Figure 7.
Hepatocytes Expressing Oncogenic URI Produce α-Ketoglutarate and Galectin-3 to Preserve an HPC Undifferentiated State
(A) Schematic representation of intermediary metabolism induced in hURI-tetOFFhep mice. Data were obtained and analyzed from previous iTRAQ data obtained from livers from 1- or 8-week-old hURI-tetOFFhep mice (Tummala et al., 2014). Fold changes are represented within the brackets. Upregulated enzymes are represented in red, and downregulated enzymes are represented in green.
(B) Histogram representing α-ketoglutarate levels from livers from either NR-treated or untreated 12-week-old hURI-tetOFFhep mice as indicated. n = 6 for controls and mutants; n = 5 for controls and mutants treated with NR.
(C) Histogram representing α-ketoglutarate levels from livers from 4-week-old hURI-tetOFFhep mice (mutant) and DEN-treated C57BL/6 mice. Mice were treated with either GlcNAc or 4-F-GalNAc for 3 weeks. n = 3 for mutants treated with GlcNAc.
(D) Schematic representation of AML-12 and BMOL cell treatment. Media from HA-URI-overexpressing AML-12 cells treated either with DMSO or BPTES (10 μM) was used to culture BMOL cells for 3 days.
(E) Cell number of BMOL cells after 0, 24, 48, and 72 hr of incubation with media from AML-12 cells from (D).
(F) Histogram representing α-ketoglutarate levels in media from AML-12 cells stably overexpressing EGFP or HA-URI.
(G) qRT-PCR of BMOL cells cultured with media from AML-12 cells as described in (D).
(H) WB of BMOL cells after 72 hr incubation with media from AML-12 cells stably overexpressing EGFP or HA-URI and transfected with either siCTR or siGal3. Membrane was blotted with the indicated antibodies.
(I) qRT-PCR of BMOL cells cultured with media from AML-12 cells as described in (H).
(J) Scheme representing hepatocytes expressing oncogenic URI-secreted galectin-3 to induce HPC proliferation, whereas α-ketoglutarate preserves an HPC undifferentiated state during expansion.
Data are presented as mean ± SEM. ns, non significant; ∗p ≤ 0.05; ∗∗∗p ≤ 0.001.