Gene Expression 17(1) Abstracts

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Gene Expression, Vol. 17, pp. 7–17, 2016
1052-2166/16 $90.00
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DOI: http://dx.doi.org/10.3727/105221616X691712
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Zinc Fingers and Homeoboxes 2 (Zhx2) Regulates Sexually Dimorphic Cyp Gene Expression in the Adult Mouse Liver

Kate Townsend Creasy,*1 Jieyun Jiang,† Hui Ren,† Martha L. Peterson,†‡ and Brett T. Spear†‡

*Department of Pharmacology and Nutritional Sciences, University of Kentucky, Lexington, KY, USA
†Department of Microbiology, Immunology and Molecular Genetics, University of Kentucky, Lexington, KY, USA
‡Markey Cancer Center, University of Kentucky, Lexington, KY, USA

The mammalian cytochrome P450 (Cyp) gene family encodes a large number of structurally related enzymes that catalyze a variety of metabolic and detoxification reactions. The liver is the primary site of Cyp expression in terms of expression levels and number of expressed genes, consistent with this organ’s essential role in metabolism of endogenous and xenobiotic compounds. Many Cyp genes exhibit sexually dimorphic expression. For example, Cyp2a4 is expressed significantly higher in the adult liver of female mice compared to male mice. An exception to this pattern is seen in BALB/cJ mice, where male hepatic Cyp2a4 mRNA levels are substantially elevated compared to male mice of other strains. The Zinc fingers and homeoboxes 2 (Zhx2) protein governs the silencing of several genes in the postnatal liver, including a-fetoprotein, H19, and glypican 3. Zhx2 also regulates numerous hepatic genes that govern lipid homeostasis. We previously showed that the Zhx2 gene is mutated in BALB/cJ mice, which led us to consider whether elevated male hepatic Cyp2a4 levels in this strain are due to this Zhx2 mutation. Using mice with a conditional Zhx2 deletion, we show here that the absence of Zhx2 in hepatocytes results in increased Cyp2a4 expression in adult male liver. We extend this finding to show that additional Cyp genes are disregulated in the absence of Zhx2. We also show that mRNA levels of Cyp2a4 and several other female-biased Cyp genes are increased, and male-biased Cyp4a12 is decreased in mouse liver tumors. These data indicate that Zhx2 is a novel regulator of sex-biased Cyp gene expression in the normal and diseased liver.

Key words: Transcription; Development; Hepatocyte; Regeneration; Knockout mice; Cancer

1Current address: Cardiovascular Research Institute, University of California, San Francisco, 555 Mission Bay Boulevard South, Room 282T, San Francisco, CA 94158, USA.
Address correspondence to Brett T. Spear, Department of Microbiology, Immunology and Molecular Genetics, University of Kentucky College of Medicine, 800 Rose Street, Lexington, KY 40536, USA. Tel: (859) 257-5167; Fax: (859) 257-8994; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Gene Expression, Vol. 17, pp. 19–34, 2016
1052-2166/16 $90.00
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DOI: http://dx.doi.org/10.3727/105221616X691631
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Thyroid Hormone Receptor β Agonist Induces β-Catenin-Dependent Hepatocyte Proliferation in Mice: Implications in Hepatic Regeneration

Tamara Feliciano Alvarado,* Elisabetta Puliga,†‡ Morgan Preziosi,† Minakshi Poddar,† Sucha Singh,† Amedeo Columbano,‡ Kari Nejak-Bowen,† and Satdarshan P. S. Monga†§

*Division of Gastroenterology, Department of Pediatrics, Children’s Hospital of Pittsburgh of UPMC, Pittsburgh, PA, USA
†Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
‡Department of Biomedical Sciences, University of Cagliari, Cagliari, Italy
§Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

