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Effect of retinoblastoma gene on human osteosarcoma cell line OS732
1Department of Pathology, Sun Yat-sen University of Medical Sciences, Guangzhou 510080, China (Email: qiaohui99@163.net)
Objective:
To investigate the effect of growth suppression and gap junctional intercellular communication (GJIC) on osteosarcoma cell line OS732 transduced with the exogenous N-terminal truncated retinoblastoma (Rb) gene.
Methods:
Recombinant eukaryotic expression plasmid harbouring 1.65 kb Rb gene was constructed and transduced into OS732 cells. Rb mRNA expression was detected by RT-PCR and Northern blot. The cell count, cell cycle analysis and soft agar colony formation test were used to observe growth features. RT-PCR and Lucifer yellow dye transfer were carried out to measure GJIC.
Results:
Both exogenous and endogenous Rb gene could be detected in OS732 cell transduced with the N-terminal truncated Rb gene. Compared with the parental cells, change in morphology of Rb-transduced cells were found; growth rate decreased; amount of colony formation reduced; cell cycle arrested at G(0)/G(1) checkpoint; Connexin43 expression and GJIC enhanced.
Conclusions:
The malignant phenotype of OS732 is inhibited and GJIC is increased by transduction of N-terminal truncated Rb gene.
Insights
Introducing the retinoblastoma (Rb) gene into osteosarcoma cells suppressed growth and enhanced gap junctional intercellular communication (GJIC). This study highlights Rb gene therapy potential for osteosarcoma treatment.
Area of Science:
- Oncology
- Molecular Biology
- Cell Biology
Background:
- Osteosarcoma is a primary bone malignancy with limited treatment options.
- The retinoblastoma (Rb) gene plays a crucial role in cell cycle regulation.
- Gap junctional intercellular communication (GJIC) is vital for cellular coordination and tissue homeostasis.
Purpose of the Study:
- To investigate the impact of introducing an N-terminal truncated retinoblastoma (Rb) gene into the OS732 osteosarcoma cell line.
- To assess the effects on cell growth and GJIC.
Main Methods:
- Constructed a recombinant plasmid with the 1.65 kb Rb gene for transduction into OS732 cells.
- Utilized RT-PCR and Northern blot to confirm Rb mRNA expression.
- Assessed cell growth via cell count, cell cycle analysis, and soft agar colony formation.
- Measured GJIC using RT-PCR and Lucifer yellow dye transfer.
Main Results:
- Transduced OS732 cells expressed both exogenous and endogenous Rb gene.
- Rb gene transduction resulted in altered cell morphology, reduced growth rate, and decreased colony formation.
- Cell cycle arrest was observed at the G(0)/G(1) checkpoint.
- Connexin43 expression and GJIC were significantly enhanced in Rb-transduced cells.
Conclusions:
- Transduction of the N-terminal truncated Rb gene effectively inhibits the malignant phenotype of OS732 osteosarcoma cells.
- This intervention leads to a notable increase in GJIC, suggesting a potential therapeutic mechanism.