• HBV-related hepatocellular carcinoma
• hepatitis B virus
• liver
• liver regeneration

• Animals
• Chromatin Assembly and Disassembly
• Epigenesis, Genetic
• Humans
• Liver/cytology
• Liver/metabolism
• Liver Diseases/genetics
• Liver Diseases/pathology
• Liver Regeneration

The liver is a unique organ that has a phenomenal capacity to regenerate after injury. Different surgical procedures, including partial hepatectomy (PH), intraoperative portal vein ligation (PVL), and associated liver partition and portal vein ligation for staged hepatectomy (ALPPS) show clinically distinct recovery patterns and regeneration. The observable clinical differences likely mirror some underlying variations in the patterns of gene activation and regeneration pathways. In this study, we provided a comprehensive comparative transcriptomic analysis of gene regulation in regenerating rat livers temporally spaced at 24 h and 96 h after PH, PVL, and ALPPS. The time-dependent factors appear to be the most important determinant of post-injury alterations of gene expression in liver regeneration. Gene expression profile after ALPPS showed more similar expression pattern to the PH than the PVL at the early phase of the regeneration. Early transcriptomic changes and predicted upstream regulators that were found in all three procedures included cell cycle associated genes (E2F1, CCND1, FOXM1, TP53, and RB1), transcription factors (Myc, E2F1, TBX2, FOXM1), DNA replication regulators (CDKN1A, EZH2, RRM2), G1/S-transition regulators (CCNB1, CCND1, RABL6), cytokines and growth factors (CSF2, IL-6, TNF, HGF, VEGF, and EGF), ATM and p53 signaling pathways. The functional pathway, upstream, and network analyses revealed both unique and overlapping molecular mechanisms and pathways for each surgical procedure. Identification of molecular signatures and regenerative signaling pathways for each surgical procedure further our understanding of key regulators of liver regeneration as well as patient populations that are likely to benefit from each procedure.

To study the anti-inflammatory and liver regenerative effects of adipose-derived mesenchymal stem cells (ADSCs) on a porcine model of ischemia-reperfusion (IR) and hemihepatectomy.

Eighteen healthy Bama miniature pigs were randomly divided into the sham-operated (sham), untreated IR injury (IRI), and ADSC-transplanted (ADSC) groups. Hepatic IR was established by laparoscopic hemihepatectomy. ADSCs were transplanted directly into the liver parenchyma after the surgery. Hepatic inflammation and liver regeneration were evaluated by histopathological examination and assessment of relevant cytokines and other factors.

ADSC transplantation successfully ameliorated the IRI-induced histopathological damage and the high levels of pro-inflammatory cytokines like IL-1β, IL-6, and TNF-α. In addition, the ADSCs enhanced the expression of the anti-inflammatory IL-10, regenerative factors including HGF, Cyclin D1, and proliferating cell nuclear antigen (PCNA), and angiogenic factors like VEGF, ANG-1, and ANG-2.

ADSCs attenuated the hepatic IRI-induced inflammatory response and promoted liver regeneration.

• anti-oxidative stress
• cytokine
• liver regeneration
• partial hepatectomy
• placenta

• DNA methylation
• H3K27me3
• UHRF1
• epigenetic compensation
• epigenomics
• liver biology
• partial hepatectomy
• tissue regeneration
• transposons

• Animals
• CCAAT-Enhancer-Binding Proteins/physiology
• DNA Methylation
• Epigenomics
• Gene Expression Profiling
• Gene Expression Regulation
• Hepatocytes/cytology
• Hepatocytes/physiology
• Liver Regeneration
• Male
• Mice
• Mice, Knockout
• Ubiquitin-Protein Ligases/physiology

• R01 CA154683 NCI NIH HHS
• R01 DK080789 NIDDK NIH HHS
• R37 CA230636 NCI NIH HHS
• S10 OD018522 NIH HHS

• Adaptive lasso
• Gene selection
• Group lasso
• Rat hepatocyte proliferation
• Weighted gene co-expression network