Triiodothyronine (T3) induces hepatocyte proliferation in rodents. Recent work has shown molecular mechanism for T3’s mitogenic effect to be through activation of β-catenin signaling. Since systemic side effects of T3 may preclude its clinical use, and hepatocytes mostly express T3 hormone receptor β (TRβ), we investigated if selective TRβ agonists like GC-1 may also have β-catenin-dependent hepatocyte mitogenic effects. Here we studied the effect of GC-1 and T3 in conditional knockouts of various Wnt pathway components. We also assessed any regenerative advantage of T3 or GC-1 when given prior to partial hepatectomy in mice. Mice administered GC-1 showed increased pSer675-β-catenin, cyclin D1, BrdU incorporation, and PCNA. No abnormalities in liver function tests were noted. GC-1-injected liver-specific β-catenin knockouts (β-catenin LKO) showed decreased proliferation when compared to wild-type littermates. To address if Wnt signaling was required for T3- or GC-1-mediated hepatocyte proliferation, we used LRP5-6-LKO, which lacks the two redundant Wnt coreceptors. Surprisingly, decreased hepatocyte proliferation was also evident in LRP5-6-LKO in response to T3 and GC-1, despite increased pSer675-β-catenin. Further, increased levels of active β-catenin (hypophosphorylated at Ser33, Ser37, and Thr41) were evident after T3 and GC-1 treatment. Finally, mice pretreated with T3 or GC-1 for 7 days followed by partial hepatectomy showed a significant increase in hepatocyte proliferation both at the time (T0) and 24 h after surgery. In conclusion, like T3, TRb-selective agonists induce hepatocyte proliferation through β-catenin activation via both PKA- and Wnt-dependent mechanisms and confer a regenerative advantage following surgical resection. Hence, these agents may be useful regenerative therapies in liver transplantation or other surgical settings.

Key words: Wnt pathway; Regeneration; Hepatocyte proliferation; Liver transplantation

Address correspondence to Satdarshan P. S. Monga, M.D., Department of Gastroenterology, Hepatology and Nutrition, University of Pittsburgh School of Medicine, 200 Lothrop Street S-422 BST, Pittsburgh, PA 15261, USA. Tel: (412) 648-9966; Fax: (412) 648-1916; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Gene Expression, Vol. 17, pp. 35–45, 2016
1052-2166/16 $90.00
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DOI: http://dx.doi.org/10.3727/105221616X691578
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Liver-Specific Deletion of Integrin-Linked Kinase in Mice Attenuates Hepatotoxicity and Improves Liver Regeneration After Acetaminophen Overdose

Bharat Bhushan,* Genea Edwards,* Aishwarya Desai,* George K. Michalopoulos,† and Udayan Apte*

*Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA
†Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

Acetaminophen (APAP) overdose is the major cause of acute liver failure in the US. Prompt liver regeneration is critical for recovery after APAP hepatotoxicity, but mechanisms remain elusive. Extracellular matrix (ECM)-mediated signaling via integrin-linked kinase (ILK) regulates liver regeneration after surgical resection. However, the role of ECM signaling via ILK in APAP toxicity and compensatory regeneration is unknown, which was investigated in this study using liver-specific ILK knockout (KO) mice. ILK KO and wild-type (WT) mice were treated with 300 mg/kg APAP, and injury and regeneration were studied at 6 and 24 h after APAP treatment. ILK KO mice developed lower liver injury after APAP overdose, which was associated with decreased JNK activation (a key mediator of APAP toxicity). Further, higher glutathione levels after APAP treatment and lower APAP protein adducts levels, along with lower levels of CYP2E1, suggest decreased metabolic activation of APAP in ILK KO mice. Interestingly, despite lower injury, ILK KO mice had rapid and higher liver regeneration after APAP overdose accompanied with increased b-catenin signaling. In conclusion, liver-specific deletion of ILK improved regeneration, attenuated toxicity after APAP overdose, and decreased metabolic activation of APAP. Our study also indicates that ILK-mediated ECM signaling plays a role in the regulation of CYP2E1 and may affect toxicity of several centrilobular hepatotoxicants including APAP.

Key words: Extracellular matrix (ECM); Drug-induced liver injury; Hepatocyte proliferation; β-Catenin; Cytochrome P450 2E1

Address correspondence to Udayan Apte, Ph.D., DABT, Department of Pharmacology, Toxicology and Therapeutics, and Department of Cancer Biology, University of Kansas Medical Center, 3901 Rainbow Boulevard, MS1018, Kansas City, KS 66160, USA. Tel: (913) 588-9247; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Gene Expression, Vol. 17, pp. 47–59, 2016
1052-2166/16 $90.00
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DOI: http://dx.doi.org/10.3727/105221616X692135
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Enhanced Steatosis and Fibrosis in Liver of Adult Offspring Exposed to Maternal High-Fat Diet

