Hepatocyte-specific Epidermal Growth Factor Receptor Deletion Promotes Fibrosis but has no Effect on Steatosis in Fast-food Diet Model of Metabolic Dysfunction-associated Steatotic Liver Disease.

Background & Aims: Metabolic dysfunction-associated steatotic liver disease (MASLD) has become the most prevalent chronic liver disorder, with no approved treatment. Our previous work demonstrated the efficacy of a pan-ErbB inhibitor, Canertinib, in reducing steatosis and fibrosis in a murine fast-food diet (FFD) model of MASLD. The current study explores the effects of hepatocyte-specific ErbB1 (ie, epidermal growth factor receptor [EGFR]) deletion in the FFD model.

Antagonizing Activin A/p15 Signaling as Therapeutic Strategy for Liver Disease.

Background/aim: Activin A is involved in the pathogenesis of human liver diseases, but its therapeutic targeting is not fully explored. Here, we tested the effect of novel, highly specific small-molecule-based activin A antagonists (NUCC-474/555) in improving liver regeneration following partial hepatectomy and halting fibrosis progression in models of chronic liver diseases (CLDs).

Methods: Cell toxicity of antagonists was determined in rat hepatocytes and Huh-7 cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay.

Mechanism and Effect of HNF4α Decrease in a Rat Model of Cirrhosis and Liver Failure.

Background & Aims: HNF4α, a master regulator of liver development and the mature hepatocyte phenotype, is down-regulated in chronic and inflammatory liver disease. We used contemporary transcriptomics and epigenomics to study the cause and effects of this down-regulation and characterized a multicellular etiology.

Methods: Progressive changes in the rat carbon tetrachloride model were studied by deep RNA sequencing and genome-wide chromatin immunoprecipitation sequencing analysis of transcription factor (TF) binding and chromatin modification.

Hepatocyte Smoothened Activity Controls Susceptibility to Insulin Resistance and Nonalcoholic Fatty Liver Disease.

Background & Aims: Nonalcoholic steatohepatitis (NASH), a leading cause of cirrhosis, strongly associates with the metabolic syndrome, an insulin-resistant proinflammatory state that disrupts energy balance and promotes progressive liver degeneration. We aimed to define the role of Smoothened (Smo), an obligatory component of the Hedgehog signaling pathway, in controlling hepatocyte metabolic homeostasis and, thereby, susceptibility to NASH.

Methods: We conditionally deleted Smo in hepatocytes of healthy chow-fed mice and performed metabolic phenotyping, coupled with single-cell RNA sequencing (RNA-seq), to characterize the role of hepatocyte Smo in regulating basal hepatic and systemic metabolic homeostasis.

Dysregulation of Lipid and Glucose Homeostasis in Hepatocyte-Specific SLC25A34 Knockout Mice.

Nonalcoholic fatty liver disease (NAFLD) is an epidemic affecting 30% of the US population. It is characterized by insulin resistance, and by defective lipid metabolism and mitochondrial dysfunction in the liver. SLC25A34 is a major repressive target of miR-122, a miR that has a central role in NAFLD and liver cancer.

Gadd45 in the Liver: Signal Transduction and Transcriptional Mechanisms.

Injury and growth stimulation both remarkably increase the hepatic expression of Gadd45β. This contrasts with expression in liver cancer, where promoter methylation frequently silences Gadd45β, due to a suppressive function that is often proapoptotic. In normal hepatocytes, Gadd45β facilitates cell survival, growth, and proliferation.

Lymphocyte-Specific Protein-1 Suppresses Xenobiotic-Induced Constitutive Androstane Receptor and Subsequent Yes-Associated Protein-Activated Hepatocyte Proliferation.

Activation of constitutive androstane receptor (CAR) transcription factor by xenobiotics promotes hepatocellular proliferation, promotes hypertrophy without liver injury, and induces drug metabolism genes. Previous work demonstrated that lymphocyte-specific protein-1 (LSP1), an F-actin binding protein and gene involved in human hepatocellular carcinoma, suppresses hepatocellular proliferation after partial hepatectomy. The current study investigated the role of LSP1 in liver enlargement induced by chemical mitogens, a regenerative process independent of tissue loss.

Contributes to Mid-Hindbrain Patterning by Regulation of mdDA Neuronal Cell-Fate and Repression of a Hindbrain-Specific Cell-Fate.

is a member of the NK homeobox family and resembles both in homology and expression pattern. However, while is required for development of serotonergic neurons, the role of in the mid-hindbrain region is still ill-defined. We have previously shown that expression is downregulated upon loss of during development.

Cubilin-, megalin-, and Dab2-dependent transcription revealed by CRISPR/Cas9 knockout in kidney proximal tubule cells.

