Association between HNF4A rs1800961 polymorphisms and gallstones in a Taiwanese population.

BACKGROUND AND AIM: A large genetic effect of a novel gallstone-associated genetic variant, the hepatocyte nuclear factor 4α (HNF4A) rs1800961 polymorphism, has been identified through recent genome-wide association studies. However, this effect has not been validated in Asian populations. We investigated the association between the rs1800961 variant and gallstones among a Taiwanese population. METHODS: A total of 20 405 participants aged between 30 and 70 years voluntarily enrolled in the Taiwan Biobank. Self-report questionnaires, physical examinations, biochemical tests, and genotyping were used for analysis. The association of the HNF4A rs1800961 variant and other metabolic risks with gallstone disease was analyzed using multiple logistic regression models. RESULTS: The minor T allele of HNF4A rs1800961 was associated with an increased risk of gallstone, and the association remained significant even after adjustment for other risk factors including age, body mass index (BMI), diabetes, hyperlipidemia, hypertension, and cigarette smoking (adjusted odds ratio [OR] = 1.90, 95% confidence interval [CI] = 1.31 to 2.75) in male participants. When further stratified by BMI and age, the lithogenic effect was the most significant in male participants with obesity (adjusted OR = 3.55, 95% CI = 1.92 to 6.56) and who were younger (adjusted OR = 2.45, 95% CI = 1.49 to 4.04). CONCLUSION: The novel gallstone-associated HNF4A rs1800961 variant was associated with the risk of gallstone in the Taiwanese men. Screening for the rs1800961 polymorphism may be particularly useful in assessing the risk of gallstone formation in younger or obese men.

Transcription factor HNF1ß controls a transcriptional network regulating kidney cell structure and tight junction integrity.

Mutations in the hepatocyte nuclear factor (HNF)1ß gene (HNF1B) cause autosomal dominant tubulointerstitial kidney disease, a rare and heterogeneous disease characterized by renal cysts and/or malformation, maturity-onset diabetes of the young, hypomagnesemia, and hypokalemia. The electrolyte disturbances may develop in the distal part of the nephron, which is important for fine-tuning of Mg2+ and Ca2+ reabsorption. Therefore, we aimed to study the transcriptional network directed by HNF1ß in the distal part of the nephron. We combined HNF1ß chromatin immunoprecipitation-sequencing and mRNA expression data to identify direct targets of HNF1ß in a renal distal convoluted tubule cell line (mpkDCT). Gene Ontology term pathway analysis demonstrated enrichment of cell polarity, cell-cell junction, and cytoskeleton pathways in the dataset. Genes directly and indirectly regulated by HNF1ß within these pathways included members of the apical and basolateral polarity complexes including Crumbs protein homolog 3 (Crb3), partitioning defective 6 homolog-ß (Pard6b), and LLGL Scribble cell polarity complex component 2 (Llgl2). In monolayers of mouse inner medullary collecting duct 3 cells expressing dominant negative Hnf1b, tight junction integrity was compromised, as observed by reduced transepithelial electrical resistance values and increased permeability for fluorescein (0.4 kDa) compared with wild-type cells. Expression of dominant negative Hnf1b also led to a decrease in height (30%) and an increase in surface (58.5%) of cells grown on membranes. Moreover, three-dimensional spheroids formed by cells expressing dominant negative Hnf1b were reduced in size compared with wild-type spheroids (30%). Together, these findings demonstrate that HNF1ß directs a transcriptional network regulating tight junction integrity and cell structure in the distal part of the nephron.NEW & NOTEWORTHY Genetic defects in transcription factor hepatocyte nuclear factor (HNF)1ß cause a heterogeneous disease characterized by electrolyte disturbances, kidney cysts, and diabetes. By combining RNA-sequencing and HNF1ß chromatin immunoprecipitation-sequencing data, we identified new HNF1ß targets that were enriched for cell polarity pathways. Newly discovered targets included members of polarity complexes Crb3, Pard6b, and Llgl2. Functional assays in kidney epithelial cells demonstrated decreased tight junction integrity and a loss of typical cuboidal morphology in mutant Hnf1b cells.

Hepatocyte Nuclear Factor-1α Increases Fibrinogen Gene Expression in Liver and Plasma Fibrinogen Concentration in Rats with Experimental Chronic Renal Failure.