• ALT
• IL6
• RT-PCR
• TNFα
• monoterpenoid alcohol

Liver regeneration after partial hepatectomy (PHx) is a time-dependent process, which is tightly regulated by multiple signaling cascades. Failure of this complex process leads to posthepatectomy liver failure (PHLF), which is associated with a high rate of mortality. Thus, it is extremely important to establish a useful biomarker of liver regeneration to help prevent PHLF. Here, we hypothesized that alterations in the plasma peptide profile may predict liver regeneration following PHx and hence we set up a diagnostic platform for monitoring posthepatectomy outcome. We chronologically analyzed plasma peptidomic profiles of 5 partially hepatectomized microminipigs using the ClinProt^TM system, which consists of magnetic beads and MALDI-TOF/TOF MS. We identified endogenous circulating peptides specific to each phase of the postoperative course after PHx in pigs. Notably, peptide fragments of histones were detected immediately after PHx; the presence of these fragments may trigger liver regeneration in the very acute phase after PHx. An N-terminal fragment of hemoglobin subunit α (3627 m/z) was detected as an acute-phase-specific peptide. In the recovery phase, the short N-terminal fragments of albumin (3028, 3042 m/z) were decreased, whereas the long N-terminal fragment of the protein (8926 m/z) was increased. To further validate and extract phase-specific biomarkers using plasma peptidome after PHx, plasma specimens of 4 patients who underwent PHx were analyzed using the same method as we applied to pigs. It revealed that there was also phase-specificity in peptide profiles, one of which was represented by a fragment of complement C4b (2378 m/z). The strategy described herein is highly efficient for the identification and characterization of peptide biomarkers of liver regeneration in a swine PHx model. This strategy is feasible for application to human biomarker studies and will yield clues for understanding liver regeneration in human clinical trials.

Liver biology and function, drug-induced liver injury (DILI) and liver diseases are difficult to study using current in vitro models such as primary human hepatocyte (PHH) monolayer cultures, as their rapid de-differentiation restricts their usefulness substantially. Thus, we have developed and extensively characterized an easily scalable 3D PHH spheroid system in chemically-defined, serum-free conditions. Using whole proteome analyses, we found that PHH spheroids cultured this way were similar to the liver in vivo and even retained their inter-individual variability. Furthermore, PHH spheroids remained phenotypically stable and retained morphology, viability, and hepatocyte-specific functions for culture periods of at least 5 weeks. We show that under chronic exposure, the sensitivity of the hepatocytes drastically increased and toxicity of a set of hepatotoxins was detected at clinically relevant concentrations. An interesting example was the chronic toxicity of fialuridine for which hepatotoxicity was mimicked after repeated-dosing in the PHH spheroid model, not possible to detect using previous in vitro systems. Additionally, we provide proof-of-principle that PHH spheroids can reflect liver pathologies such as cholestasis, steatosis and viral hepatitis. Combined, our results demonstrate that the PHH spheroid system presented here constitutes a versatile and promising in vitro system to study liver function, liver diseases, drug targets and long-term DILI.

• Bioinformatics
• Gene profiles
• Liver
• Liver cirrhosis
• Microarray analysis

• Animals
• Binding Sites
• Computational Biology
• Ethanol/pharmacology
• Gene Expression Regulation
• Genome
• Hepatectomy
• Liver Regeneration/drug effects
• Liver Regeneration/genetics
• NF-kappa B/metabolism
• Promoter Regions, Genetic
• Protein Binding/drug effects
• Rats, Sprague-Dawley

• R01 AA018873 NIAAA NIH HHS
• R21 AA016919 NIAAA NIH HHS
• R21 AA022417 NIAAA NIH HHS
• T32 AA007463 NIAAA NIH HHS

Wangensteen et al. employed a parallel screen to test the impact of 43 selected genes on liver repopulation in the Fah^−/− mouse model of hereditary tyrosinemia. The transcription factor Foxa3 was a strong promoter of liver regeneration, while tumor necrosis factor receptor 1 (TNFR1) was the most significant suppressor of repopulation among all of the genes tested.

The fundamental question of which genes are most important in controlling liver regeneration remains unanswered. We employed a parallel screen to test the impact of 43 selected genes on liver repopulation in the Fah^−/− mouse model of hereditary tyrosinemia. We discovered that the transcription factor Foxa3 was a strong promoter of liver regeneration, while tumor necrosis factor receptor 1 (TNFR1) was the most significant suppressor of repopulation among all of the genes tested. Our approach enabled the identification of these factors as important regulators of liver repopulation and potential drug targets for the promotion of liver repopulation.

