the TGF-β1 levels of TECs (n = 20). (H–K) The distances from Tregs (H and I), TNFRSF4+ Tregs (J), or Ki67+ Tregs (K) to TGF-β1+TEC. non-VETC HCCs, n = 9 (B–D and I–K), 6 (E), 10 (F and G); VETC tumors: n = 65 (A), 12 (B–D), 6 (E), 10 (F and G), 9 (I–K). Multiplex immunofluorescent stainings were performed for (B–D, and F–K), scRNA-seq data were analyzed in E, Scale bars: 50 μm. Data are shown as mean ± SEM. *P < 0.05; **P < 0.01; ****P < 0.0001, by Pearson’s correlation test (A and G), Mann-Whitney U test (B–E), 2-tailed Student’s t test (F and I–K).