Chronic kidney disease (CKD) is associated with elevated plasma fibrinogen concentration. However, the underlying molecular mechanism for elevated plasma fibrinogen concentration in CKD patients has not yet been clarified. We recently found that HNF1α was significantly upregulated in the liver of chronic renal failure (CRF) rats, an experimental model of CKD in patients. Given that the promoter region of the fibrinogen gene possesses potential binding sites for HNF1α, we hypothesized that the upregulation of HNF1α can increase fibrinogen gene expression and consequently plasma fibrinogen concentration in the experimental model of CKD. Here, we found the coordinated upregulation of Aα-chain fibrinogen and Hnfα gene expression in the liver and elevated plasma fibrinogen concentrations in CRF rats, compared with pair-fed and control animals. Liver Aα-chain fibrinogen and HNF1α mRNAs levels correlated positively with (a) liver and plasma fibrinogen levels and (b) liver HNF1α protein levels. The positive correlation between (a) liver Aα-chain fibrinogen mRNA level, (b) liver Aα-chain fibrinogen level, and (c) serum markers of renal function suggest that fibrinogen gene transcription is closely related to the progression of kidney disease. Knockdown of Hnfα in the HepG2 cell line by small interfering RNA (siRNA) led to a decrease in fibrinogen mRNA levels. Clofibrate, an anti-lipidemic drug that reduces plasma fibrinogen concentration in humans, decreased both HNF1α and Aα-chain fibrinogen mRNAs levels in (a) the liver of CRF rats and (b) HepG2 cells. The obtained results suggest that (a) an elevated level of liver HNF1α can play an important role in the upregulation of fibrinogen gene expression in the liver of CRF rats, leading to an elevated concentration of plasma fibrinogen, a protein related to the risk of cardiovascular disease in CKD patients, and (b) fibrates can decrease plasma fibrinogen concentration through inhibition of HNF1α gene expression.

Relative DNA Methylation and Demethylation Efficiencies during Postnatal Liver Development Regulate Hepatitis B Virus Biosynthesis.

Hepatitis B virus (HBV) transcription and replication increase progressively throughout postnatal liver development with maximal viral biosynthesis occurring at around 4 weeks of age in the HBV transgenic mouse model of chronic infection. Increasing viral biosynthesis is associated with a corresponding progressive loss of DNA methylation. The loss of DNA methylation is associated with increasing levels of 5-hydroxymethylcytosine (5hmC) residues which correlate with increased liver-enriched pioneer transcription factor Forkhead box protein A (FoxA) RNA levels, a rapid decline in postnatal liver DNA methyltransferase (Dnmt) transcripts, and a very modest reduction in ten-eleven translocation (Tet) methylcytosine dioxygenase expression. These observations are consistent with the suggestion that the balance between active HBV DNA methylation and demethylation is regulated by FoxA recruitment of Tet in the presence of declining Dnmt activity. These changes lead to demethylation of the viral genome during hepatocyte maturation with associated increases in viral biosynthesis. Consequently, manipulation of the relative activities of these two counterbalancing processes might permit the specific silencing of HBV gene expression with the loss of viral biosynthesis and the resolution of chronic HBV infections.IMPORTANCE HBV biosynthesis begins at birth and increases during early postnatal liver development in the HBV transgenic mouse model of chronic infection. The levels of viral RNA and DNA synthesis correlate with pioneer transcription factor FoxA transcript plus Tet methylcytosine dioxygenase-generated 5hmC abundance but inversely with Dnmt transcript levels and HBV DNA methylation. Together, these findings suggest that HBV DNA methylation during neonatal liver development is actively modulated by the relative contributions of FoxA-recruited Tet-mediated DNA demethylation and Dnmt-mediated DNA methylation activities. This mode of gene regulation, mediated by the loss of DNA methylation at hepatocyte-specific viral and cellular promoters, likely contributes to hepatocyte maturation during liver development in addition to the postnatal activation of HBV transcription and replication.

The molecular functions of hepatocyte nuclear factors - In and beyond the liver.

