Neonatal thyroxine activation modifies epigenetic programming of the liver

The type 2 deiodinase (D2) in the neonatal liver accelerates local thyroid hormone triiodothyronine (T3) production and expression of T3-responsive genes. Here we show that this surge in T3 permanently modifies hepatic gene expression. Liver-specific Dio2 inactivation (Alb-D2KO) transiently increases H3K9me3 levels during post-natal days 1–5 (P1–P5), and results in methylation of 1,508 DNA sites (H-sites) in the adult mouse liver. These sites are associated with 1,551 areas of reduced chromatin accessibility (RCA) within core promoters and 2,426 within intergenic regions, with reduction in the expression of 1,363 genes. There is strong spatial correlation between density of H-sites and RCA sites. Chromosome conformation capture (Hi-C) data reveals a set of 81 repressed genes with a promoter RCA in contact with an intergenic RCA ~300 Kbp apart, within the same topologically associating domain (χ2 = 777; p < 0.00001). These data explain how the systemic hormone T3 acts locally during development to define future expression of hepatic genes.

Generation of mouse hepatobiliary organoids from hepatocyte progenitors and cholangiocytes isolated from healthy adult mouse liver

Number of liver organoids have been reported, though it is not clearly shown whether the functional connection between hepatocytes and cholangiocytes is recapitulated in those organoids. Here, we report generation of a hepatobiliary tubular organoid (HBTO) using mouse hepatocyte progenitors called small hepatocytes (SHs) and cholangiocytes. SHs differentiate and form the bile canalicular network in HBTOs and secret metabolites into the canaliculi, which are then transported into the biliary structure. Hepatocytes in the organoid acquire and maintain metabolic functions including albumin secretion and cytochrome P450 activities, over the long term. We provide the step-by-step protocol for induction of HBTO including isolation of cholangiocytes and SHs and co-culture of these two types of cell to generate functional connections between hepatocytes and cholangiocytes.

Liver type 1 innate lymphoid cells develop locally via an interferon-γ–dependent loop

The pathways that lead to the development of tissue-resident lymphocytes, including liver type 1 innate lymphoid cells (ILC1s), remain unclear. We show here that the adult mouse liver contains Lin−Sca-1+Mac-1+ hematopoietic stem cells derived from the fetal liver. This population includes Lin−CD122+CD49a+ progenitors that can generate liver ILC1s but not conventional natural killer cells. Interferon-γ (IFN-γ) production by the liver ILC1s themselves promotes the development of these cells in situ, through effects on their IFN-γR+ liver progenitors. Thus, an IFN-γ–dependent loop drives liver ILC1 development in situ, highlighting the contribution of extramedullary hematopoiesis to regional immune composition within the liver.

Hepatocyte-specific deletion of XBP1 sensitizes mice to liver injury through hyperactivation of IRE1α

X-box binding protein-1 (XBP1) is a transcription factor that plays a central role in controlling cellular responses to endoplasmic reticulum (ER) stress. Under stress conditions, the transcriptionally active form of XBP1 is generated by unique splicing of Xbp1 mRNA by the ER-resident protein inositol-requiring enzyme-1 (IRE1α). Genetic deletion of XBP1 has multiple consequences: some resulting from the loss of the transcription factor per se, and others related to compensatory upstream activation of IRE1α. The objective of the current study was to investigate the effects of XBP1 deletion in adult mouse liver and determine to what extent they are direct or indirect. XBP1 was deleted specifically from hepatocytes in adult Xbp1fl/fl mice using AAV8-Transthyretin-Cre (Xbp1Δhep). Xbp1Δhep mice exhibited no liver disease at baseline, but developed acute biochemical and histologic liver injury in response to a dietary challenge with fructose for 4 wk. Fructose-mediated liver injury in Xbp1Δhep mice coincided with heightened IRE1α activity, as demonstrated by cJun phosphorylation and regulated IRE1α -dependent RNA decay (RIDD). Activation of eIF2α was also evident, with associated up-regulation of the pro-apoptotic molecules CHOP, BIM and PUMA. To determine whether the adverse consequences of liver-specific XBP1 deletion were due to XBP1 loss or heightened IRE1α activity, we repeated a fructose challenge in mice with liver-specific deletion of both XBP1 and IRE1α (Xbp1Δhep;IRE1αΔhep). Xbp1Δhep;IRE1αΔhep mice were protected from fructose-mediated liver injury and failed to exhibit any of the signs of ER stress seen in mice lacking XBP1 alone. The protective effect of IRE1α deletion persisted even with long-term exposure to fructose. Xbp1Δhep mice developed liver fibrosis at 16 wk, but Xbp1Δhep;IRE1αΔhep mice did not. Overall, the results indicate that the deleterious effects of hepatocyte-specific XBP1 deletion are due primarily to hyperactivation of IRE1α. They support further exploration of IRE1α as a contributor to acute and chronic liver diseases.

Collapse of the hepatic gene regulatory network in the absence of FoxA factors

The FoxA transcription factors are critical for liver development through their pioneering activity, which initiates a highly complex regulatory network thought to become progressively resistant to the loss of any individual hepatic transcription factor via mutual redundancy. To investigate the dispensability of FoxA factors for maintaining this regulatory network, we ablated all FoxA genes in the adult mouse liver. Remarkably, loss of FoxA caused rapid hepatocyte dedifferentiation manifested by a massive reduction in the expression of key liver genes. Interestingly, expression of these genes was reduced back to the low levels of the fetal prehepatic endoderm stage, leading to necrosis and lethality within days. Mechanistically, we found FoxA proteins to be required for maintaining enhancer activity, chromatin accessibility, nucleosome positioning and binding by HNF4α. Thus, the FoxA factors act continuously, guarding hepatic enhancer activity throughout life.