• Foxa3
• TNFR1
• injury
• liver
• regeneration
• repopulation

• rat liver regeneration
• erythropoietin
• rat genome 230 2.0 microarray
• gene expression profiles
• gene synergetic effect

• Algorithms
• Animals
• Cell Proliferation/genetics
• Cells, Cultured
• Erythropoietin/genetics
• Extracellular Signal-Regulated MAP Kinases/genetics
• Gene Expression Profiling
• Gene Regulatory Networks
• Hepatectomy/methods
• Hepatocytes/metabolism
• Liver Regeneration/genetics
• Male
• Models, Genetic
• Oligonucleotide Array Sequence Analysis
• Phosphatidylinositol 3-Kinases/genetics
• Proto-Oncogene Proteins c-akt/genetics
• Rats, Sprague-Dawley
• Reverse Transcriptase Polymerase Chain Reaction
• Signal Transduction/genetics
• Time Factors
• Transcription Factors/genetics
• ras Proteins/genetics

• EGR1
• GRO1
• massive hepatectomy
• microarray
• splenectomy

The recovery of liver mass is mainly mediated by proliferation of hepatocytes after 2/3 partial hepatectomy (PH) in rats. Studying the gene expression profiles of hepatocytes after 2/3 PH will be helpful to investigate the molecular mechanisms of liver regeneration (LR). We report here the first application of weighted gene co-expression network analysis (WGCNA) to analyze the biological implications of gene expression changes associated with LR. WGCNA identifies 12 specific gene modules and some hub genes from hepatocytes genome-scale microarray data in rat LR. The results suggest that upregulated MCM5 may promote hepatocytes proliferation during LR; BCL3 may play an important role by activating or inhibiting NF-kB pathway; MAPK9 may play a permissible role in DNA replication by p38 MAPK inactivation in hepatocytes proliferation stage. Thus, WGCNA can provide novel insight into understanding the molecular mechanisms of LR.

• Animals
• Gene Expression Profiling
• Gene Regulatory Networks
• Hepatectomy
• Hepatocytes/cytology
• Hepatocytes/metabolism
• Liver Regeneration/genetics
• Molecular Sequence Annotation
• Rats
• Rats, Sprague-Dawley
• Signal Transduction/genetics

• early growth response 1
• heat shock protein
• heme oxygenase-1
• hepatectomy
• liver regeneration
• microarray
• reperfusion injury
• splenectomy

AIM: To investigate the effect of lentiviral-mediated delivery of shRNA targeting the SMAD3 gene on liver regeneration in rats.

METHODS: Sixty male rats were randomly divided into three groups: SMAD3 shRNA group (n = 20), shRNA control group (n = 20), and normal saline group (n = 20). Intrasplenic injection of lentivirus carrying the SMAD3 shRNA or control shRNA at a dose of 1.0 × 10⁸ TU per rat was performed in rats of the SMAD3 shRNA group and shRNA control group, respectively, while the normal saline group was given equal volume of normal saline. After 96 h, 2/3 partial hepatectomy was performed to develop a rat model of liver regeneration. Seven rats of each group were sacrificed at 96 h, and the rest were sacrificed at 144 h after hepatectomy. Liver tissue specimens were collected. SMAD3 expression was detected using real-time PCR and immunohistochemistry. Proliferation of hepatocytes was evaluated by detecting Ki67 by immunohistochemistry. The ratio of liver weight to body weight was determined to observe its role in liver regeneration in vivo.

RESULTS: At 96 h and 144 h after hepatectomy, SMAD3 mRNA expression decreased by 73% and 63% in the SMAD3 shRNA group compared to the shRNA control group. The expression of SMAD3 protein also decreased obviously. More active proliferation of hepatocytes was observed after SMAD3 down-regulation. The liver weight/body weight in the SMAD3 shRNA group was higher than those in the normal saline control group and shRNA control group (96 h: 4.50 ± 0.43 vs 3.97 ± 0.55 vs 3.98 ± 0.40, 144 h: 4.66 ± 0.54 vs 4.15 ± 0.51 vs 4.20 ± 0.34), but there was no statistically significance between them (all P > 0.05).

CONCLUSION: Down-regulation of SMAD3 expression could moderately promote liver regeneration in rats.

