SLC10A1 Enhances N'-Nitrosonornicotine-Mediated Regulation of Hepatocellular Carcinoma Cell Proliferation, Migration, and Cisplatin Resistance via PI3K/AKT and MAPK/ERK Signaling Pathways

Bacground: Hepatocellular carcinoma (HCC) is a highly prevalent and malignant form of cancer. SLC10A1, a member of the solute carrier family 10 (SLC10), exhibits lower expression levels in HCC tissues compared to adjacent paracancerous tissues. N'-Nitrosonornicotine (NNN), a well-known carcinogenic component of tobacco, has an undefined role in the context of HCC development and progression. Aims: The primary objectives of our study were to elucidate the regulatory function of SLC10A1 in the proliferation, migration, adhesion, and colony formation of HCC cells, and to identify the underlying signaling pathways involved. Additionally, we sought to determine the impact of SLC10A1 on cisplatin resistance in HCC cells and to explore the role of NNN in the development of HCC. Methods: We utilized HepG2 and HCCLM3 cell lines that stably overexpressed SLC10A1. To assess cell migration, we employed both the scratch wound healing assay and the Transwell migration assay. Cell proliferation was evaluated using the CCK8 assay, while the colony formation ability was also measured with the CCK8 assay. The adhesion of cells to the extracellular matrix was determined through a cell adhesion assay. Furthermore, the effect of SLC10A1 overexpression on the cisplatin resistance of HCC cells was examined via the CCK8 assay. The intracellular signaling pathways regulated by SLC10A1 were investigated using western blot analysis. Finally, we treated HCC cells with NNN at a concentration of 1 mM and monitored cell proliferation. Results: The CCK-8 assay demonstrated that the overexpression of SLC10A1 significantly suppressed the proliferation of both HepG2 and HCCLM3 cells. The wound healing and Transwell assays revealed that SLC10A1 overexpression led to a notable decrease in cell migration capacity. Moreover, the overexpression of SLC10A1 inhibited the adhesion ability of both cell lines to various extracellular matrix components, including type I collagen, fibronectin, and polylysine. Significantly fewer colonies were formed in the SLC10A1 overexpression group compared to the control group. In the cisplatin treatment experiments, the overexpression of SLC10A1 reduced the half maximal inhibitory concentration (IC ₅₀ ) of cisplatin treatment. Additionally, HCC cells treated with NNN exhibited enhanced proliferation capacity. Western blot analysis showed that SLC10A1 overexpression decreased the phosphorylation levels of AKT and ERK, effectively inhibiting the PI3K/AKT and MAPK/ERK signaling pathways. Conclusion: Our in vitro findings indicate that SLC10A1 exerts inhibitory effects on the proliferation, migration, adhesion, and colony formation of HCC cells. These inhibitory effects are mediated by the suppression of the PI3K/AKT and MAPK/ERK signaling pathways. Moreover, the overexpression of SLC10A1 reduces the cisplatin resistance of HCC cells. In contrast, NNN promotes the progression of HCC by enhancing cell proliferation.