Common Transcriptional Program of Liver Fibrosis in Mouse Genetic Models and Humans.

Multifactorial metabolic diseases, such as non-alcoholic fatty liver disease, are a major burden to modern societies, and frequently present with no clearly defined molecular biomarkers. Herein we used system medicine approaches to decipher signatures of liver fibrosis in mouse models with malfunction in genes from unrelated biological pathways: cholesterol synthesis-, notch signaling-, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling-, and unknown lysosomal pathway-. Enrichment analyses of Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome and TRANScription FACtor (TRANSFAC) databases complemented with genome-scale metabolic modeling revealed fibrotic signatures highly similar to liver pathologies in humans.

The lysosomal transporter MFSD1 is essential for liver homeostasis and critically depends on its accessory subunit GLMP.

Lysosomes are major sites for intracellular, acidic hydrolase-mediated proteolysis and cellular degradation. The export of low-molecular-weight catabolic end-products is facilitated by polytopic transmembrane proteins mediating secondary active or passive transport. A number of these lysosomal transporters, however, remain enigmatic.

Receptor-Mediated Endocytosis of VEGF-A in Rat Liver Sinusoidal Endothelial Cells.

Background And Aims: Vascular endothelial growth factor (VEGF) receptors (VEGFR1 and VEGFR2) bind VEGF-A with high affinity. This study sought to determine the relative contributions of these two receptors to receptor-mediated endocytosis of VEGF-A and to clarify their endocytic itineraries in rat liver sinusoidal endothelial cells (LSECs).

Methods: Isolated LSECs and radiolabeled VEGF-A were used to examine surface binding and receptor-mediated endocytosis.

Age-dependent development of liver fibrosis in Glmp (gt/gt) mice.

Background: Mice lacking glycosylated lysosomal membrane protein (Glmp (gt/gt) mice) have liver fibrosis as the predominant phenotype due to chronic liver injury. The Glmp (gt/gt) mice grow and reproduce at the same rate as their wild-type siblings. Life expectancy is around 18 months.

Increased glucose utilization and decreased fatty acid metabolism in myotubes from Glmp(gt/gt) mice.

Glycosylated lysosomal membrane protein (GLMP) has been reported to enhance the expression from a peroxisome proliferator-activated receptor alpha (PPARα) responsive promoter, but also to be an integral lysosomal membrane protein. Using myotubes established from wild-type and Glmp(gt/gt) mice, the importance of GLMP in skeletal muscle was examined. Glmp(gt/gt) myotubes expressed a more glycolytic phenotype than wild-type myotubes.

Lack of the Lysosomal Membrane Protein, GLMP, in Mice Results in Metabolic Dysregulation in Liver.

Ablation of glycosylated lysosomal membrane protein (GLMP, formerly known as NCU-G1) has been shown to cause chronic liver injury which progresses into liver fibrosis in mice. Both lysosomal dysfunction and chronic liver injury can cause metabolic dysregulation. Glmp gt/gt mice (formerly known as Ncu-g1gt/gt mice) were studied between 3 weeks and 9 months of age.

Loss of lysosomal membrane protein NCU-G1 in mice results in spontaneous liver fibrosis with accumulation of lipofuscin and iron in Kupffer cells.

Human kidney predominant protein, NCU-G1, is a highly conserved protein with an unknown biological function. Initially described as a nuclear protein, it was later shown to be a bona fide lysosomal integral membrane protein. To gain insight into the physiological function of NCU-G1, mice with no detectable expression of this gene were created using a gene-trap strategy, and Ncu-g1(gt/gt) mice were successfully characterized.

Human NCU-G1 can function as a transcription factor and as a nuclear receptor co-activator.

Background: Novel, uncharacterised proteins represent a challenge in biochemistry and molecular biology. In this report we present an initial functional characterization of human kidney predominant protein, NCU-G1.

Results: NCU-G1 was found to be a highly conserved nuclear protein rich in proline with a molecular weight of approximately 44 kDa.

ABCA1, ABCG1 and SR-BI: hormonal regulation in primary rat hepatocytes and human cell lines.