The multiligand receptors megalin () and cubilin () and their endocytic adaptor protein Dab2 () play essential roles in maintaining the integrity of the apical endocytic pathway of proximal tubule (PT) cells and have complex and poorly understood roles in the development of chronic kidney disease. Here, we used RNA-sequencing and CRISPR/Cas9 knockout (KO) technology in a well-differentiated cell culture model to identify PT-specific transcriptional changes that are directly consequent to the loss of megalin, cubilin, or Dab2 expression. KO of had the greatest transcriptional effect, and nearly all genes whose expression was affected in KO and KO cells were also changed in KO cells.

Deletion of conserved non-coding sequences downstream from NKX2-1: A novel disease-causing mechanism for benign hereditary chorea.

Background: Benign hereditary chorea (BHC) is an autosomal dominant disorder characterized by early-onset non-progressive involuntary movements. Although NKX2-1 mutations or deletions are the cause of BHC, some BHC families do not have pathogenic alterations in the NKX2-1 gene, indicating that mutations of non-coding regulatory elements of NKX2-1 may also play a role.

Methods And Results: By using whole-genome microarray analysis, we identified a 117 Kb founder deletion in three apparently unrelated BHC families that were negative for NKX2-1 sequence variants.

Transcriptional Programs Driving Shear Stress-Induced Differentiation of Kidney Proximal Tubule Cells in Culture.

Cultured cell models are an essential complement to dissecting kidney proximal tubule (PT) function in health and disease but do not fully recapitulate key features of this nephron segment. We recently determined that culture of opossum kidney (OK) cells under continuous orbital shear stress (OSS) significantly augments their morphological and functional resemblance to PTs . Here we used RNASeq to identify temporal transcriptional changes upon cell culture under static or shear stress conditions.

Yes-Associated Protein Is Crucial for Constitutive Androstane Receptor-Driven Hepatocyte Proliferation But Not for Induction of Drug Metabolism Genes in Mice.

Background And Aims: Constitutive androstane receptor (CAR) agonists, such as 1,4-bis [2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), are known to cause robust hepatocyte proliferation and hepatomegaly in mice along with induction of drug metabolism genes without any associated liver injury. Yes-associated protein (Yap) is a key transcription regulator that tightly controls organ size, including that of liver. Our and other previous studies suggested increased nuclear localization and activation of Yap after TCPOBOP treatment in mice and the potential role of Yap in CAR-driven proliferative response.

Targeted Inhibition of the E3 Ligase SCF Has Antitumor Activity in -Deficient Human and Mouse Small-Cell Lung Cancer.

The tumor suppressor gene is mutated in highly aggressive tumors including small-cell lung cancer (SCLC), where its loss, along with , is required and sufficient for tumorigenesis. While -mutant cells fail to arrest at G-S in response to cell-cycle restriction point signals, this information has not led to effective strategies to treat -deficient tumors, as it is challenging to develop targeted drugs for tumors that are driven by the loss of gene function. Our group previously identified Skp2, a substrate recruiting subunit of the SCF-Skp2 E3 ubiquitin ligase, as an early repression target of pRb whose knockout blocked tumorigenesis in Rb1-deficient prostate and pituitary tumors.

Biliary Obstruction Promotes Multilineage Differentiation of Hepatic Stem Cells.

Because of their high regenerative potential, stem cells are an ideal resource for development of therapies that replace injured tissue mass and restore function in patients with end-stage liver diseases. Using a rat model of bile duct ligation (BDL) and biliary fibrosis, we investigated cell engraftment, liver repopulation, and ectopic tissue formation after intrasplenic transplantation of epithelial stem/progenitor cells. Fetal liver cells were infused into the spleens of Fisher 344 rats with progressing biliary fibrosis induced by common BDL or rats without BDL.

Pharmacologic Inhibition of Epidermal Growth Factor Receptor Suppresses Nonalcoholic Fatty Liver Disease in a Murine Fast-Food Diet Model.

Epidermal growth factor receptor (EGFR) is a critical regulator of hepatocyte proliferation and liver regeneration. Our recent work indicated that EGFR can also regulate lipid metabolism during liver regeneration after partial hepatectomy. Based on these findings, we investigated the role of EGFR in a mouse model of nonalcoholic fatty liver disease (NAFLD) using a pharmacological inhibition strategy.

Shear stress and oxygen availability drive differential changes in opossum kidney proximal tubule cell metabolism and endocytosis.

Kidney proximal tubule (PT) cells have high-metabolic demands to drive the extraordinary ion and solute transport, water reabsorption, and endocytic uptake that occur in this nephron segment. Increases in renal blood flow alter glomerular filtration rate and lead to rapid mechanosensitive adaptations in PT transport, impacting metabolic demand. Although the PT reabsorbs essentially all of the filtered glucose, PT cells rely primarily on oxidative metabolism rather than glycolysis to meet their energy demands.

