Context 3,5,3'-L-triiodothyronine (T3) is a potent inducer of hepatocyte proliferation via the Wnt/β-catenin signaling pathway. Previous studies suggested the involvement of rapid noncanonical thyroid hormone receptor (TR)β signaling, directly activating hepatic Wnt/β-catenin signaling independent from TRβ DNA binding. However, the mechanism by which T3 increases Wnt/β-catenin signaling in hepatocytes has not yet been determined. Objective We aimed to determine whether DNA-binding of TRβ is required for stimulation of hepatocyte proliferation by T3. Methods Wild type (WT) mice, TRβ knockout mice (TRβ KO), TRβ mutant mice with either specifically abrogated DNA-binding (TRβ GS) or abrogated direct PI3K activation (TRβ 147F) were treated with T3 for 6h or 7 days. Hepatocyte proliferation was assessed by Ki67 staining and apoptosis by TUNEL-assay. Activation of β-catenin signaling was measured in primary murine hepatocytes. Gene expression was analyzed by microarray, gene set enrichment analysis (GSEA) and qRT-PCR. Results T3 induced hepatocyte proliferation with an increased number of Ki67-positive cells in WT and TRβ 147F mice (9.2±6.5% and 10.1±2.9%) compared to TRβ KO and TRβ GS mice (1.2±1.1% and 1.5±0.9%). Microarray analysis and GSEA showed that genes of the Wnt/β-catenin pathway, among them Fzd8 (Frizzled receptor 8) and Ctnnb1 (β-catenin), were positively enriched only in T3-treated WT and TRβ 147F mice while anti-proliferative factor Btg2 was repressed. Consequently, expression of Ccnd1 (CyclinD1) was induced. Conclusions Instead of directly activating Wnt signaling, T3 and TRβ induce key genes of the Wnt/β-catenin pathway, ultimately stimulating hepatocyte proliferation via CyclinD1. Thus, canonical transcriptional TRβ action is necessary for T3-mediated stimulation of hepatocyte proliferation.

Thyroid hormones (THs) and TH receptor-beta (TRβ) reduce hepatic triglycerides, indicating a therapeutic potential for TH analogs in liver steatosis. To avoid adverse extrahepatic, especially TRα-mediated effects such as tachycardia and bone loss, TH analogs with combined TRβ and hepatocyte specificity are desired. MGL-3196 is a new TH analog that supposedly meets these criteria. Here, we characterize the thyromimetic potential of MGL-3196 in cell-based assays and address its cellular uptake requirements. We studied the contribution of liver-specific organic anion transporters (OATP)1B1 and 1B3 to MGL-3196 action. The TR isoform-specific efficacy of MGL-3196 compared with 3,5,3′-triiodothyronine (T3) was determined with luciferase assays and gene expression analysis in OATP1B1 and OATP1B3 and TRα- or TRβ-expressing cells and in primary murine hepatocytes (PMHs) from wild-type and TRβ knockout mice. We measured the oxygen consumption rate to compare the effects of MGL-3196 and T3 on mitochondrial respiration. We identified OATP1B1 as the primary transporter for MGL-3196. MGL-3196 had a high efficacy (90% that of T3) in activating TRβ, while the activation of TRα was only 25%. The treatment of PMHs with T3 and MGL-3196 at EC50 resulted in a similar induction of Dio1 and repression of Serpina7. In HEK293 cells stably expressing OATP1B1, MGL-3196 had comparable effects on mitochondrial respiration as T3. These data indicate that MGL-3196’s hepatic thyromimetic action, the basis for its therapeutic use, results from a combination of hepatocyte-specific transport by OATP1B1 and the selective activation of TRβ over TRα.