The hepatocyte nuclear factors (HNFs) namely HNF1α/ß, FOXA1/2/3, HNF4α/γ and ONECUT1/2 are expressed in a variety of tissues and organs, including the liver, pancreas and kidney. The spatial and temporal manner of HNF expression regulates embryonic development and subsequently the development of multiple tissues during adulthood. Though the HNFs were initially identified individually based on their roles in the liver, numerous studies have now revealed that the HNFs cross-regulate one another and exhibit synergistic relationships in the regulation of tissue development and function. The complex HNF transcriptional regulatory networks have largely been elucidated in rodent models, but less so in human biological systems. Several heterozygous mutations in these HNFs were found to cause diseases in humans but not in rodents, suggesting clear species-specific differences in mutational mechanisms that remain to be uncovered. In this review, we compare and contrast the expression patterns of the HNFs, the HNF cross-regulatory networks and how these liver-enriched transcription factors serve multiple functions in the liver and beyond, extending our focus to the pancreas and kidney. We also summarise the insights gained from both human and rodent studies of mutations in several HNFs that are known to lead to different disease conditions.

Clinical Management of Women with Monogenic Diabetes During Pregnancy.

PURPOSE OF REVIEW: Monogenic diabetes accounts for 1-2% of all diabetes cases, but is frequently misdiagnosed as type 1, type 2, or gestational diabetes. Accurate genetic diagnosis directs management, such as no pharmacologic treatment for GCK-MODY, low-dose sulfonylureas for HNF1A-MODY and HNF4A-MODY, and high-dose sulfonylureas for KATP channel-related diabetes. While diabetes treatment is defined for the most common causes of monogenic diabetes, pregnancy poses a challenge to management. Here, we discuss the key issues in pregnancy affected by monogenic diabetes. RECENT FINDINGS: General recommendations for pregnancy affected by GCK-MODY determine need for maternal insulin treatment based on fetal mutation status. However, a recent study suggests macrosomia and miscarriage rates may be increased with this strategy. Recent demonstration of transplacental transfer of sulfonylureas also raises questions as to when insulin should be initiated in sulfonylurea-responsive forms of monogenic diabetes. Pregnancy represents a challenge in management of monogenic diabetes, where factors of maternal glycemic control, fetal mutation status, and transplacental transfer of medication must all be taken into consideration. Guidelines for pregnancy affected by monogenic diabetes will benefit from large, prospective studies to better define the need for and timing of initiation of insulin treatment.

Hepatocyte Nuclear Factor-4α P2 Promoter Variants Are Associated With the Risk of Metabolic Syndrome and Testosterone Deficiency in Aging Taiwanese Men.

BACKGROUND: Hepatocyte nuclear factor-4α (HNF4A) can influence the risk of insulin resistance that is postulated to be an important link between metabolic syndrome (MetS) and testosterone deficiency (TD) in men. AIM: To investigate the relationship between single-nucleotide polymorphisms (SNPs) of HNF4A and the risk of developing MetS and TD in a population of aging Taiwanese men. METHODS: A free health screening of men over 40 years of age was conducted in a medical center in Kaohsiung City, Taiwan. All participants underwent a physical examination, answered a questionnaire on demographics and medical history, completed the Androgen Deficiency in The Aging Male questionnaire to assess clinical symptoms of TD, and provided 20-mL whole blood samples for biochemical, hormonal, and genetic evaluation. MAIN OUTCOME MEASURE: 3 common SNPs (rs11574736, rs1884613, and rs2144908) of HNF4A were selected and identified using a TaqMan 5' allelic discrimination assay. RESULTS: 559 men were enrolled for this study (mean age, 55.8± 4.9 years). Prevalence of TD was significantly higher (P = .031) in subjects with MetS (16.8%) than those without MetS (10.1%). In SNP rs1884613 of HNF4A, subjects with the C allele carried a 1.31- and 1.50-times higher risk of developing MetS and TD, respectively, compared to those with the G allele, after adjusting for potential covariates. In addition, subjects with the CC genotype were exposed to a 1.91- and 2.20-times higher risk of developing MetS and TD, respectively, compared to those with the GG genotype. CLINICAL IMPLICATIONS: Our findings may point to the importance of the role played by insulin resistance in the link between MetS and TD. STRENGTH & LIMITATIONS: Our current work is the first report with adequate sample size to evaluate the role of genetic variants of HNF4A on the risk of both MetS and TD in men. The limitations included subjects enrolled from a free health screening and single measurement of serum testosterone levels. CONCLUSION: The rs1884613 SNP marker of HNF4A is significantly associated with an increased risk for developing both MetS and TD in aging Taiwanese men. Further population-based studies utilizing larger samples of different ethnicities may be needed to confirm these preliminary results. Liu C-C, Lee Y-C, Hung S-P. Hepatocyte Nuclear Factor-4α P2 Promoter Variants Are Associated With the Risk of Metabolic Syndrome and Testosterone Deficiency in Aging Taiwanese Men. J Sex Med 2018;15:1527-1536.

