Analysis of mitotic stress-induced activation of the cGAS/STING signaling pathway in human hepatocytes

Hepatocellular carcinoma (HCC) has been classified as a "genomically unstable" cancer due to the typical presentation of chromosome breakage and aneuploidy. Recent investigations demonstrate that micronuclei, which are formed as a consequence of genotoxic stress, have the capacity to activate innate immune signaling pathways via the cGAS-STING pathway.

The objective of this study was to investigate mitotic stress-induced activation of the cGAS-STING signaling pathway in THLE-5B cells, a SV40 LT immortalized human hepatocyte cell line. In contrast to cancer-derived cell lines these cells are near diploid and express all proteins of the cGAS-STING cascade, rendering them an optimal model for examining this pathway in the context of mitotic stress.

Inactivation of the spindle assembly checkpoint by CFI402257, an inhibitor of the MPS1 kinase, was employed to induce mitotic errors in THLE-5B cells, resulting in the accumulation of aneuploid cells and formation of micronuclei and chromosomal bridges. Confocal microscopy demonstrated the colocalization of cGAS with mitotic abnormalities, and Western blot analysis revealed the activation of the cGAS-STING signaling pathway.

Furthermore, CFI402257-treated THLE-5B cells exhibited the characteristics of a senescence-associated secretory phenotype (SASP) as indicated by SA-β-Gal activity and expression of pro-inflammatory cytokines, including IFN-β, CCL2, and IL-1β.

While SASP enhances immune surveillance to target damaged cells, it may also promote chronic inflammation, which can have adverse effects, including fostering a tumor-promoting environment or accelerating cellular aging and chronic inflammation. The study emphasizes the potential of THLE-5B cells for investigating the cGAS-STING signaling pathway in the context of chromosomal dysregulation in hepatocytes.

Publication History

Article published online:
20 January 2025