Background: Scavenger receptor type B class I (SR-BI), ABC transporter A1 (ABCA1) -and G1 (ABCG1) all play important roles in the reverse cholesterol transport. Reverse cholesterol transport is a mechanism whereby the body can eliminate excess cholesterol. Here, the regulation of SR-BI, ABCA1, and ABCG1 by dexamethasone (a synthetic glucocorticoid) and insulin were studied in order to gain more insight into the role of these two hormones in the cholesterol metabolism.

Bile acids reduce SR-BI expression in hepatocytes by a pathway involving FXR/RXR, SHP, and LRH-1.

Hepatic SR-BI mediates uptake of circulating cholesterol into liver hepatocytes where a part of the cholesterol is metabolised to bile acids. In the hepatocytes, bile acids reduce their own synthesis by a negative feedback loop to prevent toxic high levels of bile acids. Bile acid-activated FXR/RXR represses expression of CYP7A1, the rate-limiting enzyme during bile acid synthesis, by inducing the expression of SHP, which inhibits LXR/RXR and LRH-1-transactivation of CYP7A1.

Glucocorticoid response and promoter occupancy of the mouse LXRalpha gene.

The liver X receptors alpha and beta (LXRalpha and LXRbeta) are members of the nuclear receptor superfamily of proteins which are highly expressed in metabolically active tissues. They regulate gene expression of critical genes involved in cholesterol catabolism and transport, lipid and triglyceride biosynthesis, and carbohydrate metabolism in response to distinct oxysterol intermediates in the cholesterol metabolic pathway. Several LXR target genes have been identified, but there is limited information on how expression of the LXRs themselves is controlled.

Akt/PTEN signaling mediates estrogen-dependent proliferation of primordial germ cells in vitro.

Testicular tumors in humans are reported to be significantly increasing in incidence. Embryo exposure to environmental estrogens has been proposed as one of the possible underlying causes. In mice, genetic, immunological, and experimental evidence suggest that germ cell testicular tumors may derive from primordial germ cells (PGCs), the embryonic precursors of gametes.

Oxysterol-activated LXRalpha/RXR induces hSR-BI-promoter activity in hepatoma cells and preadipocytes.

SR-BI mediates exchange of cholesterol between HDL and cells, and is a crucial factor in the transport of excessive cellular cholesterol from extrahepatic tissues to the liver ("reverse cholesterol transport") and, therefore, also for cholesterol homeostasis. Hepatic SR-BI mediates transfer of HDL-cholesterol to the hepatocytes where cholesterol may be metabolised to bile acids. LXR and SREBP are key factors in the regulation of cholesterol metabolism.

Different regulation of the LXRalpha promoter activity by isoforms of CCAAT/enhancer-binding proteins.

LXRs have recently been shown to regulate key enzymes in cholesterol degradation, reverse transport of cholesterol from peripheral cells, cholesterol uptake and lipogenesis. The LXRalpha promoter was thus studied to investigate if LXRalpha gene expression is under the regulation of transcription factors involved in adipogenesis. We report that the C/EBP transcription factor interacts with the promoter of the LXRalpha gene.

Members of the nuclear factor 1 family reduce the transcriptional potential of the nuclear receptor LXRalpha promoter.

Expression of the LXRalpha nuclear receptor in liver is predicted to affect cholesterol and lipid metabolism. Here we show that a short fragment from the LXRalpha gene promoter spanning the region from -144 to +43 relative to the mRNA initiation site can drive transcription of a reporter gene. Under basal conditions, in vitro DNase I footprinting demonstrated interaction between nuclear proteins and an NF1 recognition site in close vicinity to the transcriptional initiation.

Evidence for independent control at the mRNA and protein levels of cellular retinol binding protein 1 in rat Sertoli cells.

Cellular retinol binding protein 1 (CRBP1) is the cytosolic carrier for retinol. It is expressed in many tissues, but the concentrations vary considerably. In Sertoli cells from immature rat testis, CRBP1 is highly expressed.

Vitamin A-sensitive tissues in transgenic mice expressing high levels of human cellular retinol-binding protein type I are not altered phenotypically.

