[Preliminary study on the biological role of EF-hand domain-containing protein 2 in hepatocellular carcinoma].

This study investigates the expression pattern and functional significance of EF-hand domain-containing protein 2 (EFHD2) in hepatocellular carcinoma (HCC), with particular focus on its regulatory effects on tumor proliferation, migration, and invasion. Cellular experimental study was completed from June 2024 to January 2025 in the Basic Laboratory of the General Hospital of Southern Theater Command. TCGA database to determine EFHD2 expression and its clinicopathological correlations. GSCA database to assess methylation patterns and immune infiltration. Model of transient overexpression and knockdown of EFHD2 was constructed in hepatocellular carcinoma cells Hep3B, then RT-qPCR and Western blot were applied to verify the transfection efficiency. CCK-8 and colony formation assays for proliferation assessment, Transwell chambers for migration/invasion quantification. Protein-protein interaction networks were constructed via STRING, followed by GO/KEGG enrichment analysis. Statistical analysis was performed using the two independent samples -test. The results showed that EFHD2 demonstrated significant upregulation in HCC tissues versus normal controls (<0.05). Elevated EFHD2 expression correlated with advanced clinical stage (<0.05) and poor differentiation (<0.05). In the CCK-8 assay, the EFHD2 overexpression group demonstrated significantly higher cell viability than the control group, as evidenced by 450 nm relative absorbance values on Day 1 (0.529±0.019 0.515±0.016, =0.041, =0.320), Day 2 (1.356±0.019 1.094±0.042, =3.833, <0.001), Day 3 (2.817±0.049 2.143±0.124, =3.833, <0.001), and Day 4 (3.848±0.015 3.430±0.021, =0.469, <0.001). The EFHD2 knockdown group showed reduced cell viability compared to controls: Day 1 (0.541±0.020 0.552±0.015, =0.098, =0.423), Day 2 (1.154±0.009 1.326±0.029, =2.485, <0.001), Day 3 (2.453±0.041 2.653±0.031, =0.479, <0.001), and Day 4 (3.685±0.038 3.836±0.021, =6.804, <0.001). In colony formation assays, the overexpression group displayed a significant increase in colony numbers (254.667±23.861 186.000±16.703, =0.865, =0.015), whereas the knockdown group exhibited decreased colony formation (229.000±24.637 306.667±36.501, =0.988, =0.038). In Transwell assays, the EFHD2 overexpression group revealed enhanced migratory capacity [ (605.000±72.670) cells (472.667±28.095) cells, =2.462, =0.042] and invasive potential [(767.333±21.221) cells (414.333±16.623) cells, =0.331, <0.001]. The knockdown group showed attenuated migration [(311.000±71.084) cells (479.667±50.846) cells, =0.718, =0.029] and invasion [(247.667±48.263) cells (345.667±32.130) cells, =0.727, =0.043] compared to controls. The network of EFHD2-interacting proteins was further constructed by the STRING database, and the GO and KEGG analysis were used to perform bioinformatics analysis reveal that EFHD2 is mainly involved in actin cytoskeleton regulation. In conclusion, EFHD2 is highly expressed in HCC and is involved in the process of proliferation, migration and invasion of HCC.