Michael D. Thompson,* Mary J. Cismowski,† Aaron J. Trask,† Scott W. Lallier,‡ Amanda E. Graf,‡ Lynette K. Rogers,‡ Pamela A. Lucchesi,†§ and David R. Brigstock

*Division of Endocrinology, Nationwide Children’s Hospital, Columbus, OH, USA
†Center for Cardiovascular and Pulmonary Research, Nationwide Children’s Hospital, Columbus, OH, USA
‡Center for Perinatal Research, Nationwide Children’s Hospital, Columbus, OH, USA
§Department of Basic Sciences, The Commonwealth Medical College, Scranton, PA, USA
¶Center for Clinical and Translational Research, Nationwide Children’s Hospital, Columbus, OH, USA

Early life exposures can increase the risk of developing chronic diseases including nonalcoholic fatty liver disease. Maternal high-fat diet increases susceptibility to development of steatosis in the offspring. We determined the effect of maternal high-fat diet exposure in utero and during lactation on offspring liver histopathology, particularly fibrosis. Female C57Bl/6J mice were fed a control or high-fat diet (HFD) for 8 weeks and bred with lean males. Nursing dams were continued on the same diet with offspring sacrificed during the perinatal period or maintained on either control or high-fat diet for 12 weeks. Increased hepatocyte proliferation and stellate cell activation were observed in the liver of HFD-exposed pups. Offspring exposed to perinatal high-fat diet and high-fat diet postweaning showed extensive hepatosteatosis compared to offspring on high-fat diet after perinatal control diet. Offspring exposed to perinatal high-fat diet and then placed on control diet for 12 weeks developed steatosis and pericellular fibrosis. Importantly, we found that exposure to perinatal high-fat diet unexpectedly promotes more rapid disease progression of nonalcoholic fatty liver disease, with a sustained fibrotic phenotype, only in adult offspring fed a postweaning control diet.

Key words: Developmental origins of disease; Fatty liver; High-fat diet (HFD); Fibrosis; Stellate cell activation

Address correspondence to Michael D. Thompson, M.D., Ph.D., Division of Endocrinology, Nationwide Children’s Hospital, 700 Children’s Drive, Columbus, OH 43205, USA. Tel: (614) 722-3153; Fax: (614) 722-4440; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Gene Expression, Vol. 17, pp. 61–77, 2016
1052-2166/16 $90.00
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DOI: http://dx.doi.org/10.3727/105221616X691730
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

Myeloid Mixed Lineage Kinase 3 Contributes to Chronic Ethanol-Induced Inflammation and Hepatocyte Injury in Mice

Rebecca L. McCullough,* Paramananda Saikia,* Katherine A. Pollard,* Megan R. McMullen,* Laura E. Nagy,*†‡ and Sanjoy Roychowdhury*‡

*Department of Pathobiology, Center for Liver Disease Research, Cleveland Clinic, Cleveland, OH, USA
†Department of Gastroenterology, Center for Liver Disease Research, Cleveland Clinic, Cleveland, OH, USA
‡Department of Molecular Medicine, Case Western Reserve University, Cleveland, OH, USA

Proinflammatory activity of hepatic macrophages plays a key role during progression of alcoholic liver disease (ALD). Since mixed lineage kinase 3 (MLK3)-dependent phosphorylation of JNK is involved in the activation of macrophages, we tested the hypothesis that myeloid MLK3 contributes to chronic ethanol-induced inflammatory responses in liver, leading to hepatocyte injury and cell death. Primary cultures of Kupffer cells, as well in vivo chronic ethanol feeding, were used to interrogate the role of MLK3 in the progression of liver injury. Phosphorylation of MLK3 was increased in primary cultures of Kupffer cells isolated from ethanol-fed rats compared to cells from pair-fed rats. Kupffer cells from ethanol-fed rats were more sensitive to LPS
stimulated cytokine production; this sensitization was normalized by pharmacological inhibition of MLK3. Chronic ethanol feeding to mice increased MLK3 phosphorylation robustly in F4/80+ Kupffer cells, as well as in isolated nonparenchymal cells. MLK3−/− mice were protected from chronic ethanol-induced phosphorylation of MLK3 and JNK, as well as multiple indicators of liver injury, including increased ALT/AST, inflammatory cytokines, and induction of RIP3. However, ethanol-induced steatosis and hepatocyte apoptosis were not affected by MLK3. Finally, chimeric mice lacking MLK3 only in myeloid cells were also protected from chronic ethanol-induced phosphorylation of JNK, expression of inflammatory cytokines, and increased ALT/AST. MLK3 expression in myeloid cells contributes to phosphorylation of JNK, increased cytokine production, and hepatocyte injury in response to chronic ethanol. Our data suggest that myeloid MLK3 could be targeted for developing potential therapeutic strategies to suppress liver injury in ALD patients.

