Expression Of Hepatic Transcription Factors Research Articles

PAS Kinase: A Nutrient and Energy Sensor "Master Key" in the Response to Fasting/Feeding Conditions.

The protein kinase with PAS domains (PASK) is a nutrient and energy sensor located in the cells of multiple organs. Many of the recent findings for understanding PASK functions in mammals have been reported in studies involving PASK-deficient mice. This minireview summarizes the PASK role in the control of fasting and feeding responses, focusing especially on the hypothalamus and liver. In 2013, PASK was identified in the hypothalamic areas involved in feeding behavior, and its expression was regulated under fasting/refeeding conditions. Furthermore, it plays a role in coordinating the activation/inactivation of the hypothalamic energy sensors AMPK and mTOR/S6K1 pathways in response to fasting. On the other hand, PASK deficiency prevents the development of obesity and non-alcoholic fatty liver in mice fed with a high-fat diet. This protection is explained by the re-establishment of several high-fat diet metabolic alterations produced in the expression of hepatic transcription factors and key enzymes that control the main metabolic pathways involved in maintaining metabolic homeostasis in fasting/feeding responses. This minireview covers the effects of PASK inactivation in the expression of certain transcription factors and target enzymes in several metabolic pathways under situations such as fasting and feeding with either a standard or a high-fat diet.

A Multiple-Cell Microenvironment in a 3-Dimensional System Enhances Direct Cellular Reprogramming Into Hepatic Organoids

ObjectivesThe difficulty in proliferation and availability and the rapid loss functions of primary human hepatocytes highlight the need to develop an alternative, preferably renewable source of human induced hepatocytes in regenerative medicine. Liver organoids generated on a multiple-cell microenvironment in a 3-dimensional (3D) system can provide a highly efficient solution to this issue. MethodsHuman hepatocytes were induced from fibroblasts by the lentiviral expression of FOXA3, HNF1A, and HNF4A. Together with these induced hepatocytes, human umbilical vein endothelial cells and mesenchymal stem cells in a 3D system were used to produce liver organoids. Liver-related gene and protein expression of liver organoids and induced hepatocytes were tested using a 2-dimensional (2D) system. ResultsLiver organoids notably increased the expression of hepatic transcription factors, marker genes, transporter genes, and liver metabolism enzyme genes, while it decreased the specific gene expression of fibroblasts. Liver organoids expressed comparable liver-specific proteins, such as ALB, AAT, and HNF4A in the 3D system. ConclusionDirect reprogramming in multiple-cell microenvironments in 3D systems is more controllable and efficient than cell reprogramming in 2D systems. Liver organoids have the potential for use in disease modeling, pharmaceutical applications, and cellular transplantation.

Histone Modifications Regulate the Developmental Expression of Human Hepatic UDP-Glucuronosyltransferase 1A1.

Human UDP-glucuronosyltransferase 1A1 (UGT1A1) is a unique enzyme involved in bilirubin conjugation. We previously characterized the hepatic expression of transcription factors affecting UGT1A1 expression during development. Accordingly, in this study, we characterized the ontogenetic expression of hepatic UGT1A1 from the perspective of epigenetic regulation. We observed significant histone-3-lysine-4 dimethylation (H3K4me2) enrichment in the adult liver and histone-3-lysine-27 trimethylation (H3K27me3) enrichment in the fetal liver, indicating that dynamic alterations of histone methylation were associated with ontogenetic UGT1A1 expression. We further showed that the transcription factor hepatocyte nuclear factor 1α (HNF1A) affects histone modifications around the UGT1A1 locus. In particular, we demonstrated that by recruiting HNF1A the cofactors mixed-lineage leukemia 1, the transcriptional coactivator p300, and nuclear receptor coactivator 6 aggregate at the UGT1A1 promoter, thereby regulating histone modifications and subsequent UGT1A1 expression. In this study, we proposed new ideas for the developmental regulation of metabolic enzymes via histone modifications, and our findings will potentially contribute to the development of age-specific therapies.

Hepatic expression of transcription factors affecting developmental regulation of UGT1A1 in the Han Chinese population.

