Objective: To study the methylation status of the SLC5A8 and TMS1/ASC genes, candidate tumor-inhibiting genes closely related to the central nervous system, in the promoter regions, the mRNA expression of these 2 genes, and their correlation with the clinical characteristics in human glioma.
Methods: The methylation status of SLC5A8 and TMS1/ASC genes in the promoter regions was studied by methylation specific PCR (MSP) in the specimens of primary astrocytoma from 88 patients, 55 males and 33 females, aged 12 - 81, and 10 specimens of normal brain tissue, all obtained during operation, and in the human glioma cells of the lines U251 and SHG-44. The mRNA expression levels of SLC5A8 and TMS1/ASC genes in 30 specimens of primary glioma and 10 specimens of normal brain tissue were determined by conventional RT-PCR and real-time PCR. 5-Aza-2'-deoxycytidine (5-Aza-CdR), a demethylating agent, was added into the culture fluid of the U251 and SHG-44 cells, and then real-time PCR was used to the methylation status and mRNA expression levels of the SLC5A8 and TMS1/ASC genes.
Results: MSP showed that the SLC5A8 promoter region was hypermethylated in 62 of the 88 specimens of astrocytoma (70.45%) and the TMS1/ASC promoter region was hypermethylated in 51 of the88 specimens of astrocytoma (57.95%). But no methylation of SLC5A8 and TMS1/ASC promoter was detected in the 10 specimens of normal brain tissue. The mRNA expression of SLC5A8 gene and the mRNA expression of TMS1/ASC gene in the specimens of astrocytoma of different pathological grades were all significantly decreased compared to the specimens of normal brain tissue (all P < 0.05). The mRNA expression of SLC5A8 gene was not significantly related to the age and sex, however, the mRNA expression of TMS1/ASC was significantly higher in the age group > 60 than in other age groups (all P < 0.05). Both U251 and SHG-44 glioma cells showed methylation of SLC5A8 and TMS1/ASC genes and after the treatment of 5-Aza-CdR both genes showed reactivated mRNA expression.
Conclusion: Hypermethylation of SLC5A8 and TMS1/ASC genes in the promoter regions may play an important role in the down-regulation of their mRNA levels in glioma. The methylation frequency and mRNA levels of SLC5A8 or TMS1/ASC genes are closely related to the malignant development of glioma.