• SMAD3
• ShRNA
• Lentiviral vector
• Liver regeneration

• microrna
• alcohol
• partial hepatectomy
• gene expression

• integrin signaling pathway
• rat
• liver regeneration
• partial hepatectomy
• immunochemistry
• hepatocyte proliferation
• rat genome 230 2.0 array
• rt-pcr
• gene expression profile
• gene synergy value

• Animals
• Computational Biology
• Gene Expression Profiling
• Hepatocytes/metabolism
• Integrins/genetics
• Integrins/metabolism
• Liver Regeneration
• Oligonucleotide Array Sequence Analysis
• Rats
• Rats, Sprague-Dawley
• Signal Transduction

• sympathectomy combining partial hepatectomy (sph)
• vagotomy combining partial hepatectomy (vph)
• total liver denervation combining partial hepatectomy (tdph)
• rat liver regeneration (lr)
• gene expression profiles
• biological processes

• Animals
• Autonomic Nervous System/metabolism
• Biological Phenomena/genetics
• Denervation
• Gene Expression Profiling
• Gene Expression Regulation
• Hepatectomy
• Liver Regeneration/genetics
• Liver Regeneration/physiology
• Models, Animal
• Oligonucleotide Array Sequence Analysis
• Rats
• Rats, Sprague-Dawley
• Reproducibility of Results
• Reverse Transcriptase Polymerase Chain Reaction

• Animals
• Gene Expression Profiling
• Hepatectomy
• Hepatocytes/metabolism
• Liver/cytology
• Liver/physiology
• Polymerase Chain Reaction
• Rats
• Rats, Sprague-Dawley
• Regeneration

• hpa, hypothalamic–pituitary–adrenal
• acth, adrenocorticotropic hormone
• 3αhsd, 3α-hydroxysteroid dehydrogenase
• fxr, farnesoid x receptor
• cyp7a1, cholesterol 7α-hydroxylase
• 11βhsd, 11β-hydroxysteroid dehydrogenase
• bdl, bile duct ligation
• thb, tetrahydrocorticosterone
• dhb, dihydrocorticosterone
• gcms, gas chromatography mass spectrometry
• cdca, chenodeoxycholic acid
• ca, cholic acid
• dca, deoxycholic acid
• gcdca, glyco-cdca
• k_i, inhibitor constant
• tcdca, tauro-cdca
• dmem, dulbecco’s modified eagle’s medium
• pcr, polymerase chain reaction
• ff, fat-free
• ecrp, endoscopic retrograde cholangiopancreatography
• anova, analysis of variance
• alt, alanine transaminase
• alp, alkaline phosphatase
• cyp11b1, 11β-hydroxylase
• nefa, non-esterified fatty acids
• sem, standard error of mean
• bile acid
• glucocorticoid
• 5β-reductase
• adrenal
• jaundice

• 3-Hydroxysteroid Dehydrogenases/genetics
• Animals
• Base Sequence
• Bile Acids and Salts/pharmacology
• Bile Acids and Salts/therapeutic use
• Bile Ducts/physiology
• Cytosol/enzymology
• Female
• Glucocorticoids/metabolism
• Humans
• Hydrocortisone/metabolism
• Hypothalamo-Hypophyseal System/drug effects
• Hypothalamo-Hypophyseal System/physiology
• Jaundice, Obstructive/drug therapy
• Jaundice, Obstructive/metabolism
• Jaundice, Obstructive/urine
• Kinetics
• Ligation
• Liver/enzymology
• Male
• Pituitary-Adrenal System/drug effects
• Pituitary-Adrenal System/physiology
• Promoter Regions, Genetic/drug effects
• Promoter Regions, Genetic/genetics
• Rats
• Rats, Wistar
• Tetrahydrocortisol/urine
• Transcription, Genetic/drug effects

• Wellcome Trust
• G0400874 Medical Research Council
• British Heart Foundation

Although 70% (or 2/3) partial hepatectomy (PH) is the most studied model for liver regeneration, the hepatic protein expression profile associated with lower volume liver resection (such as 50% PH) has not yet been reported. Therefore, the aim of this study was to determine the global protein expression profile of the regenerating mouse liver following 50% PH by differential proteomics, and thereby gaining some insights into the hepatic regeneration mechanism(s) under this milder but clinically more relevant condition.

