Hepatology
Abstract
Authors
Jing Zeng, Derrick Zhao, Grayson Way, Andrew Fagan, Michael Fuchs, Puneet Puri, Brian C. Davis, Xuan Wang, Emily C. Gurley, Phillip B. Hylemon, Jian-Gao Fan, Masoumeh Sikaroodi, Patrick M. Gillevet, Huiping Zhou, Jasmohan S. Bajaj
Abstract
Due to the limitations of available in vitro systems and animal models, we lack a detailed understanding of the pathogenetic mechanisms and have minimal treatment options for liver fibrosis. Therefore, we engineered a live cell imaging system that assesses fibrosis in a human multi-lineage hepatic organoid in a microwell (i.e., microHOs). Transcriptomic analysis revealed that TGFβ1 converted mesenchymal cells in microHOs into myofibroblast-like cells resembling those in fibrotic human liver tissue. When pro-fibrotic intracellular signaling pathways were examined, the anti-fibrotic effect of receptor-specific tyrosine kinase inhibitors was limited to the fibrosis induced by the corresponding growth factor, which indicates their anti-fibrotic efficacy would be limited to fibrotic diseases solely mediated by that growth factor. Based upon transcriptomic and transcription factor activation analyses in microHOs, GSK3β and p38 MAPK inhibitors were identified as potential new broad-spectrum therapies for liver fibrosis. Other new therapies could subsequently be identified using the microHO system.
Authors
Yuan Guan, Zhuoqing Fang, Angelina Hu, Sarah Roberts, Meiyue Wang, Wenlong Ren, Patrik K. Johansson, Sarah C. Heilshorn, Annika Enejder, Gary Peltz
Abstract
Biliary obstruction and cholangiocyte hyperproliferation are important features of cholangiopathies affecting the large extrahepatic bile duct (EHBD). The mechanisms underlying obstruction-induced cholangiocyte proliferation in the EHBD remain poorly understood. Developmental pathways, including WNT signaling, are implicated in regulating injury responses in many tissues, including the liver. To investigate the contribution of WNT signaling to obstruction-induced cholangiocyte proliferation in the EHBD, we used complementary in vivo and in vitro models with pharmacologic interventions and transcriptomic analyses. To model obstruction, we used bile duct ligation (BDL) in mice. Human and mouse biliary organoids and mouse biliary explants were used to investigate the effects of WNT activation and inhibition in vitro. We observed an upregulation of WNT ligand expression associated with increased biliary proliferation following obstruction. Cholangiocytes were identified as both WNT ligand-expressing and WNT responsive cells. Inhibition of WNT signaling decreased cholangiocyte proliferation in vivo and in vitro, while activation increased proliferation. WNT effects on cholangiocyte proliferation were β-catenin-dependent, and we showed a direct effect of WNT7B on cholangiocyte growth. Our studies suggested that cholangiocyte-derived WNT ligands can activate WNT signaling to induce proliferation after obstructive injury. These findings implicated the WNT pathway in injury-induced cholangiocyte proliferation within the EHBD.
Authors
Ashley N. Calder, Mirabelle Q. Peter, John W. Tobias, Nureen H. Mohamad Zaki, Theresa M. Keeley, Timothy L. Frankel, Linda C. Samuelson, Nataliya Razumilava
Abstract
Hepatic macrophages and regulatory T cells (Tregs) play an important role in the maintenance of liver immune homeostasis, but the mechanism by which hepatic macrophages regulate Tregs in acute liver injury remains largely unknown. Here, we found that the hepatic Treg proportion and β-catenin expression in hepatic macrophages were associated with acetaminophen (APAP) and D-galactosamine (D-GalN)/ lipopolysaccharide (LPS)-induced acute liver injury. Interestingly, β-catenin was markedly upregulated only in infiltrating macrophages, but not in resident Kupffer cells. Myeloid-specific β-catenin knockout mice showed an increased inflammatory cell infiltration and hepatocyte apoptosis. Moreover, myeloid β-catenin deficiency decreased the hepatic Treg proportion in the injured liver. Mechanistically, in vitro co-culture experiments revealed that macrophage β-catenin modulated its exosome composition, and influenced Treg differentiation. Using mass spectrometry-based proteomics, we identified that macrophage β-catenin activation increased the level of exosomal α-SNAP, which in turn promoted Treg differentiation. Overall, our findings demonstrated a molecular mechanism that macrophage β-catenin regulated the Treg proportion in the liver by enhancing the expression of exosomal α-SNAP, providing insights into the pathophysiology of acute liver injury.
