The sequence of events triggering liver regeneration after acute loss of hepatic mass has been the subject of much investigation during the past 20 years. 1 Rapid changes in gene expression and activation of receptors and transcription factors occur immediately after partial hepatectomy (PHx). 2,3 Several potential signaling stimuli are released in the liver or in circulation after the loss of hepatic parenchyma. These stimuli have effects on liver and on hepatocytes (or other hepatic cells) in culture. Such stimuli include cytokines (eg, tumor necrosis factor α, interleukin-6, norepinephrine, transforming growth factor [TGF]-β 1) and several growth factors. Cytokines are not direct mitogens for hepatocytes in culture and, after removal of their effects (eg, in mouse genetic models or by pharmacologic agents), liver regeneration is retarded or decreased, but hepatic mass is eventually restored nonetheless. Although this may suggest that the role of cytokines is not critical or essential, this should not imply that the events they mediate are not important or that they do not actually occur. The preponderance of evidence suggests that the cytokine effects facilitate optimal timing and orchestration of the early signaling events after PHx, summarized under the term priming of hepatocytes. 4The effects of growth factors are mediated through their receptors. Two receptor-ligand and growth factor signaling systems appear to be mainly involved in liver regeneration: hepatocyte growth factor and its receptor (Met) and the epidermal growth factor receptor (EGFR) and its relatively large family of ligands and coreceptors. The receptor for EGFR was the first one that was shown to play a role in liver regeneration. A seminal publication by Earp et al 5 showed that EGFR was phosphorylated and downregulated after PHx, suggesting binding of ligands and endocytosis.The signaling scenario related to EGFR family members and its ligands is quite complex. The EGFR itself (also known as ErbB or HER) is a member of a family of four. The other members are ErbB-2 (HER-2, NEU), ErbB-3 (HER-3), and ErbB-4 (HER-4). 6 Of these, HER-4 is expressed in only a limited number of tissues and it does not appear to be expressed in adult or embryonic liver. There are many ligands for EGFR, including epidermal growth factor (EGF), TGF-α, amphiregulin, heparin-binding EGF ( HB-EGF), cripto, epiregulin, and betacellulin.All of these considerations are relevant in putting in context the significant findings presented by Berasain et al 7 in this issue of Gastroenterology. The authors found that amphiregulin, a ligand for EGFR, is expressed early (within 30 minutes) during liver regeneration after PHx. In addition, they also found that liver regeneration is substantially decreased and retarded in mice with homozygous deletion of amphiregulin. The findings suggest a unique role for amphiregulin that cannot be substituted by the other ligands for EGFR. The time course of expression of amphiregulin corresponds well with the pattern of tyrosine phosphorylat...

Evidence from human histopathology and experimental studies with rodents and zebrafish has shown that hepatocytes and cholangiocytes may function as facultative stem cells for each other in conditions of impaired regeneration. The interpretation of the findings derived from these studies has generated considerable discussion and some controversies. This review examines the evidence obtained from the different experimental models and considers implications that these studies may have for human liver disease. Few topics of liver tissue biology have attracted as much attention as the existence of liver-specific tissue stem cells. Routine liver histology reveals two types of epithelial cells, hepatocytes and cholangiocytes (also known as biliary epithelial cells). Endothelial cells line the hepatic capillaries (sinusoids), with macrophages (Kupffer cells) interspersed along the sinusoid lumen. Stellate cells exist under the sinusoids and in close proximity to hepatocytes. None of these cells appears to have functions of a fully committed tissue specific stem cell, analogous to the cells of the intestinal crypts, the basal layer of the epidermis, bone marrow stem cells, etc. Hepatocytes and cholangiocytes can be easily identified based on their morphology and cell-specific biomarkers. Hepatocytes and cholangiocytes, however, often have mutually mixed expression of biomarkers in pathologic conditions. In patients with fulminant hepatic failure (FHF), there is rampant proliferation of cholangiocytes organized in ductular structures (“ductular reaction”1, 2). Many of these cholangiocytes (known as ductular hepatocytes) express biomarkers associated with hepatocytes, (HNF4, albumin, HEPPAR3, etc.). They are seen surrounding cells ranging in size from small to typical hepatocytes, and with a gradient of expression of cholangiocyte-associated biomarkers (e.g. EpCAM) decreasing from the periphery to the center (Regenerative Clusters: see Figure 1). It is not clear in FHF whether cholangiocytes give rise to hepatocytes or vice versa. Most cells in liver tissues from patients with FHF, however, are typical cholangiocytes, so it is likely that these are the source of hepatocytes detected in the (more rarely seen) regenerative clusters. The term “progenitor” cells (used in tissue biology to describe the immediate progeny of stem cells) is most often used to collectively cover these proliferating cells with mixed hepatobiliary biomarkers in rats, mice, humans and fish. This may be inappropriate because it implies that such cells are generated by tissue-specific stem cells, even though such stem cells are not identifiable in the liver. Though the term “progenitor cells” does not fulfill criteria used in other tissues, it does imply a transition from one type of cell differentiation to another. Thus, the term has persisted in hepatic biology, even though it is not entirely appropriate. However, in most of the scenarios below, hepatocytes and cholangiocytes appear to function as “facultative stem cells” for each other. Thus the ter...