Proteins from sham-operated mouse livers and livers regenerating for 24 h after 50% PH were separated by SDS-PAGE and analyzed by nanoUPLC-Q-Tof mass spectrometry. Compared to sham-operated group, there were totally 87 differentially expressed proteins (with 50 up-regulated and 37 down-regulated ones) identified in the regenerating mouse livers, most of which have not been previously related to liver regeneration. Remarkably, over 25 differentially expressed proteins were located at mitochondria. Several of the mitochondria-resident proteins which play important roles in citric acid cycle, oxidative phosphorylation and ATP production were found to be down-regulated, consistent with the recently-proposed model in which the reduction of ATP content in the remnant liver gives rise to early stress signals that contribute to the onset of liver regeneration. Pathway analysis revealed a central role of c-Myc in the regulation of liver regeneration.

Our study provides novel evidence for mitochondria as a pivotal organelle that is connected to liver regeneration, and lays the foundation for further studies on key factors and pathways involved in liver regeneration following 50% PH, a condition frequently used for partial liver transplantation and conservative liver resection.

AIM: To investigate the molecular mechanisms involved in coagulation factor expression and/or function during direct hyperplasia (DH)-mediated liver regeneration.

METHODS: Direct hyperplasia-mediated liver regeneration was induced in female C57BL/6 mice by administering 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), a representative hepatomitogen. Mice were weighed and sacrificed at various time points [Day 0 (D0: prior to injection), 3 h, D1, D2, D3, and D10] after TCPOBOP administration to obtain liver and blood samples. Using the RNA samples extracted from the liver, a comprehensive analysis was performed on the hepatic gene expression profiling of coagulation-related factors by real-time RT-PCR (fibrinogen, prothrombin, factors V, VII, VIII, IX, X, XI, XII, XIIIβ, plasminogen, antithrombin, protein C, protein S, ADAMTS13, and VWF). The corresponding plasma levels of coagulation factors (fibrinogen, prothrombin, factors V, VII, VIII, IX, X, XI, XII, XIII, and VWF) were also analyzed and compared with their mRNA levels.

RESULTS: Gavage administration of TCPOBOP (3 mg/kg body weight) resulted in a marked and gradual increase in the weight of the mouse livers relative to the total body weight to 220% by D10 relative to the D0 (control) ratios. At the peak of liver regeneration (D1 and D2), the gene expression levels for most of the coagulation-related factors (fibrinogen, prothrombin, factors V, VII, VIII, IX, XI, XII, XIIIβ, plasminogen, antithrombin, protein C, ADAMTS13, VWF) were found to be down-regulated in a time-dependent manner, and gradually recovered by D10 to the basal levels. Only mRNA levels of factor X and protein S failed to show any decrease during the regenerative phase. As for the plasma levels, 5 clotting factors (prothrombin, factors VIII, IX, XI, and XII) demonstrated a significant decrease (P < 0.05) during the regeneration phase compared with D0. Among these 5 factors, factor IX and factor XI showed the most dramatic decline in their activities by about 50% at D2 compared to the basal levels, and these reductions in plasma activity for both factors were consistent with our RT-PCR findings. In contrast, the plasma activities of the other coagulation factors (fibrinogen, factors V, VII, XIII, and VWF) were not significantly reduced, despite the reduction in the liver mRNA levels. Unlike the other factors, FX showed a temporal increase in its plasma activity, with significant increases (P < 0.05) detected at D1.

CONCLUSION: Investigating the coagulation cascade protein profiles during liver regeneration by DH may help to better understand the basic biology of the liver under normal and pathological conditions.

• Coagulation factor
• 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene
• Direct hyperplasia
• Liver regeneration

• Adenoviridae/genetics
• Animals
• Cell Proliferation
• Fibroblast Growth Factors/genetics
• Fibroblast Growth Factors/metabolism
• Follistatin/biosynthesis
• Follistatin/genetics
• Follistatin/metabolism
• Gene Expression Profiling/methods
• Gene Expression Regulation
• Gene Transfer Techniques
• Growth Differentiation Factor 15/genetics
• Growth Differentiation Factor 15/metabolism
• Liver/metabolism
• Liver/pathology
• Male
• Oligonucleotide Array Sequence Analysis
• Organ Size
• RNA, Messenger/metabolism
• Rats
• Rats, Sprague-Dawley
• Reverse Transcriptase Polymerase Chain Reaction
• Time Factors