Binding of Drug-Activated CAR/Nr1i3 Alters Metabolic Regulation in the Liver.

The constitutive androstane receptor (CAR/Nr1i3) regulates detoxification of drugs and other xenobiotics by the liver. Binding of these compounds, activating ligands, causes CAR to translocate to the nucleus and stimulate genes of detoxification. However, CAR activation also changes metabolism and induces rapid liver growth.

Constitutive Activation of the Human Aryl Hydrocarbon Receptor in Mice Promotes Hepatocarcinogenesis Independent of Its Coactivator Gadd45b.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or dioxin, is a potent liver cancer promoter through its sustained activation of the aryl hydrocarbon receptor (Ahr) in rodents. However, the carcinogenic effect of TCDD and AHR in humans has been controversial. It has been suggested that the inter-species difference in the carcinogenic activity of AhR is largely due to different ligand affinity in that TCDD has a 10-fold lower affinity for the human AHR compared with the mouse Ahr.

Loss of hepatocyte β-catenin protects mice from experimental porphyria-associated liver injury.

Background & Aims: Porphyrias result from anomalies of heme biosynthetic enzymes and can lead to cirrhosis and hepatocellular cancer. In mice, these diseases can be modeled by administration of a diet containing 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC), which causes accumulation of porphyrin intermediates, resulting in hepatobiliary injury. Wnt/β-catenin signaling has been shown to be a modulatable target in models of biliary injury; thus, we investigated its role in DDC-driven injury.

Hdac1 Regulates Differentiation of Bipotent Liver Progenitor Cells During Regeneration via Sox9b and Cdk8.

Background & Aims: Upon liver injury in which hepatocyte proliferation is compromised, liver progenitor cells (LPCs), derived from biliary epithelial cells (BECs), differentiate into hepatocytes. Little is known about the mechanisms of LPC differentiation. We used zebrafish and mouse models of liver injury to study the mechanisms.

Activin A is a prominent autocrine regulator of hepatocyte growth arrest.

Activin A, a multifunctional cytokine, plays an important role in hepatocyte growth suppression and is involved in liver size control. The present study was aimed to determine the cell location of activin A in the normal rat liver microenvironment and the contribution of activin A signaling to the hepatocyte phenotype to obtain insight into molecular mechanisms. Immunohistochemical and hybridization analyses identified hepatocytes as the major activin A-positive cell population in normal liver and identified mast cells as an additional activin A source.

β-Catenin regulation of farnesoid X receptor signaling and bile acid metabolism during murine cholestasis.

Unlabelled: Cholestatic liver diseases result from impaired bile flow and are characterized by inflammation, atypical ductular proliferation, and fibrosis. The Wnt/β-catenin pathway plays a role in bile duct development, yet its role in cholestatic injury remains indeterminate. Liver-specific β-catenin knockout mice and wild-type littermates were subjected to cholestatic injury through bile duct ligation or short-term exposure to 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet.

The transcriptome of the opossum kidney OK proximal tubule cell line.

The OK cell line derived from the kidney of a female opossum has proven to be a useful model in which to investigate the unique regulation of ion transport and membrane trafficking mechanisms in the proximal tubule (PT). Sequence data and comparison of the transcriptome of this cell line to eutherian mammal PTs would further broaden the utility of this culture model. However, the genomic sequence for is not available and although a draft genome sequence for the opossum (sequenced in 2012 by the Broad Institute) exists, transcripts sequenced from both species show significant divergence.

Triple Staining Including FOXA2 Identifies Stem Cell Lineages Undergoing Hepatic and Biliary Differentiation in Cirrhotic Human Liver.

Recent investigations have reported many markers associated with human liver stem/progenitor cells, "oval cells," and identified "niches" in diseased livers where stem cells occur. However, there has remained a need to identify entire lineages of stem cells as they differentiate into bile ducts or hepatocytes. We have used combined immunohistochemical staining for a marker of hepatic commitment and specification (FOXA2 [Forkhead box A2]), hepatocyte maturation (Albumin and HepPar1), and features of bile ducts (CK19 [cytokeratin 19]) to identify lineages of stem cells differentiating toward the hepatocytic or bile ductular compartments of end-stage cirrhotic human liver.

Biliary fibrosis drives liver repopulation and phenotype transition of transplanted hepatocytes.

Background & Aims: Current research focuses on developing alternative strategies to restore decreased liver mass prior to the onset of end-stage liver disease. Cell engraftment/repopulation requires regeneration in normal liver, but we have shown that severe liver injury stimulates repopulation without partial hepatectomy (PH). We have now investigated whether a less severe injury, secondary biliary fibrosis, would drive engraftment/repopulation of ectopically transplanted mature hepatocytes.