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13 August 2019 : Laboratory Research  

Knockdown of the Sonic Hedgehog (SHH) Gene Inhibits Proliferation of Hep3B and SMMC-7721 Hepatocellular Carcinoma Cells via the PI3K/Akt/PCK1 Signaling Pathway

Youping Chen1ABCDE, Weifang Zhu1ABEF*

DOI: 10.12659/MSM.914768

Med Sci Monit 2019; 25:6023-6033

Abstract

BACKGROUND: The PCK1 gene encodes phosphoenolpyruvate carboxykinase (PEPCK), which has been shown have a role in metabolic events in hepatocellular carcinoma (HCC). This study aimed to investigate the role of the SHH gene and its encoded protein, sonic hedgehog (SHH), in two human hepatocellular carcinoma (HCC) cell lines.

MATERIAL AND METHODS: The human HCC cell lines Hep3B and SMMC-7721 were cultured. Cells were transfected with plasmids carrying specific SHH gene short-hairpin RNA (shRNA) and negative control (NC) shRNA. The effects of knockdown of expression levels of the SHH gene were studied on cell survival, cell apoptosis, the cell cycle, gluconeogenesis, and the expression of PCK1. Anchorage-independent growth, a characteristic of transformed cells, was detected by the colony formation assay. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot were performed 24 h after transfection.

RESULTS: Knockdown of expression levels of the SHH gene reduced cell proliferation and growth of HCC cells and induced cell apoptosis and G1 cell cycle arrest in both HCC cell lines. Knockdown of the SHH gene decreased the levels of glycolysis products and increased the production of glucose and reduced the phosphorylation of PI3K and Akt but induced the expression of PCK1.

CONCLUSIONS: Knockdown of the SHH gene reduced cell survival of HCC cells by increasing apoptosis, reducing cell proliferation, inducing G1 cell cycle arrest, and restoring gluconeogenesis, and was associated with the inhibition of the PI3K/Akt axis and induced the expression of PCK1.

Keywords: Hedgehog Proteins, Liver Neoplasms, Phosphatidylinositol 3-Kinases, Phosphoenolpyruvate Carboxykinase (ATP), Apoptosis, Carcinoma, Hepatocellular, Cell Movement, Cell Proliferation, G1 Phase Cell Cycle Checkpoints, Gene Expression Regulation, Neoplastic, Glycolysis, Hep G2 Cells, Intracellular Signaling Peptides and Proteins, Phosphoenolpyruvate Carboxykinase (GTP), Proto-Oncogene Proteins c-akt, Signal Transduction

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750