Authors
Ruobin Zong, Yujie Liu, Mengya Zhang, Buwei Liu, Wei Zhang, Hankun Hu, Changyong Li
Abstract
Excessive fructose intake is a risk factor for the development of obesity and its complications. Targeting ketohexokinase (KHK), the first enzyme of fructose metabolism, has been investigated for the management of MASLD. We compared the effects of systemic, small molecule inhibitor of KHK enzymatic activity to hepatocyte-specific, GalNAc-siRNA mediated knockdown of KHK in mice on a HFD. We measured KHK enzymatic activity, extensively quantified glycogen accumulation, performed RNAseq analysis, and enumerated hepatic metabolites using mass spectrometry. Both KHK siRNA and KHK inhibitor led to an improvement in liver steatosis, however, via substantially different mechanisms. KHK knockdown decreased the de novo lipogenesis pathway, whereas the inhibitor increased the fatty acid oxidation pathway. Moreover, KHK knockdown completely prevented hepatic fructolysis and improved glucose tolerance. Conversely, the KHK inhibitor only partially reduced fructolysis, but it also targeted triokinase, mediating the third step of fructolysis. This leads to the accumulation of fructose-1 phosphate, resulting in glycogen accumulation, hepatomegaly, and impaired glucose tolerance. Overexpression of wild-type, but not kinase-dead KHK in cultured hepatocytes increased hepatocyte injury and glycogen accumulation when treated with fructose. The differences between KHK inhibition and knockdown are, in part, explained by the kinase-dependent and independent effects of KHK on hepatic metabolism.
Authors
Se-Hyung Park, Taghreed Fadhul, Lindsey R. Conroy, Harrison A. Clarke, Ramon C. Sun, Kristina Wallenius, Jeremie Boucher, Gavin O'Mahony, Alessandro Boianelli, Marie Persson, Sunhee Jung, Cholsoon Jang, Analia S. Loria, Genesee J. Martinez, Zachary A. Kipp, Evelyn A. Bates, Terry D. Hinds, Jr., Senad Divanovic, Samir Softic
Abstract
Cell cycle inhibitors have a long history as cancer treatment. Here, we reported that these inhibitors combated cancer partially via Stimulator of IFN genes (STING) signaling pathway. We demonstrated that Paclitaxel (microtubule stabilizer), Palbociclib (cyclin dependent kinase 4/6 inhibitor), AZD1152 and GSK1070916 (aurora kinase B inhibitors) have anti-cancer functions beyond arresting cell cycle. They consistently caused cytosolic DNA accumulation and DNA damage, which inadvertently triggered the cytosolic DNA sensor DEAD-box helicase 41 (DDX41) and activated STING to secrete pro-inflammatory senescence-associated secretory phenotype factors (SASPs). Interestingly, we found that DDX41 was a transcriptional target of HIF. Hypoxia induced expression of DDX41 through HIF-1, making hypoxic HCC cells more sensitive to the anti-mitotic agents in STING activation and SASP production. The SASPs triggered immune cell infiltration in tumors for cancer clearance. The treatment of cell cycle inhibitors, especially Paclitaxel, extends survival by perturbing mouse HCC growth when used in combination with anti-PD-1. We observed a trend that Paclitaxel suppressed STINGWT HCC more effectively than STINGKO HCC, suggesting that STING might contribute to the anti-tumor effects of Paclitaxel. Our study revealed the immune-mediated tumor-suppressing properties of cell cycle inhibitors and suggested combined treatment with immunotherapy as a potential therapeutic approach.
Authors
Po Yee Wong, Cerise Yuen Ki Chan, Helen Do Gai Xue, Chi Ching Goh, Jacinth Wing Sum Cheu, Aki Pui Wah Tse, Misty Shuo Zhang, Yan Zhang, Carmen Chak Lui Wong
Abstract
The complexity of the mechanisms underlying non-alcoholic fatty liver disease (NAFLD) progression remains a significant challenge for the development of effective therapeutics. miRNAs have shown great promise as regulators of biological processes and as therapeutic targets for complex diseases. Here, we study the role of hepatic miR-33, an important regulator of lipid metabolism, during the progression of NAFLD and the development of hepatocellular carcinoma (HCC). We report that miR-33 is elevated in the livers of humans and mice with NAFLD and that its deletion in hepatocytes (miR-33 HKO) improves multiple aspects of the disease, including steatosis and inflammation, limiting the progression to non-alcoholic steatohepatitis (NASH), fibrosis and HCC. Mechanistically, hepatic miR-33 deletion reduces lipid synthesis and promotes mitochondrial fatty acid oxidation, reducing lipid burden. Additionally, absence of miR-33 alters the expression of several known miR-33 target genes involved in metabolism and results in improved mitochondrial function and reduced oxidative stress. The reduction in lipid accumulation and liver injury resulted in decreased YAP/TAZ pathway activation, which may be involved in the reduced HCC progression in HKO livers. Together, these results suggest suppressing hepatic miR-33 may be an effective therapeutic approach to temper the development of NAFLD, NASH, and HCC in obesity
Authors
Pablo Fernández-Tussy, Magdalena P. Cardelo, Hanming Zhang, Jonathan Sun, Nathan L. Price, Nabil E. Boutagy, Leigh Goedeke, Martí Cadena-Sandoval, Chrysovalantou E. Xirouchaki, Wendy A. Brown, Xiaoyong Yang, Oscar Pastor-Rojo, Rebecca A. Haeusler, Anton M. Bennett, Tony Tiganis, Yajaira Suárez, Carlos Fernández-Hernando
Abstract
Fibrinogen-like protein 1 (FGL1) contributes to the proliferation and metabolism of hepatocytes; however, as a major ligand of the immune checkpoint, its role in the liver regional immune microenvironment is poorly understood. Hepatocytes specifically and highly expressed FGL1 under normal physiological conditions. Increases in hepatic CD8+ T and NK cell numbers and functions were found in Fgl1-deficient (Fgl1–/–) mice, but not in the spleen or lymph node, similar to findings in anti-FGL1 mAb–treated wild-type mice. Furthermore, Fgl1 deficiency or anti-FGL1 mAb blockade restrained liver metastasis and slowed the growth of orthotopic tumors, with significantly prolonged survival of tumor-bearing mice. Tumor-infiltrating hepatic CD8+ T and NK cells upregulated the expression of lymphocyte activation gene-3 (LAG-3) and exhibited stronger antitumor activities after anti-FGL1 treatment. The antitumor efficacy of FGL1 blockade depended on cytotoxic T lymphocytes and NK cells, demonstrated by using a cell-deficient mouse model and cell transfer in vivo. In vitro, FGL1 directly inhibited hepatic T and NK cells related to the receptor LAG-3. In conclusion, hepatocyte-derived FGL1 played critical immunoregulatory roles in the liver and contributed to liver metastasis and tumor growth by inhibiting CD8+ T and NK cell functions via the receptor LAG-3, providing a new strategy for liver cancer immunotherapy.
