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Int J Biol Sci 2022; 18(6):2345-2361. doi:10.7150/ijbs.70137 This issue Cite

Research Paper

MGP promotes CD8⁺ T cell exhaustion by activating the NF-κB pathway leading to liver metastasis of colorectal cancer

Dawei Rong^1,2#, Guangshun Sun^3#, Zhiying Zheng^4#, Li Liu^5,6#, Xiaoyuan Chen^1,2, Fan Wu³, Yichao Gu², Yongjiu Dai², Weizhe Zhong², Xiaopei Hao², Chuanyong Zhang^2✉, Xiongxiong Pan^4✉, Jinhai Tang^7✉, Weiwei Tang^2✉, Xuehao Wang^1,2✉

1. School of Medicine, Southeast University, Nanjing 210000, Jiangsu, P. R. China.
2. Hepatobiliary/Liver Transplantation Center, the First Affiliated Hospital of Nanjing Medical University, Key Laboratory of Living Donor Transplantation, Chinese Academy of Medical Sciences, Nanjing 210000, Jiangsu, P. R. China.
3. Department of General Surgery, Nanjing First Hospital, Nanjing Medical University, Nanjing 210000 Jiangsu, P. R. China.
4. Department of Anesthesiology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China.
5. First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin 301617, P. R. China.
6. State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, P. R. China.
7. Department of General Surgery, The First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China.
#These authors contributed equally to this work.

✉ Corresponding authors: Xuehao Wang, E-mail: wangxhedu.cn; Weiwei Tang, E-mail: 1243773473twwwcom; Jinhai Tang, E-mail: jhtangedu.cn; Xiongxiong Pan, E-mail: xiongxiongedu.cn; Chuanyong Zhang, E-mail: zcy13951673178com.

Abstract

Matrix Gla protein (MGP) was originally reported as a physiological suppressor of ectopia calcification and has also been reported to be associated with cancer. However, the relation between the biological functions of MGP and the immune response in colorectal cancer (CRC) remains unclear. Here, we investigated the regulatory role of MGP in the immune microenvironment of CRC. MGP expression in CRC samples was assessed by single-cell RNA sequencing and the Gene Expression Omnibus (GEO) database, and confirmed by quantitative real-time Polymerase Chain Reaction (qRT-PCR) and immunohistochemistry analysis of human CRC samples. The effect of MGP on proliferation and invasion of CRC cells was evaluated by in vitro assays involving MGP knockdown and overexpression. Luciferase reporter assay and chromatin immunoprecipitation (ChIP)-qPCR assay were performed to identify transcriptional regulatory sites of the nuclear factor kappa-B (NF-κB) and programmed cell death ligand 1 (PD-L1). In vivo experiments were performed in mouse model of CRC liver metastasis established via spleen injection. The results revealed that MGP was significantly upregulated in cancer cell clusters from the primary CRC or liver metastases, compared with that in the corresponding paracancerous tissues via single-cell RNA sequencing. MGP enriched intracellular free Ca²⁺ levels and promoted NF-κB phosphorylation, thereby activated PD-L1 expression to promote CD8⁺ T cell exhaustion in CRC. The luciferase reporter assay and ChIP-qPCR assay indicated that the transcriptional regulation of NF-κB upregulated PD-L1 expression. In vivo, MGP inhibition significantly decreased the rate of CRC liver metastasis, which was further reduced after combined therapy with αPD1 (anti-PD1). In conclusions, this study revealed that MGP can facilitate CD8⁺ T cell exhaustion by activating the NF-κB pathway, leading to liver metastasis of CRC. The combination of MGP knockdown and αPD1 can synergistically resist liver metastasis of CRC.

Keywords: MGP, colorectal cancer, immune escape, liver metastasis, PD-L1

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