Complete or partial inactivity of UGT1A1, the unique enzyme responsible for bilirubin glucuronidation, is commonly associated with hyperbilirubinemia. We investigated the dynamic expression of UGT1A1, and that of the transcription factors (TFs) involved in its developmental regulation, during human hepatic growth in Han Chinese individuals. Eighty-eight prenatal, pediatric, and adult liver samples were obtained from Han Chinese individuals. Quantitative real-time polymerase chain reaction was used to evaluate mRNA expression of UGT1A1 and TFs including PXR, CAR, HNF1A, HNF4A, PPARA, etc. UGT1A1 protein levels and metabolic activity were determined by western blotting and high-performance liquid chromatography. Direct sequencing was employed to genotype UGT1A1*6 (211G˃A) and UGT1A1*28 (TA6˃TA7) polymorphisms. UGT1A1 expression was minimal in prenatal samples, but significantly elevated during pediatric and adult stages. mRNA and protein levels and metabolic activity were prominently increased (120-, 20-, and 10-fold, respectively) in pediatric and adult livers compared to prenatal samples. Furthermore, expression did not differ appreciably between pediatric and adult periods. Dynamic expression of TFs, including PXR, CAR, HNF1A, HNF4A, and PPARA, was consistent with UGT1A1 levels at each developmental stage. A pronounced correlation between expression of these TFs and that of UGT1A1 (P < 0.001) was observed. Moreover, UGT1A1*6 and UGT1A1*28 polymorphisms reduced levels of UGT1A1 by up to 40-60%. Hepatic expression of transcription factors is associated with developmental regulation of UGT1A1 in the Han Chinese population. Moreover, UGT1A1 polymorphisms are associated with reduced expression of UGT1A1 mRNA and protein, as well as enzyme activity.

Evaluation of a new standardized enzymatic isolation protocol for human umbilical cord-derived stem cells

The umbilical cord (UC) represents an important source of mesenchymal stem cells (MSC). These human UC-derived MSC (UC–MSC) have already been isolated using a protocol based on the migratory and plastic adhesive properties of MSC (UC–MSC–Mig). The UC–MSC–Mig isolation method, however, is difficult to standardize. Therefore, we developed an enzymatic isolation protocol (UC–MSC–Enz) to overcome the above mentioned disadvantages. First, we investigated the UC–MSC–Enz for their MSC properties. We found that UC–MSC–Enz express the MSC markers CD73, CD90 and CD105 and are able to differentiate into osteoblasts, adipocytes and chondroblasts fulfilling the MSC criteria of the International Society for Cellular Therapy. Previously we found that UC–MSC–Mig are unique among MSCs due to their significant expression of several hepatic (progenitor) markers. Therefore, we also investigated the expression of hepatic transcription factors and other hepatic markers in UC–MSC–Enz at both the mRNA and protein level. We found that the expression of hepatic transcription factors (GATA4, GATA6, SOX9 and SOX17) and hepatic markers (AFP, DPP4, CX43, DKK1, DSG2, KRT18 and KRT19) in UC–MSC–Enz was not significantly different from those of UC–MSC–Mig. Consequently, this optimized enzyme-based method represents a fast, robust and standardized way to isolate UC–MSC for a broad range of applications.

Differential Regulation of Hepatic Transcription Factors in the Wistar Rat Offspring Born to Dams Fed Folic Acid, Vitamin B12 Deficient Diets and Supplemented with Omega-3 Fatty Acids

Nutritional status of the mother is known to influence various metabolic adaptations required for optimal fetal development. These may be mediated by transcription factors like peroxisome proliferator activated receptors (PPARs), which are activated by long chain polyunsaturated fatty acids. The objective of the current study was to examine the expression of different hepatic transcription factors and the levels of global methylation in the liver of the offspring born to dams fed micronutrient deficient (folic acid and vitamin B12) diets and supplemented with omega-3 fatty acids. Female rats were divided into five groups (n = 8/group) as follows; control, folic acid deficient (FD), vitamin B12 deficient (BD) and omega-3 fatty acid supplemented groups (FDO and BDO). Diets were given starting from pre-conception and continued throughout pregnancy and lactation. Pups were dissected at the end of lactation. Liver tissues were removed; snap frozen and stored at −80°C. Maternal micronutrients deficiency resulted in lower (p<0.05) levels of pup liver docosahexaenoic acid (DHA) and arachidonic acid (ARA) as compared to the control group. Pup liver PPARα and PPARγ expression was lower (p<0.05) in the BD group although there were no differences in the expression of SREBP-1c, LXRα and RXRα expression. Omega-3 fatty acids supplementation to this group normalized (p<0.05) levels of both PPARα and PPARγ but reduced (p<0.05) SREBP-1c, LXRα and RXRα expression. There was no change in any of the transcription factors in the pup liver in the FD group. Omega-3 fatty acids supplementation to this group reduced (p<0.05) PPARα, SREBP-1c and RXRα expression. Pup liver global methylation levels were higher (p<0.01) in both the micronutrients deficient groups and could be normalized (p<0.05) by omega-3 fatty acid supplementation. Our novel findings suggest a role for omega-3 fatty acids in the one carbon cycle in influencing the hepatic expression of transcription factors in the offspring.

