Single-Cell RNA Sequencing Revealed That the Enrichment of TPI1⁺ Malignant Hepatocytes Was Linked to HCC Metastasis and Immunosuppressive Microenvironment

Abstract

Background

Tumor metastasis is the leading cause of high mortality in hepatocellular carcinoma (HCC). The metastasis-related HCC microenvironment is characterized by high heterogeneity. Single-cell RNA sequencing (scRNA-seq) may aid in determining specific cell clusters involved in regulating the immune microenvironment of HCC.

Methods

The scRNA-seq data of 10 HCC samples were collected from the Gene Expression Omnibus (GEO) database GSE124395. Correlations between key gene expression and clinicopathological data were determined using public databases. HCC tissues and matched tumor-adjacent and normal tissue samples were obtained by surgical resection at Sichuan Cancer Hospital. Immune cell infiltration analysis was performed and verified by immunohistochemistry and immunofluorescent staining.

Results

Nine malignant hepatocyte clusters with different marker genes and biological functions were identified. C3_Hepatocyte-SERF2 and C6_Hepatocyte-IL13RA2 were mainly involved in the regulation of the immune microenvironment, which was also a significant pathway in regulating HCC metastasis. Key genes in malignant hepatocyte clusters that associated with HCC metastasis were further screened by LASSO regression analysis. TPI1, a key gene in C6_Hepatocyte-IL13RA2 and HCC metastasis, could participate in regulating the HCC immune microenvironment in The Cancer Genome Atlas (TCGA) and Tumor Immune Estimation Resource (TIMER) databases. Moreover, immunohistochemistry analysis demonstrated that TPI1 expression was positively correlated with HCC metastasis and poor prognosis, while negatively correlated with CD8⁺ T cell infiltration. The negative correlation between TPI1 expression and CD8⁺ T cell infiltration was further confirmed by immunofluorescence staining.

Conclusion

In summary, a cluster of TPI1⁺ malignant hepatocytes was associated with the suppression of CD8⁺ T cell infiltration and HCC metastasis, providing novel insights into potential biomarkers for immunotherapy in HCC.

Keywords:

• single-cell RNA sequencing
• malignant hepatocyte
• hepatocellular carcinoma
• immune microenvironment
• metastasis

Data Sharing Statement

The data supporting the findings of our study are available from the corresponding author under reasonable requirements.

Ethics Approval and Consent to Participate

The study was approved by the Clinical Research Ethics Committee of Sichuan Cancer Hospital, and written informed consent was received from all participants. All procedures were carried out in accordance with the ethical guidelines of the Helsinki Declaration.

Disclosure

The authors declare no potential conflicts of interest in this work.

Additional information

Funding

This study was supported by Sichuan Province Science and Technology Support Program (No. 2018HH0062), and National Basic Research Program of China (2015CB964703).