Key words: Alcoholic liver disease (ALD); Kupffer cells; Necroptosis; Toll-like receptor 4 (TLR4); Cytokines

Address correspondence to Laura E. Nagy, Cleveland Clinic Foundation, Lerner Research Institute/NE40, 9500 Euclid Avenue, Cleveland, OH 44195, USA. Tel: 216-444-4120; Fax: 216-636-1493; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it  or Sanjoy Roychowdhury, Cleveland Clinic Foundation, Lerner Research Institute/NE40, 9500 Euclid Avenue, Cleveland, OH 44195, USA. Tel: 216-444-4120; Fax: 216-636-1493; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it


Gene Expression, Vol. 17, pp. 79–88, 2016
1052-2166/16 $90.00
+ .00
DOI: http://dx.doi.org/10.3727/105221616X691604
E-ISSN 1555-3884
Copyright ©
2016 Cognizant Comm. Corp.
Printed in the USA. All rights reserved

FAK Kinase Activity Is Required for the Progression of c-MET/β-Catenin-Driven Hepataocellular Carcinoma

Na Shang,* Maribel Arteaga,* Ali Zaidi,* Scott J. Cotler,† Peter Breslin,‡ Xianzhong Ding,§ Paul Kuo,* Michael Nishimura,* Jiwang Zhang,§ and Wei Qiu*

*Department of Surgery and Oncology Institute, Loyola University Chicago Stritch School of Medicine, Maywood, IL, USA
†Department of Medicine, Loyola University Chicago Stritch School of Medicine, Maywood, IL, USA
‡Department of Molecular/Cellular Physiology, Loyola University Chicago Stritch School of Medicine, Maywood, IL, USA
§Department of Pathology, Loyola University Chicago Stritch School of Medicine, Maywood, IL, USA

There is an urgent need to develop new and more effective therapeutic strategies and agents to treat hepatocellular carcinoma (HCC). We have recently found that deletion of Fak in hepatocytes before tumors form inhibits tumor development and prolongs survival of animals in a c-MET (MET)/β-catenin (CAT)-driven HCC mouse model. However, it has yet to be determined whether FAK expression in hepatocytes promotes MET/CAT-induced HCC progression after tumor initiation. In addition, it remains unclear whether FAK promotes HCC development through its kinase activity. We generated hepatocyte-specific inducible Fak-deficient mice (Alb-creERT2; Fakflox/flox) to examine the role of FAK in HCC progression. We reexpressed wild-type and mutant FAK in Fak-deficient mice to determine FAK’s kinase activity in HCC development. We also examined the efficacy of a FAK kinase inhibitor PF-562271 on HCC inhibition. We found that deletion of Fak after tumors form significantly repressed MET/CAT-induced tumor progression. Ectopic FAK expression restored HCC formation in hepatocyte-specific Fak-deficient mice. However, overexpression of a FAK kinase-dead mutant led to reduced tumor load compared to mice that express wild-type FAK. Furthermore, PF-562271 significantly suppressed progression of MET/CAT-induced HCC. Fak kinase activity is important for MET/CAT-induced HCC progression. Inhibiting FAK kinase activity provides a potential therapeutic strategy to treat HCC.

Key words: Hepatocellular carcinoma (HCC); Sleeping Beauty Transposon system; Proliferation; AKT; Extracellular receptor kinase (ERK)

Address correspondence to Wei Qiu, Ph.D., Department of Surgery and Oncology Institute, Loyola University Chicago Stritch School of Medicine, 2160 South 1st Avenue, Building 112 Room 338, Maywood, IL 60153, USA. Tel: 708-327-8191; Fax: 708-327-3342l; E-mail: This e-mail address is being protected from spambots. You need JavaScript enabled to view it