Dynamics of HBV cccDNA expression and transcription in different cell growth phase

BackgroundThe covalently closed-circular DNA (cccDNA) of hepatitis B virus (HBV) is associated with viral persistence in HBV-infected hepatocytes. However, the regulation of cccDNA and its transcription in the host cells at different growth stages is not well understood.MethodsWe took advantages of a stably HBV-producing cell line, 1.3ES2, and examine the dynamic changes of HBV cccDNA, viral transcripts, and viral replication intermediates in different cellular growth stages.ResultsIn this study, we showed that cccDNA increased suddenly in the initial proliferation phase of cell growth, probably attributable to its nuclear replenishment by intracellular nucleocapsids. The amount of cccDNA then decreased dramatically in the cells during their exponential proliferation similar to the loss of extrachromosomal plasmid DNA during cell division, after which it accumulated gradually while the host cells grew to confluency. We found that cccDNA was reduced in dividing cells and could be removed when proliferating cells were subjected to long term of lamivudine (3TC) treatment. The amounts of viral replicative intermediates were rapidly reduced in these proliferating cells and were significantly increased after cells reaching confluency. The expression levels of viral transcripts were increased in parallel with the elevated expression of hepatic transcription factors (HNF4α, CEBPα, PPARα, etc.) during cell growth confluency. The HBV transcripts were transcribed from both integrated viral genome and cccDNA, however the transcriptional abilities of cccDNA was less efficient then that from integrated viral genome in all cell growth stages. We also noted increases in the accumulation of intracellular viral particles and the secretion of mature virions as the cells reached confluency and ceased to grow.ConclusionsBased on the dynamics of HBV replication, we propose that HBV replication is modulated differently in the different stages of cell growth, and can be divided into three phases (initial proliferation phase, exponential proliferation phase and growth confluency phase) according to the cell growth curve. The regulation of cccDNA in different cell growth phase and its importance regarding HBV replication are discussed.

Impairment of hepatic Stat-3 activation and reduction of PPARα activity in fructose-fed rats

Fructose makes up a significant proportion of energy intake in westernized diets; its increased consumption has paralleled the growing prevalence of obesity and metabolic syndrome over the past two decades. In the current study, we demonstrate that fructose administration (10% wt/vol) in the drinking water of rats reduces the trans-activating and trans-repressing activity of the hepatic peroxisome proliferator-activated receptor alpha (PPARalpha). As a consequence, fructose decreases hepatic fatty oxidation and increases pro-inflammatory transcription factor nuclear factor kappaB (NF-kappaB) activity. These changes were not observed in glucose-administered rats (10% wt/vol), although both carbohydrates produced similar changes in plasma adiponectin and in the hepatic expression of transcription factors and enzymes involved in fatty acid synthesis. Fructose-fed, but not glucose-fed, rats were hyperleptinemic and exhibited increased tyrosine phosphorylation of the signal transducer and activator of transcription-3 (STAT-3) transcription factor, although they did not present a similar increase in the serine phosphorylation of nuclear STAT3. Thus, an impairment in the hepatic transduction of the leptin signal could be responsible for the observed alterations in PPARalpha activity in fructose-fed rats. Because PPARalpha activity is lower in human than in rodent liver, fructose ingestion in humans should cause even worse effects, which would partly explain the link between increased consumption of fructose and widening epidemics of obesity and metabolic syndrome. Hypertriglyceridemia and hepatic steatosis induced by fructose ingestion result from a reduction in the hepatic catabolism of fatty acids driven by a state of leptin resistance.