Oct4 Is Crucial for Transdifferentiation of Hepatocytes to Biliary Epithelial Cells in an In Vitro Organoid Culture Model.

Hepatocyte to biliary transdifferentiation has been documented in various models of bile duct injury. In this process, mature hepatocytes transform into mature biliary epithelial cells by acquiring biliary phenotypic markers. Several signaling pathways including PI3 kinase, Notch, Hes1, Sox9, and Hippo are shown to be involved in the process. However, whether Oct4 is involved in hepatocyte to biliary transdifferentiation is unknown. We investigated the role of Oct4 in hepatocyte to biliary transdifferentiation utilizing an in vitro organoid culture system as a model of transdifferentiation. Oct4 was inhibited using adenovirus containing Oct4 shRNA. Hepatocyte-specific HNF-4α and biliary-specific HNF-1ß and CK19 expression were assessed to gauge the extent of transdifferentiation. Oct4 was induced during hepatocyte to biliary transdifferentiation. Oct4 inhibition significantly downregulated the appearance of biliary cells from hepatocytes. This was accompanied by a significant downregulation of signaling pathways including Notch, Sox9, and Hippo. Our findings suggest that Oct4 is crucial for hepatocyte to biliary transdifferentiation and maturation and that it acts upstream of Notch, Sox9, and Hippo signaling in this model. This finding identifies new signaling through Oct4 in plasticity between hepatocytes and biliary epithelial cells, which can be potentially utilized to identify new strategies in chronic biliary diseases.

Genetic basis of early-onset, maturity-onset diabetes of the young-like diabetes in Japan and features of patients without mutations in the major MODY genes: Dominance of maternal inheritance.

BACKGROUND: Causative mutations cannot be identified in the majority of Asian patients with suspected maturity-onset diabetes of the young (MODY). OBJECTIVES: To elucidate the genetic basis of Japanese patients with MODY-like diabetes and gain insight into the etiology of patients without mutations in the major MODY genes. SUBJECTS: A total of 263 Japanese patients with early-onset, non-obese, MODY-like diabetes mellitus referred to Osaka City General Hospital for diagnosis. METHODS: Mutational analysis of the four major MODY genes (GCK, HNF1A, HNF4A, HNF1B) by Sanger sequencing. Mutation-positive and mutation-negative patients were further analyzed for clinical features. RESULTS: Mutations were identified in 103 (39.2%) patients; 57 mutations in GCK; 29, HNF1A; 7, HNF4A; and 10, HNF1B. Contrary to conventional diagnostic criteria, 18.4% of mutation-positive patients did not have affected parents and 8.2% were in the overweight range (body mass index [BMI] >85th percentile). HOMA-IR at diagnosis was elevated (>2) in 15 of 66 (22.7%) mutation-positive patients. Compared with mutation-positive patients, mutation-negative patients were significantly older (P = 0.003), and had higher BMI percentile at diagnosis (P = 0.0006). Interestingly, maternal inheritance of diabetes was significantly more common in mutation-negative patients (P = 0.0332) and these patients had significantly higher BMI percentile as compared with mutation-negative patients with paternal inheritance (P = 0.0106). CONCLUSIONS: Contrary to the conventional diagnostic criteria, de novo diabetes, overweight, and insulin-resistance are common in Japanese patients with mutation-positive MODY. A significant fraction of mutation-negative patients had features of early-onset type 2 diabetes common in Japanese, and non-Mendelian inheritance needs to be considered for these patients.

[Endoplasmic reticulum stress in INS-1-3 cell associated with the expression changes of MODY gene pathway].