Many signals involved in pathophysiology are controlled by hypoxia-inducible factors (HIFs), transcription factors that induce expression of hypoxiaresponsive genes. HIFs are post-translationally regulated by a family of O 2 -dependent HIF hydroxylases: four prolyl 4-hydroxylases and an asparaginyl hydroxylase. Most of these enzymes are abundant in resting liver, which is itself unique because of its physiological O 2 gradient, and they can exist in both nuclear and cytoplasmic pools. In this study, we analyzed the cellular localization of endogenous HIFs and their regulatory hydroxylases in primary rat hepatocytes cultured under hypoxia-reoxygenation conditions. In hepatocytes, hypoxia targeted HIF-1␣ to the peroxisome, rather than the nucleus, where it co-localized with von Hippel-Lindau tumor suppressor protein and the HIF hydroxylases. Confocal immunofluorescence microscopy demonstrated that the HIF hydroxylases translocated from the nucleus to the cytoplasm in response to hypoxia, with increased accumulation in peroxisomes on reoxygenation. These results were confirmed via immunotransmission electron microscopy and Western blotting. Surprisingly, in resting liver tissue, perivenous localization of the HIF hydroxylases was observed, consistent with areas of low pO 2 . In conclusion, these studies establish the peroxisome as a highly relevant site of subcellular localization and function for the endogenous HIF pathway in

Purpose of ReviewThe aim of the study is to review the liver disease caused by alpha-1 antitrypsin deficiency (A1ATD), including pathogenesis, epidemiology, diagnostic testing, and recent therapeutic developments.Recent FindingsTherapeutic approaches target several intracellular pathways to reduce the cytotoxic effects of the misfolded mutant globular protein (ATZ) on the hepatocyte. These include promoting ATZ transport out of the endoplasmic reticulum (ER), enhancing ATZ degradation, and preventing ATZ globule-aggregation.SummaryA1ATD is the leading genetic cause of liver disease among children. It is a protein-folding disorder in which toxic insoluble ATZ proteins aggregate in the ER of hepatocytes leading to inflammation, fibrosis, cirrhosis, and increased risk of hepatocellular carcinoma. The absence of the normal A1AT serum protein also predisposes patients to pan lobar emphysema as adults. At this time, the only approved therapy for A1ATD-associated liver disease is orthotopic liver transplantation, which is curative. However, there has been significant recent progress in the development of small molecule therapies with potential both to preserve the native liver and prevent hepatotoxicity.

In rodents, complete bile duct ligation (cBDL) of the common bile duct is an established surgical technique for studying obstructive cholestasis and bile duct proliferation. However, long-term experiments can lead to increased morbidity and mortality. In select mouse strains with underlying liver disease, meaningful comparisons can be made even with ligation of a single lobe of the liver, which can reduce animal losses and expenses. Here, we describe partial bile duct ligation (pBDL) in the mouse, in which only the left hepatic bile duct is ligated, causing biliary obstruction in the left lobe but not the remaining lobes. With careful microsurgical technique, pBDL experiments can be cost-effective, since the unligated lobe serves as an internal control to the ligated lobes, when subjected to the same conditions in the same animal. Unlike cBDL, a separate sham-operated control group is not necessary. pBDL is highly useful to directly compare localized versus systemic effects of cholestasis and other retained bile components. pBDL can also be repurposed as a novel method to investigate mechanisms related to medications and cell migration.

