Arsenic trioxide (
As(2)O(3); ATO, TRISENOX®) is used to treat patients with refractory or relapsed acute promyelocytic leukaemia while its application for treatment of solid
cancers like
glioblastoma is still under evaluation. In the present study, we investigated the interaction of
arsenic trioxide with
metallothionein (MT)
isoforms as a possible (protective response) resistance of
glioblastoma cells to
arsenic-induced cytotoxicity. Special attention was focused on MT3, the
isoform expressed mainly in the brain. MT3 has low
metal inducibility, fast
metal binding/releasing properties and outstanding neuronal inhibitory activity. The human
astrocytoma (
glioblastoma) cell line U87 MG was treated with 0.6, 2 and 6-7 μM
arsenic (equivalent to 0.3, 1 and 3-3.5 μM
As(2)O(3)) for 12, 24 or 48 h and gene expression for different MT
isoforms, namely MT2A, MT1A, MT1F, MT1X, MT1E and MT3, was measured by real time qPCR using
SYBR Green I and Taqman® gene expression assays. TfR,
18S rRNA, GAPDH and AB were tested as reference genes, and the last two evaluated to be appropriate in conditions of low (GAPDH) and high (AB)
arsenic exposure. The gene expression of MT3 gene was additionally tested and confirmed by restriction
enzyme analysis with PvuII. In the given conditions the mRNAs of six MT
isoforms were identified in human
glioblastoma cell line U87 MG. Depending on
arsenic exposure conditions, an increase or decrease of MT gene expression was observed for each
isoform, with the highest increase for
isoforms MT1X, MT1F and MT2A
mRNA (up to 13-fold) and more persistent decreases for MT1A, MT1E and MT3
mRNA. Despite the common assumption of the noninducibility of MT3, the evident MT3
mRNA increase was observed during high As exposure (up to 4-fold). In conclusion, our results clearly demonstrate the influence of As on MT
isoform gene expression. The MT1X, MT1F and MT2A increase could represent brain tumour acquired resistance to As cytotoxicity while the MT3 increase is more enigmatic, with its possible involvement in
arsenic-related induction of type II cell death.