Abstract:

Objective

To investigate the expression level of CCAAT enhancer binding protein ζ antisense strand （CEBPZOS） in hepatocellular carcinoma （HCC） tissues and the mechanism of CEBPZOS in promoting the progression of HCC.

Methods

Based on TCGA and GTEx databases， the correlation between the expression of CEBPZOS and clinical stages and prognosis was analyzed. Hepa1-6 cell lines with CEBPZOS knockdown/overexpression were constructed. The effect of CEBPZOS knockdown/overexpression on the proliferation and migration of Hepa1-6 cells was assessed by CCK-8 assay， colony formation assay，Transwell migration chamber and scratch assay， respectively. Hepa1-6-luc cell lines with CEBPZOS knockdown/overexpression and mouse models with subcutaneous HCC were constructed. The changes of tumor radiation intensity were dynamically monitored by in vivo imaging of small animals. Meantime，the changes in body weight of mice were also recorded. The experimental data between two groups were compared by one-way ANOVA. Survival analysis was performed by Kaplan-Meier method and Log-rank test.

Results

Based on TCGA and GTEx database analyses， the expression level of CEBPZOS mRNA in liver cancer tissues was significantly higher than that in normal tissues adjacent to liver cancer （P＜0.05）. With the progression of tumors （stageⅠ-Ⅲ）， the expression level of CEBPZOS mRNA showed an increasing trend（F=6.41， P＜0.05）. The overall survival and disease-free survival rates in the CEBPZOS overexpression group were significantly lower than those in the CEBPZOS knockdown group （HR=2.00，1.70； both P＜0.05）. CCK-8 and colony formation assays in vitro showed that the proliferation ability of Hepa1-6 cells was significantly decreased after CEBPZOS knockdown， whereas that of Hepa1-6 cells with CEBPZOS overexpression was enhanced （F=148.60， 223.80， both P＜0.05）. Scratch test and Transwell migration chamber revealed that the migration ability of Hepa1-6 cells was decreased after CEBPZOS knockdown， while that of Hepa1-6 cells was increased after CEBPZOS overexpression （F=387.50，80.97； both P＜0.05）. Animal experiments showed that the tumor radiation intensity of mice in the CEBPZOS overexpression group was significantly enhanced over time （F=142.80， P＜0.05）. On the 5th， 9th and 14th d， the tumor radiation intensity was the highest in the CEBPZOS overexpression group， while that in the CEBPZOS knockdown group was the lowest （F=5.24，52.77， 111.00； all P＜0.05）. The body weight of mice in the CEBPZOS overexpression group was significantly increased on the 10th， 12th and 14th d （F=10.27， 28.70， 29.25； all P＜0.05）.

Conclusions

CEBPZOS is highly expressed in HCC， which is closely associated with rapid progression of tumors， high TNM stage and poor prognosis. CEBPZOS can promote the progression of HCC by accelerating the proliferation and migration of tumor cells， which is a potential prognostic biomarker and therapeutic target.

Key words: Carcinoma,hepatocellular, CCAAT enhancer binding protein zeta opposite strand, Transcriptional regulation, Tumor proliferation, Prognosis, Biomarker