Objective: Study of liver explants of biliary atresia (BA) patients with successful Kasai portoenterostomy (KP). Methods: Pathology and medical records of BA liver explants from January 2009 to June 2018 with successful KP were reviewed along with appropriate controls. Results: Fourteen out of 68 (20.6%) BA patients with LT had a successful KP. Median age at BA diagnosis, KP and LT was 60.5 days, 61 days, and 10 years, respectively, with conjugated bilirubin (c-bil) normalizing at 12.5 weeks after KP. Advanced fibrosis was diffuse in 2/14 (14.3%) explants, limited to periphery in 11/14 (78.6%) and absent in 1. Hilar partial nodular transformation (PNT) was seen in 11 explants (78.6%) and diffuse nodular regenerative hyperplasia (NRH) in 2 (14.3%). Areas of PNT and NRH showed diffuse portal sclerosis (100%), complete and incomplete portal vein (PV) stenosis (100%), PV herniation (100%), hypervascular portal tracts (20%), periportal abnormal vessels (100%), abundant lymphatic collaterals (100%), mild medial hepatic arterial hypertrophy (100%), and delicate fibrous septae (100%). Extrahepatic PVs showed variable luminal occlusion with mean PV intima to full thickness ratio of 0.6 +/− 0.11; significantly higher than age-matched noncirrhotic (n = 27, 0.08 +/− 0.09; P < 0.0001) and cirrhotic controls (n = 19, 0.34 +/− 0.2; P = 0.0015); and comparable to BA patients with failed KP (P = 0.82) and without KP (P = 0.04). Conclusions: BA patients with successful KP can present with obliterative portal venopathy (OPV). In the context of optimal bile drainage, portal hypertension may not be because of advanced parenchymal fibrosis but possibly because of OPV. Vascular abnormalities of the PV system should be investigated in BA patients.

Alpha-1 Antitrypsin deficiency (A1ATD) can progress to cirrhosis and hepatocellular carcinoma; however, not all patients are susceptible to severe liver disease. Liver transplantation is the only cure for A1ATD-related liver disease. A1ATD is caused by a toxic gain-of-function mutation in the human SERPINA1 gene, generating mis folded ATZ protein "globules" in hepatocytes. These insoluble aggregates overwhelm protein clearance pathways and lead to chronic intracellular stress. This review serves to summarize the basic hepatic mechanisms involved in A1ATD, as described in relevant in vitro and animal models. Potential treatments such as autophagy-enhancing agents and molecular therapies are also discussed. Clinical trials are underway to further assess some of these novel approaches in patients, but more safety and efficacy data is needed to successfully translate these interventions from the laboratory to the clinic.

Background Alpha-1 antitrypsin deficiency (A1ATD) can progress to cirrhosis and hepatocellular carcinoma; however, not all patients are susceptible to severe liver disease. In A1ATD, a toxic gain-of-function mutation generates insoluble ATZ “globules” in hepatocytes, overwhelming protein clearance mechanisms. The relationship between bile acids and hepatocytic autophagy is less clear, but may involve altered gene expression pathways. Based on previous findings that bile duct ligation (BDL) induces autophagy, we hypothesized that retained bile acids may have hepatoprotective effects in PiZZ transgenic mice, which model A1ATD. Methods We performed BDL and partial BDL (pBDL) in PiZZ mice, followed by analysis of liver tissues. Results PiZZ liver subjected to BDL showed up to 50% clearance of ATZ globules, with increased expression of autophagy proteins. Analysis of transcription factors revealed significant changes. Surprisingly nuclear TFEB, a master regulator of autophagy, remained unchanged. pBDL confirmed that ATZ globule clearance was induced by localized stimuli rather than diet or systemic effects. Several genes involved in bile metabolism were over-expressed in globule-devoid hepatocytes, compared to globule-containing cells. Conclusions Retained bile acids led to a dramatic reduction of ATZ globules, with enhanced hepatocyte regeneration and autophagy. These findings support investigation of synthetic bile acids as potential autophagy-enhancing agents.