s-adenosylhomocysteine and Glioma

S-Adenosylhomocysteine (SAH) is a risk factor for many diseases, including tumor progression and neurodegenerative disease. In this study, we examined the hypothesis that SAH may indirectly enhance the invasion of C6 glioma cells by induction of matrix metalloproteinase-2 (MMP-2) secreted from the murine microglia BV2 cells. We obtained conditioned medium (CM) by incubating BV2 cells with SAH (1-50nM) for 24 h. We found that the SAH-containing CM (SAH-BV2-CM) strongly enhanced the invasiveness of C6 glioma cells and that this effect increased with increasing concentrations of SAH in the SAH-BV2-CM. The effect of CM could be attributed to its MMP-2 activity, as a result of increased protein and messenger RNA expression of MMP-2 in BV2 cells induced by SAH. In BV2 cells treated with SAH, the binding abilities of nuclear factor-kappa B (NF-kappaB) and stimulatory protein-1 (Sp1) to the MMP-2 promoter were increased, whereas the level of NF-kappaB inhibitor was decreased. In addition, SAH significantly increased the phosphorylation of extracellular signal-regulated kinase (ERK) and phosphatidylinositol-3-kinase/serine/threonine protein kinase (or protein kinase B) (PI3K/Akt) proteins but did not affect that of c-Jun NH2-terminal kinase or p38. Pretreatment of BV2 cells with an inhibitor specific for ERK (U0126) markedly abated the expression of ERK and MMP-2. Furthermore, SAH significantly and dose dependently decreased tissue inhibitor of metalloproteinase-2 (TIMP-2) in BV2 cells. Thus, SAH may induce the invasiveness of C6 glioma cells by decreased TIMP-2 expression and increased MMP-2 expression in BV2 cells. The latter effect is likely mediated through the ERK and PI3K/Akt pathways, with increased binding activities of NF-kappaB and Sp1 to the MMP-2 gene promoter.

Topics: Animals; Base Sequence; Cell Line; Cell Line, Tumor; Chromatin Immunoprecipitation; Culture Media, Conditioned; DNA Primers; Glioma; Matrix Metalloproteinase 2; Mice; Microglia; Neoplasm Invasiveness; NF-kappa B; Protein Kinases; Reverse Transcriptase Polymerase Chain Reaction; S-Adenosylhomocysteine; Signal Transduction; Tissue Inhibitor of Metalloproteinase-2; Transcription Factors

The DNA repair enzyme O(6)-methylguanine DNA methyltransferase (MGMT) protects cells against the cytotoxic effects of alkylating agents. Therefore, modulation of MGMT expression in tumors is a possible strategy for improving the efficiency of cancer therapy. MGMT expression and activity is lost frequently in association with DNA hypermethylation of the MGMT promoter region. Since DNA and mRNA methylation are controlled by intracellular S-adenosylmethionine (AdoMet) and S-adenosylhomocysteine (AdoHcy) levels, we hypothesized a role for AdoMet/AdoHcy ratio in the regulation of MGMT promoter methylation and mRNA expression. Our initial studies showed that AdoMet/AdoHcy ratios vary over a wide range (7.0-50) in different glioblastoma and hepatoma cell lines. The studied cell lines exhibit distinct MGMT promoter methylation patterns: MGMT promoter was completely unmethylated in LN-18 and Tu 132 cells, hypermethylated in LN-229, U87-MG, and Tu 113 cells, and partially methylated in HepG2 cells. Furthermore, MGMT promoter methylation patterns and global DNA methylation are not related to intracellular AdoMet/AdoHcy ratio under control conditions. To lower AdoMet/AdoHcy ratio to values <1 we used AdoHcy hydrolase inhibitor adenosine-2',3'-dialdehyde (30 microM) and found that neither short-term (24 h) nor long-term changes (7 weeks) in AdoMet/AdoHcy ratio altered global or MGMT promoter methylation. However, experimentally elevated AdoHcy levels significantly decreased MGMT mRNA levels by >50% in all MGMT-expressing cell lines, which is most likely the result of impaired mRNA methylation. Thus, the present study suggests elevation of AdoHcy levels by AdoHcy hydrolase inhibition as a novel pharmacological approach to modulate MGMT expression and to increase the responsiveness to alkylating agents.

Topics: Adenosylhomocysteinase; Antineoplastic Agents; Cell Line, Tumor; DNA Methylation; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Glioma; Hepatoblastoma; Humans; Liver Neoplasms; O(6)-Methylguanine-DNA Methyltransferase; Promoter Regions, Genetic; RNA, Messenger; S-Adenosylhomocysteine