Objective: To profile the gene expression changes associated with endoplasmic reticulum stress in INS-1-3 cells induced by thapsigargin (TG) and tunicamycin (TM). Methods: Normal cultured INS-1-3 cells were used as a control. TG and TM were used to induce endoplasmic reticulum stress in INS-1-3 cells. Digital gene expression profiling technique was used to detect differentially expressed gene. The changes of gene expression were detected by expression pattern clustering analysis, gene ontology (GO) function and pathway enrichment analysis. Real time polymerase chain reaction (RT-PCR) was used to verify the key changes of gene expression. Results: Compared with the control group, there were 57 (45 up-regulated, 12 down-regulated) and 135 (99 up-regulated, 36 down-regulated) differentially expressed genes in TG and TM group, respectively. GO function enrichment analyses indicated that the main enrichment was in the endoplasmic reticulum. In signaling pathway analysis, the identified pathways were related with endoplasmic reticulum stress, antigen processing and presentation, protein export, and most of all, the maturity onset diabetes of the young (MODY) pathway. Conclusion: Under the condition of endoplasmic reticulum stress, the related expression changes of transcriptional factors in MODY signaling pathway may be related with the impaired function in islet beta cells.

[THE ROLE OF THE FOXA SUBFAMILY FACTORS IN THE EMBRYONIC DEVELOPMENT AND CARCINOGENESIS OF THE PANCREAS.]

The embryonic development and carcinogenesis are controlled by many transcription factors. The regulatory systems involved in embryogenesis of an organ are also involved in the tumor development in the same organ. FOX family proteins are transcription factors, which play a key role in these processes. The pioneering factors of the FOXA subfamily act at the very early stages of the embryonic development by interacting with condensed chromatin and thereby enabling the expression of the formerly silent important transcription factors. The role of these factors in tumor development is currently not fully elucidated, although recent studies indicate the important contribution of the FOXA subfamily proteins at the early stages of carcinogenesis. This review is restricted to the role of the FOXA factors in embryogenesis of the pancreas and their significance in the development of the pancreatic ductal adenocarcinoma.

Transforming growth factor ß1 represses proximal tubular cell microRNA-192 expression through decreased hepatocyte nuclear factor DNA binding.

miR (microRNA)-192 plays key roles in renal pathological and physiological responses, by repressing targets including Zeb1, Zeb2 and Wnk1. In the present study, we have assessed the regulation of miR-192 expression. We found that TGF-ß1 (transforming growth factor ß1) down-regulates miR-192 and miR-194, co-transcribed in the shared precursor pri-miR (primary miR transcript)-192/194. Luciferase reporter analysis showed constitutive promoter activity within nucleotides +21 to -223. We identified HNF (hepatocyte nuclear factor) and p53 binding sites within this region that were required for constitutive promoter activity, which was decreased by TGF-ß1 through an Alk5-dependent mechanism. TGF-ß1 treatment decreased HNF binding to the miR-194-2/192 promoter, whereas knockdown of HNF-1 inhibited mature miR-192 and miR-194 expression. miR-192, miR-194 and HNF expression were restricted to a defined subset of human tissues including kidney, small intestine, colon and liver. Our results from the present study identify co-ordinated regulation of miR-192 and miR-194, with binding of HNF and p53 transcription factors necessary for activation of transcription, and TGF-ß1-mediated repression through decreased HNF binding to its cognate promoter element.

A Hepatocyte Nuclear Factor BtabHNF4 Mediates Desiccation Tolerance and Fecundity in Whitefly (Bemisia tabaci).

Hepatocyte nuclear factor 4 (HNF4) is essential for glucose homeostasis and lipid metabolism in insects. However, little is known about the role of HNF4 in whiteflies. In the present study, we identified a hepatocyte nuclear factor protein from Bemsia tabaci (Diptera: Drosophilidae) and named it BtabHNF4. The full-length of BtabHNF4 was 3,006 bp, encoding a sequence of 434 amino acids that contains a conserved zinc-finger DNA-binding domain (DBD) and a well-conserved ligand-binding domain (LBD). The temporal and spatial expression showed that BtabHNF4 was highly expressed in the female adult stage and abdominal tissues of B. tabaci. A leaf-mediated RNA interference method was used to explore the function of BtabHNF4 in whiteflies. Our results showed that the knockdown of BtabHNF4 influences the desiccation tolerance, egg production, and egg hatching rate of whiteflies. Additionally, BtabHNF4 silencing significantly inhibited the expression level of vitellogenin. These results expand the function of HNF4 and pave the way for understanding the molecular mechanisms of HNF4 in regulating multiple physiological processes.

Pleiotropy of microRNA-192 in the kidney.