Tumor Necrosis Factor-α-Independent Downregulation of Hepatic Cholesterol 7α-Hydroxylase Gene in Mice Treated with Lead Nitrate

We previously reported that lead nitrate (LN), an inducer of hepatic tumor necrosis factor-alpha (TNF-alpha), downregulated gene expression of cholesterol 7alpha-hydroxylase. Herein, to clarify the role of TNF-alpha in LN-induced downregulation of cholesterol 7alpha-hydroxylase, effects of LN on gene expression of hepatic cholesterol 7alpha-hydroxylase (Cyp7a1) in TNF-alpha-knockout (KO) and TNF-alpha-wild-type (WT) mice were comparatively examined. Gene expression of hepatic Cyp7a1 in both WT and KO mice decreased to less than 5% of the corresponding controls at 6-12 h after treatment with LN (100 mumol/kg body weight, iv). Levels of hepatic TNF-alpha protein in either WT or KO mice were below the detection limit, although expression levels of the TNF-alpha gene markedly increased at 6 h in WT mice by LN treatment, but not in KO mice. In contrast, in both WT and KO mice, levels of hepatic IL-1beta protein, which is known to be a suppressor of the cholesterol 7alpha-hydroxylase gene in hamsters, were significantly increased 3-6 h after LN treatment. Furthermore, LN-induced downregulation of the Cyp7a1 gene did not necessarily result from altered gene expression of hepatic transcription factors, including positive regulators (liver X receptor alpha, retinoid X receptor alpha, fetoprotein transcription factor, and hepatocyte nuclear factor 4alpha) and a negative regulator small heterodimer partner responsible for expression of the Cyp7a1 gene. The present findings indicated that LN-induced downregulation of the Cyp7a1 gene in mice did not necessarily occur through a TNF-alpha-dependent pathway and might occur mainly through an IL-1beta-dependent pathway.

Introduction

The umbilical cord (UC) represents an important source of mesenchymal stem cells (MSC). These human UC-derived MSC (UC–MSC) have already been isolated using a protocol based on the migratory and plastic adhesive properties of MSC (UC–MSC–Mig). The UC–MSC–Mig isolation method, however, is difficult to standardize. Therefore, we developed an enzymatic isolation protocol (UC–MSC–Enz) to overcome the above mentioned disadvantages. First, we investigated the UC–MSC–Enz for their MSC properties. We found that UC–MSC–Enz express the MSC markers CD73, CD90 and CD105 and are able to differentiate into osteoblasts, adipocytes and chondroblasts fulfilling the MSC criteria of the International Society for Cellular Therapy. Previously we found that UC–MSC–Mig are unique among MSCs due to their significant expression of several hepatic (progenitor) markers. Therefore, we also investigated the expression of hepatic transcription factors and other hepatic markers in UC–MSC–Enz at both the mRNA and protein level. We found that the expression of hepatic transcription factors (GATA4, GATA6, SOX9 and SOX17) and hepatic markers (AFP, DPP4, CX43, DKK1, DSG2, KRT18 and KRT19) in UC–MSC–Enz was not significantly different from those of UC–MSC–Mig. Consequently, this optimized enzyme-based method represents a fast, robust and standardized way to isolate UC–MSC for a broad range of applications.

Extrahepatic biliary atresia: A disease or a phenotype?

Extrahepatic biliary atresia: A disease or a phenotype?

Expression of hepatic transcription factors during liver development and oval cell differentiation.

The oval cells are thought to be the progeny of a liver stem cell compartment and strong evidence now exists indicating that these cells can participate in liver regeneration by differentiating into different hepatic lineages. To better understand the regulation of this process we have studied the expression of liver-enriched transcriptional factors (HNF1 alpha and HNF1 beta, HNF3 alpha, HNF3 beta, and HNF3 gamma, HNF4, C/EBP, C/EBP beta, and DBP) in an experimental model of oval cell proliferation and differentiation and compared the expression of these factors to that observed during late stages of hepatic ontogenesis. The steady-state mRNA levels of four (HNF1 alpha, HNF3 alpha, HNF4, and C/EBP beta) "liver-enriched" transcriptional factors gradually decrease during the late period of embryonic liver development while three factors (HNF1 beta, HNF3 beta, and DBP) increase. In the normal adult rat liver the expression of all the transcription factors are restricted to the hepatocytes. However, during early stages of oval cell proliferation both small and large bile ducts start to express HNF1 alpha and HNF1 beta, HNF3 gamma, C/EBP, and DBP but not HNF4. At the later stages all of these factors are also highly expressed in the proliferating oval cells. Expression of HNF4 is first observed when the oval cells differentiate morphologically and functionally into hepatocytes and form basophilic foci. At that time the expression of some of the other factors is also further increased. Based on these data we suggest that the upregulation of the "establishment" factors (HNF1 and -3) may be an important step in oval cell activation. The high levels of these factors in the oval cells and embryonic hepatoblasts further substantiates the similarity between the two cell compartments. Furthermore, the data suggest that HNF4 may be responsible for the final commitment of a small portion of the oval cells to differentiate into hepatocytes which form the basophilic foci and eventually regenerate the liver parenchyma.