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Supplementary material for this article can be found on the H EPATOLOGY website ( http://interscience.wiley.com/jpages/0270-9139/suppmat/index.html ).

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suppmat_fig1.tif1773KEffect of SAMe and MTA on the steady state protein amount of Bcl-xS and Bcl-xL. In A) and B), HepG2 cells were treated with either SAMe or MTA (0 to 2.5mM for 24 hrs, or 1mM for 0 to 48 hrs). In C) and D), HepG2 cells (C) or mouse hepatocytes (D) were treated with either SAMe or MTA (1mM for 24 hrs). Total cell lysates (50µg) were subjected to Western blot analysis using anti- Bcl-xS or anti-Bcl-xL antibodies as described in Methods. The same membranes were stripped and probed with antibodies against actin to ensure equal protein loading. The right panels show densitometric changes expressed as % of control. *p<0.05 vs. respective controls.
suppmat_fig2.tif466KEffect of trichostatin A (Tricho), cycloleucine (CL), and 3-deaza-adenosine (3-dea) on SAMe (A) and MTA (B)-mediated induction of Bcl-xS. Total RNAs from HepG2 cells pretreated with trichostatin A (3µM, an inhibitor of histone deacetylase), cycloleucine (20mM, an inhibitor of methionine adenosyltransferase) or 3-deaza-adenosine (10µM, an inhibitor of methylation) for 2 hrs followed by co-treatment with SAMe (1mM) or MTA (1mM) for another 24 hrs were subjected to RT-PCR to evaluate the effect on Bcl-x gene expression as described in Methods. Representative RT-PCRs are shown.
suppmat_fig3.tif1476KEffect of SAMe and MTA on Bcl-xS expression (part A), PP1 catalytic subunit expression (part B) and apoptosis (parts C and D) in 293 cells. In part A, 293 cells were treated with SAMe or MTA (0 to 5mM for 24 hrs, or 1mM for 0 to 48 hrs) and Western blot analysis was performed using anti-Bcl-xL/S antibodies. In part B, PP1 catalytic subunit Northern blot analysis was performed after 24 hr treatment with either MTA or SAMe (1mM). The same membrane was rehybridized with the cDNA probe for b-actin to ensure equal loading. In part C, 293 cells were treated with SAMe or MTA (0 to 5mM) for 24 hrs and apoptosis was quantitated as described in Methods. In part D, 293 cells were pretreated with calyculin A (0 to10nM) for 2 hrs followed by SAMe or MTA (1mM) for 24 hrs. Apoptosis was quantitated as described in Methods. Note tha calyculin A prevented apoptosis in a dose-dependent manner as in HepG2 cells.

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