Diverse aetiologies result in significant deviation from homoeostasis in the kidney, leading to CKD (chronic kidney disease). CKD progresses to end-stage renal disease principally as a result of renal fibrosis, although the molecular mechanisms underlying this fibrotic process are still poorly understood. miRNAs (microRNAs) are a recently discovered family of endogenous short single-stranded RNAs that regulate global gene expression at the post-transcriptional level. The recent findings from our laboratory and others discussed in the present review outline pleiotropic roles for miR-192 in renal homoeostasis and in the fibrotic kidney. We describe miR-192-driven anti-and pro-fibrotic effects via the repression of ZEB1 and ZEB2 (zinc finger E-box-binding homeobox proteins 1 and 2), resulting in changes in extracellular matrix deposition and cell differentiation.

Identification of DNA regions and a set of transcriptional regulatory factors involved in transcriptional regulation of several human liver-enriched transcription factor genes.

Mammalian tissue- and/or time-specific transcription is primarily regulated in a combinatorial fashion through interactions between a specific set of transcriptional regulatory factors (TRFs) and their cognate cis-regulatory elements located in the regulatory regions. In exploring the DNA regions and TRFs involved in combinatorial transcriptional regulation, we noted that individual knockdown of a set of human liver-enriched TRFs such as HNF1A, HNF3A, HNF3B, HNF3G and HNF4A resulted in perturbation of the expression of several single TRF genes, such as HNF1A, HNF3G and CEBPA genes. We thus searched the potential binding sites for these five TRFs in the highly conserved genomic regions around these three TRF genes and found several putative combinatorial regulatory regions. Chromatin immunoprecipitation analysis revealed that almost all of the putative regulatory DNA regions were bound by the TRFs as well as two coactivators (CBP and p300). The strong transcription-enhancing activity of the putative combinatorial regulatory region located downstream of the CEBPA gene was confirmed. EMSA demonstrated specific bindings of these HNFs to the target DNA region. Finally, co-transfection reporter assays with various combinations of expression vectors for these HNF genes demonstrated the transcriptional activation of the CEBPA gene in a combinatorial manner by these TRFs.

Differential transcriptional characteristics of small and large biliary epithelial cells derived from small and large bile ducts.

Biliary epithelial cells (BEC) are morphologically and functionally heterogeneous. To investigate the molecular mechanism for their diversities, we test the hypothesis that large and small BEC have disparity in their target gene response to their transcriptional regulator, the biliary cell-enriched hepatocyte nuclear factor HNF6. The expression of the major HNF (HNF6, OC2, HNF1b, HNF1a, HNF4a, C/EBPb, and Foxa2) and representative biliary transport target genes that are HNF dependent were compared between SV40-transformed BEC derived from large (SV40LG) and small (SV40SM) ducts, before and after treatment with recombinant adenoviral vectors expressing HNF6 (AdHNF6) or control LacZ cDNA (AdLacZ). Large and small BEC were isolated from mouse liver treated with growth hormone, a known transcriptional activator of HNF6, and the effects on selected target genes were examined. Constitutive Foxa2, HNF1a, and HNF4a gene expression were 2.3-, 12.4-, and 2.6-fold, respectively, higher in SV40SM cells. This was associated with 2.7- and 4-fold higher baseline expression of HNF1a- and HNF4a-regulated ntcp and oatp1 genes, respectively. Following AdHNF6 infection, HNF6 gene expression was 1.4-fold higher (P = 0.02) in AdHNF6 SV40SM relative to AdHNF6 SV40LG cells, with a corresponding higher Foxa2 (4-fold), HNF1a (15-fold), and HNF4a (6-fold) gene expression in AdHNF6-SV40SM over AdHNF6-SV40LG. The net effects were upregulation of HNF6 target gene glucokinase and of Foxa2, HNF1a, and HNF4a target genes oatp1, ntcp, and mrp2 over AdLacZ control in both cells, but with higher levels in AdH6-SV40SM over AdH6-SV40LG of glucokinase, oatp1, ntcp, and mrp2 (by 1.8-, 3.4-, 2.4-, and 2.5-fold, respectively). In vivo, growth hormone-mediated increase in HNF6 expression was associated with similar higher upregulation of glucokinase and mrp2 in cholangiocytes from small vs. large BEC. Small and large BEC have a distinct profile of hepatocyte transcription factor and cognate target gene expression, as well as differential strength of response to transcriptional regulation, thus providing a potential molecular basis for their divergent function.

[Progress in diabetes genetics]. / Fremskritt innen diabetesgenetikk.