Authors
Fengjia Xi, Haoyu Sun, Hui Peng, Zhexiong Lian, Haiming Wei, Zhigang Tian, Rui Sun, Yongyan Chen
Abstract
Background Glycogen storage disease type IV (GSD IV) is an ultrarare autosomal recessive disorder that causes deficiency of functional glycogen branching enzyme and formation of abnormally structured glycogen termed polyglucosan. GSD IV has traditionally been categorized based on primary hepatic or neuromuscular involvement, with hepatic GSD IV subclassified as discrete subtypes: classic (progressive) and nonprogressive.Methods To better understand the progression of liver disease in GSD IV, we present clinical and histopathology data from 23 patients from around the world and characterized the liver involvement in the Gbe1ys/ys knockin mouse model.Results We propose an alternative to the established subtype-based terminology for characterizing liver disease in GSD IV and recognize 3 tiers of disease severity: (i) “severe progressive” liver disease, (ii) “intermediate progressive” liver disease, and (iii) “attenuated” liver disease. Analysis of liver pathology revealed that risk for liver failure cannot be predicted from liver biopsy findings alone in individuals affected by GSD IV. Moreover, analysis of postmortem liver pathology from an individual who died over 40 years after being diagnosed with nonprogressive hepatic GSD IV in childhood verified that liver fibrosis did not regress. Last, characterization of the liver involvement in a mouse model known to recapitulate the adult-onset neurodegenerative form of GSD IV (Gbe1ys/ys mouse model) demonstrated hepatic disease.Conclusion Our findings challenge the established subtype-based view of GSD IV and suggest that liver disease severity among patients with GSD IV represents a disease continuum.Trial registration ClinicalTrials.gov NCT02683512Funding None
Authors
Rebecca L. Koch, Bridget T. Kiely, Su Jin Choi, William R. Jeck, Leticia S. Flores, Vikrant Sood, Seema Alam, Gilda Porta, Katy LaVecchio, Claudia Soler-Alfonso, Priya S. Kishnani
Abstract
Portal hypertension (PHTN) is a severe complication of liver cirrhosis and is associated with intrahepatic sinusoidal remodeling induced by sinusoidal resistance and angiogenesis. Collagen type IV (COL4), a major component of basement membrane, forms in liver sinusoids upon chronic liver injury. However, the role, the cellular source and expression regulation of COL4 in liver diseases is unknown. Here, we examined how COL4 is produced and how it regulates sinusoidal remodeling in fibrosis and PHTN. Human cirrhotic liver sample RNA-sequencing showed increased COL4 expression, which was further confirmed via immunofluorescence staining. scRNA-sequencing identified liver sinusoidal endothelial cells (LSECs) as the predominant source of COL4 upregulation in mouse fibrotic liver. In addition, COL4 was upregulated in a tumor necrosis factor α–nuclear factor–κB dependent manner through an epigenetic mechanism in liver sinusoidal endothelial cells in vitro. Indeed, by utilizing a CRISPRi-dCas9-KRAB-mediated epigenome editing approach, epigenetic repression of the enhancer-promoter interaction showed silencing of COL4 gene expression. LSEC-specific COL4 gene mutation or repression in vivo abrogated sinusoidal resistance and angiogenesis, which thereby alleviated sinusoidal remodeling and PHTN. Our findings reveal that LSECs promote sinusoidal remodeling and PHTN during liver fibrosis through COL4 deposition.
Authors
Can Gan, Usman Yaqoob, Jianwen Lu, Man Xie, Abid A. Anwar, Nidhi Jalan-Sakrikar, Sofia Jerez, Tejasav Sehrawat, Amaia Navarro-Corcuera, Enis Kostallari, Nawras W. Habash, Sheng Cao, Vijay H. Shah
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