IBP1 Represses the Migration and Invasion of Hepatocellular Carcinoma Cells by Inhibiting SOCE-dependent Formation of Neutrophil Extracellular Traps by Targeting the Akt Pathway

Background/Aim: Considering its implication in liver disease, we herein aimed to reveal the effects of Insulin-like growth factor-binding protein 1 (IBP1) in hepatocellular carcinoma (HCC).

Materials and Methods: HCC cells with high invasiveness (MHCC97H) and low invasiveness (MHCC97L) were used and transfected with IBP1 overexpression or knockdown plasmids (with reverse-transcription quantitative PCR to validate the efficiency), followed by enzyme-linked immunosorbent assay. The formation of neutrophil extracellular traps (NETs) was confirmed based on Sytox Green and western blot assays. Further, after the co-culture with neutrophils, scratch and Transwell assays were selected to determine the effects of NETs on the migration and invasion of HCC cells under the intervention of IBP1 overexpression and Akt activator SC79. Additionally, western blot was used to test the levels of store-operated calcium entry (SOCE)-relevant factors and matrix metalloproteinase 2/9 (MMP2/9).

Results: IBP1 overexpression reduced HCC cell viability, proliferation, migration, and invasion, whereas IBP1 silencing enhanced these cellular functions. IBP1 overexpression diminished the formation of NETs and downregulated the levels of both citrullinated histone H3 (Cit-H3) and the phosphorylated Akt, whereas silencing it did the opposite. Moreover, IBP1 overexpression diminished the expression of SOCE-relevant protein markers in neutrophils and inhibited the migration and invasion of neutrophil-co-cultured HCC cells, with the decreased levels of MMP2/9. Also, IBP1 overexpression counteracted the effects of HCC cells’ culture media on the aforementioned aspects. However, such effects were all abrogated following the intervention of Akt activator SC79.

Conclusion: IBP1 inhibits the in vitro metastasis of HCC cells by blocking the SOCE-dependent formation of NETs through the Akt pathway.