BACKGROUND: Diabetes is classified as Type 1 diabetes, Type 2 diabetes, gestational diabetes and other types. Our goal was to provide an overview of new genetic knowledge of monogenic and type 2 diabetes. MATERIAL AND METHOD: The article is based on literature identified through a non-systematic search in PubMed and own experience concerning research in diabetes genetics and treatment of patients with monogenic diabetes. RESULTS: 18 genes have been found for which one single mutation may cause diabetes. The most common causes for such monogenic diabetes are mutations in the genes KCNJ11, ABCC8 and INS when the condition is diagnosed at the age 0 - 6 months, and in the genes HNF1A, GCK, HNF4A and HNF1B when the diagnosis is made later than six months of age. Genetic testing is appropriate in assessment of monogenic diabetes, because antidiabetic tablets rather that insulin injections can be used to treat patients with mutations in certain genes; i.e. KCNJ11, ABCC8, HNF1A and HNF4A. Genome-wide association studies have recently identified about 20 genetic variants that increase the risk of Type 2 diabetes, but which have a low predictive value for development of disease. How these genetic variants can cause Type 2 diabetes has not been assessed and clinical relevance remains to be shown. INTERPRETATION: So far, genetic findings only affect diagnosis and treatment of monogenic diabetes.

Interaction of hepatocyte nuclear factors in transcriptional regulation of tissue specific hormonal expression of human multidrug resistance-associated protein 2 (abcc2).

Multidrug resistance-associated protein 2 (MRP2) (ABCC2) is an ATP-binding cassette membrane protein located primarily on apical surface of hepatocytes that mediates transport of conjugated xenobiotics and endogenous compounds into bile. MRP2 is highly expressed in hepatocytes, and at lower levels in small intestines, stomach and kidney. Previous reports have characterized mammalian MRP2 promoters, but none have established the molecular mechanism(s) involved in liver enriched expression. This study aims to investigate the mechanism of hepatic MRP2 regulation. A 2130 bp of MRP2 promoter was cloned from PAC-1 clone P108G1-7, to identify putative liver specific/hormone responsive functional DNA binding sites. Using deletion analysis, site specific mutagenesis and co-transfection studies, liver specific expression was determined. MRP2 promoter-LUC constructs were highly expressed in liver cell lines compared to non-liver cells. The region extending from -3 to+458 bp of MRP2 promoter starting from AUG contained the potential binding sites for CAATT box enhancer binding protein (C/EBP), hepatocytes nuclear factor 1, 3 and 4 (HNF1, HNF3, and HNF4. Only HNF1 and HNF4 co-transfection with MRP2 luciferase increased expression. Site specific mutational analysis of HNF1 binding site indicated an important role for HNF1alpha. HNF4alpha induction of MRP2 was independent of HNF1 binding site. C/EBP, HNF3, and HNF6 inhibited HNF1alpha while HNF4alpha induced MRP2 luciferase expression and glucocorticoids stimulated MRP2 expression. This study emphasizes the complex regulation of MRP2 with HNF1alpha and HNF4alpha playing a central role. The coordinated regulation of xenobiotic transporters and oxidative conjugation may determine the adaptive responses to cellular detoxification processes.

Interdependent fibroblast growth factor and activin A signaling promotes the expression of endodermal genes in differentiating mouse embryonic stem cells expressing Src Homology 2-domain inactive Shb.

The mechanisms controlling endodermal development during stem cell differentiation have been only partly elucidated, although previous studies have suggested the participation of fibroblast growth factor (FGF) and activin A in these processes. Shb is a Src homology 2 (SH2) domain-containing adapter protein that has been implicated in FGF receptor 1 (FGFR1) signaling. To study the putative crosstalk between activin A and Shb-dependent FGF signaling in the differentiation of endoderm from embryonic stem (ES) cells, embryoid bodies (EBs) derived from mouse ES cells overexpressing wild-type Shb or Shb with a mutated SH2 domain (R522K-Shb) were cultured in the presence of activin A. We show that expression of R522K-Shb results in up-regulation of FGFR1 and FGF2 in EBs. Addition of activin A to the cultures enhances the expression of endodermal genes primarily in EBs expressing mutant Shb. Inhibition of FGF signaling by the addition of the FGFR1 inhibitor SU5402 completely counteracts the synergistic effects of R522K-Shb and activin A. In conclusion, the present results suggest that expression of R522K-Shb enhances certain signaling pathways downstream of FGF and that an interplay between FGF and activin A participates in ES cell differentiation to endoderm.