Objective To preliminarily investigate whether the expression and methylation of long non-coding RNA LINC00472 in osteosarcoma MG-63 cells are regulated by decitabine (DAC) and DNMT1.

Methods MG-63 cells were treated with 5 μmol/L DAC, a demethylation inducer, for 48 h. Real-time quantitative PCR (RT-qPCR) was used to detect the effect of DAC on LINC00472 mRNA expression level; methylation-specific PCR (MSP) and bisulfite sequencing PCR (BSP) were used to detect changes in CpG island methylation in the LINC00472 promoter region. After lentivirus-mediated silencing of DNMT1 in MG-63 cells, the effects on cell function were assessed by CCK-8 assay, colony formation assay, wound-healing assay, and Transwell migration and invasion assays. RT-qPCR was used to assess the effect of DNMT1 on the expression levels of LINC00472 mRNA in MG-63 cells, MSP and BSP were used to assess the effect of DNMT1 on the methylation levels of CpG islands in the LINC00472 promoter region of MG-63 cells.

Results RT-qPCR showed that DAC induced LINC00472 overexpression. Methylation analysis revealed that the CpG islands in the LINC00472 promoter region were at an extremely low methylation level (1.4%) in MG-63 cells, and DAC reduced this methylation from 1.4% to complete non-methylation (0%). After DNMT1 silencing, the cell proliferation, colony formation, migration, and invasion of MG-63 were inhibited， the LINC00472 mRNA expression slight increased, and CpG island methylation in the LINC00472 promoter region decreased (from 1.4% to 0.7%).

Conclusion In MG-63 cells, the LINC00472 promoter region exhibits extremely low methylation. DAC can upregulate its expression and induce complete demethylation, suggesting that methylation may be involved in its regulation. but not as the primary pathway. DNMT1 silencing inhibits cell growth but has no significant effect on LINC00472, possibly due to its inherently low methylation status.

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