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Lowered HGK expression inhibits cell invasion and adhesion in hepatocellular carcinoma cell line HepG2
Su-Xia Han1, Qing Zhu, Jin-Lu Ma
1Oncology Center of the First Affiliated Hospital, College of Medicine, Xi'an Jiaotong University, Xi'an 710061, Shaanxi Province, China. hsummer22099@yahoo.cn
Aim:
To investigate the effects of RNA interference targeting hepatocyte progenitor kinase-like kinase (HGK) in the invasion and adhesion of hepatocellular carcinoma (HCC) cell line HepG2.
Methods:
Three paired insert DNA fragments specific to HGK gene and one negative control DNA fragment were synthesized and inserted into RNAi-Ready pSIREN-RetroQ-ZsGreen vector. Western blotting assay and real-time reverse transcriptase polymerase chain reaction (RT-PCR) were used to screen the vector with a highest inhibitory rate. The vector was used to generate recombinant retrovirus specific to HGK. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2h-tetrazolium bromide (MTT) assay was used to examine cell growth; wound closure assay and cell adhesion assay were employed to investigate cell migration and adhesion respectively; and transwell assay and three-dimensional culture invasion assay were used to detect cell invasion. The expressions of matrix metalloproteinase (MMP)-2, MMP-9 and nuclear factor (NF)-κB were detected by Western blotting assay.
Results:
The real time RT-PCR and Western blotting assay showed that cells transfected with retrovirus mediating RNAi targeting of HGK (RV-shHGK)-1 vector had the strongest inhibition of HGK protein, with an inhibition rate of 76%, and this vector was used to generate recombinant retrovirus RV-shHGK-1. Cell adhesion assay and MTT assay found that cell adhesion and growth of the cells infected with RV-shHGK-1 were significantly lower than those of the control cells (P < 0.05). Wound closure assay, transwell assay and three-dimensional culture invasion assay showed that the cell invasiveness was significantly less in HGK knockdown cells than in the control cells (P < 0.05). The expressions of MMP-2, MMP-9 and NF-κB were inhibited in HepG2 cells infected with RV-shHGK-1.
Conclusion:
Down-regulation of HGK can obviously inhibit the migration and invasion of HepG2 cells in vitro. HGK may be a new therapeutic target for treatment of HCC.
Insights
Reducing hepatocyte progenitor kinase-like kinase (HGK) with RNA interference significantly inhibits hepatocellular carcinoma (HCC) cell invasion and migration. This suggests HGK is a potential therapeutic target for HCC treatment.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Hepatocellular carcinoma (HCC) is a prevalent malignancy with aggressive invasion and metastasis.
- Identifying novel therapeutic targets is crucial for improving HCC patient outcomes.
- Hepatocyte progenitor kinase-like kinase (HGK) role in HCC progression requires further elucidation.
Purpose of the Study:
- To investigate the therapeutic potential of targeting HGK in HCC.
- To evaluate the effect of HGK down-regulation on HepG2 cell invasion and adhesion.
- To explore the underlying molecular mechanisms involving MMP-2, MMP-9, and NF-κB.
Main Methods:
- RNA interference (RNAi) targeting HGK was employed using a lentiviral vector (RV-shHGK-1).
- Quantitative assessment of cell invasion, migration, adhesion, and proliferation (MTT assay, wound closure, transwell, 3D culture).
- Western blotting was used to measure protein expression levels of HGK, MMP-2, MMP-9, and NF-κB.
Main Results:
- RV-shHGK-1 effectively inhibited HGK expression by 76% in HepG2 cells.
- HGK knockdown significantly reduced HepG2 cell adhesion, growth, migration, and invasion (P < 0.05).
- Expressions of MMP-2, MMP-9, and NF-κB were significantly decreased in HGK-silenced cells.
Conclusions:
- Down-regulation of HGK inhibits migration and invasion of HCC cells in vitro.
- HGK represents a promising novel therapeutic target for HCC treatment.
- Targeting HGK may offer a new strategy for controlling HCC metastasis.
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