Stiffness-activated hepatic stellate cells boost HCC migration via TGM2/ITGB1-mediated matrix remodeling and mitochondrial transfer

Abstract

Background & Aims

High liver stiffness correlates with poor outcomes in hepatocellular carcinoma (HCC). Prior studies focused on neoplastic cells rather than the tumor microenvironment. This study investigated how the tumor microenvironment, particularly mechanosignaling in hepatic stellate cells (HSCs), drives HCC progression.

Methods

The study examined the roles of transglutaminase 2 (TGM2) and integrin β1 (ITGB1) in HSCs under mechanical stress through proteomics, cell contraction assays, and protein interactions. It also analyzed gene expression data from 178 patients with HCC and cirrhosis to assess the impact of TGM2 and ITGB1 on overall survival (OS). Mitochondrial transfer and cell migration were observed using confocal microscopy, and the effect of TGM2/ITGB1 on extracellular matrix (ECM) remodeling and HCC recurrence was studied in a rat liver cancer model.

Results

We showed that HSC activation under matrix stiffness relied on ITGB1 mechanosignaling, with high cell-surface TGM2 expression required for ITGB1 activation. This process activated downstream CAV1, which in turn stabilized ITGB1 expression. Moreover, high co-expression of TGM2/ITGB1 (R = 0.77, p <2.2 × 10^-16) was negatively correlated with OS. Interestingly, we found massive mitochondrial transfer in hybrid co-cultures between cancer-associated fibroblasts (CAFs) and Huh7 cells by tunneling nanotubes under high stiffness (p = 0.0095), which appeared to be associated with TGM2/ITGB1. Huh7 cells with CAF-derived mitochondria exhibited enhanced migration under increased substrate stiffness (p <0.0001). Accordingly, high liver stiffness activated CAFs, leading to ECM remodeling and postoperative recurrence of HCC. TGM2/ITGB1 was essential for matrix stiffness-driven HCC recurrence following surgery.

Conclusions

This study revealed a novel mechanism by which HSCs facilitate HCC progression under matrix stiffness, which may aid in the design of therapies for the clinical treatment of HCC.

Impact and Implications

Hepatic stellate cells (HSCs) undergo differentiation into cancer-associated fibroblasts (CAFs), which constitute the primary stromal cell population within the liver tumor microenvironment and are associated with poor prognosis in patients with hepatocellular cancer (HCC). The precise mechanisms through which CAFs facilitate HCC progression remain incompletely elucidated. In this study, we emphasize the role of transglutaminase 2-medated integrin β1 in the activation of HSCs induced by increased matrix stiffness. Furthermore, we explore how enhanced matrix stiffness promotes mitochondrial transfer, thereby facilitating the migration of HCC cells. These insights may inform the development of targeted therapeutic strategies for the clinical management of HCC.