The suggested function of cellular retinol-binding protein type I [CRBP(I)] is to carry retinol to esterifying or oxidizing enzymes. The retinyl esters are used in storage or transport, whereas oxidized forms such as all-trans or 9-cis retinoic acid are metabolites used in the mechanism of action of vitamin A. Thus, high expression of human CRBP(I) [hCRBP(I)] in transgenic mice might be expected to increase the production of retinoic acid in tissues, thereby inducing a phenotype resembling vitamin A toxicity.

Method of transfection affects the cAMP-mediated induction of the RIIbeta subunit of protein kinase A in Sertoli cells: inhibition of response by increase in intracellullar calcium.

mRNA for the regulatory subunit RIIbeta of cAMP-dependent protein kinase is stimulated more than 50-fold by cAMP in primary cultures of rat Sertoli cells. We have previously shown that this induction involves regulation of transcriptional activation as well as mRNA stabilization. The rat RIIbeta gene contains no cAMP response element (CRE), and the induction of RIIbeta mRNA is slow and requires on-going protein synthesis.

Calcium/phospholipid-dependent protein kinases in rat Sertoli cells: regulation of androgen receptor messenger ribonucleic acid.

The possibility that Sertoli cell responses to testosterone are modulated by the calcium/phospholipid-dependent protein kinase (protein kinase C; PKC) was examined in rat Sertoli cells in culture. Both soluble and particulate cell fractions showed low constitutive phosphotransferase activity. Incubation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA; 10(-7) M) was associated with a transient induction in both cell fractions of calcium/phosphatidylserine-dependent PKC activity, which was elevated from 15 min to 1 h.

The rat growth hormone and human cellular retinol binding protein 1 genes share homologous NF1-like binding sites that exert either positive or negative influences on gene expression in vitro.

High levels of expression for the rat growth hormone (rGH) gene are restricted to the somatotroph cells of the anterior pituitary. Previously, we have shown that rGH cell-specific repression results in part from the recognition of negatively acting silencers by a number of nuclear proteins that repress basal promoter activity. Examination of these silencers revealed the presence of binding sites for proteins that belong to the NF1 family of transcription factors.

Characterization of the 5'-flanking region of the gene for the cAMP-inducible protein kinase A subunit, RIIbeta, in Sertoli cells.

Activation of cyclic AMP-dependent protein kinases (protein kinase A, PKA) by gonadotropins and cyclic AMP (cAMP) plays an important role in the regulation of testicular functions. A regulatory subunit, RIIbeta, of PKA is transcriptionally induced in rat Sertoli cells in response to treatment with cAMP. The present study addresses regulatory mechanisms leading to increased transcription of the rat RIIbeta gene.

Retinyl ester storage is normal in transgenic mice with enhanced expression of cellular retinol-binding protein type I.

This report describes the production and characterization of transgenic mice with high expression of human cellular retinol-binding protein type I [hCRBP(I)]. In initial experiments, overexpression of hCRBP(I) was driven by the strong promoter SR(alpha), but no transgenic offspring were produced. When we used the less efficient mouse metallothionein I promoter fused to the hCRBP(I) cDNA for microinjection, we obtained 12% transgenic offspring.

Regulation of protein kinase A subunits by cyclic adenosine 3',5'-monophosphate in a mouse Sertoli cell line (MSC-1): induction of RII beta messenger ribonucleic acid is independent of continuous protein synthesis.

We report the basal and cAMP-regulated expression of protein kinase A (PKA) subunits in a mouse Sertoli cell line (MSC-1). Of the PKA subunits expressed by these cells (RI alpha, RII alpha, RII beta, C alpha, C beta), only RII beta was regulated by cAMP. An approximately 8-fold induction of RII beta mRNA and a 3-fold induction of RII beta protein was observed during 48 h of cAMP-stimulation.

Tissue distribution of the receptor for plasma retinol-binding protein.

The tissue distribution of the retinol-binding-protein receptor has been studied by using a cell-free binding assay. High binding activity was found in placenta, retina pigment epithelial cells, bone marrow and kidneys. Specific binding activity was also found in the small intestines, spleen and liver, and to a lesser extent in lung.