Cholesterol biosynthesis from lanosterol: molecular cloning, chromosomal localization, functional expression and liver-specific gene regulation of rat sterol Δ8-isomerase, a cholesterogenic enzyme with multiple functions

2001 ◽  
Vol 353 (3) ◽  
pp. 689-699 ◽  
Author(s):  
Soo-Han BAE ◽  
Jekyung SEONG ◽  
Young-Ki PAIK
Keyword(s):  
Gene Regulation ◽  
Amino Acid ◽  
Mrna Expression ◽  
Cholesterol Metabolism ◽  
Binding Protein ◽  
Cholesterol Biosynthesis ◽  
Specific Gene ◽  
Mrna Levels ◽  
H4iie Cells ◽  
Old Rats

Sterol ∆8-isomerase (SI) (EC 5.3.3.5), also known as emopamil binding protein or sigma receptor, catalyses the conversion of the 8-ene isomer into the 7-ene isomer in the cholesterol biosynthetic pathway in mammals. Recently, mutations of SI have been found to be associated with Conradi–Hünermann syndrome in humans. To investigate the invitro and invivo modes of molecular regulation of SI and its role in cholesterol biosynthesis in mammals, we isolated a full-length cDNA encoding rat SI. The deduced amino-acid sequence of rat SI predicts a 230-residue protein (26737Da) with 87% and 80% amino-acid identity to mouse and human counterparts. The rat SI gene was mapped to chromosome 12q1.2 using fluorescence insitu hybridization (FISH). The biological function of the cloned rat SI cDNA was verified by overexpressing recombinant Myc–SI in Saccharomycescerevisiae. It showed a characteristic pattern of inhibition on exposure to trans-2-[4-(1,2-diphenylbuten-1-yl)phenoxy]-N,N-dimethylethylamine (tamoxifen; IC50 = 11.2µM) and 3β-[2-(diethylamino)ethoxy]androst-5-en-17-one (U18666A; IC50 = 4.2µM), two well known potent inhibitors of SI. Northern-blot analysis of 3-week-old rats compared with 2-year-old rats showed that SI mRNA expression in both age groups was restricted to liver, where a 70% reduction in mRNA levels was observed in 2-year-old rats. The FISH studies revealed ubiquitous expression of SI mRNA in rat hepatocytes. The invitro studies showed that the SI mRNA was highly suppressed by 25-hydroxycholesterol in H4IIE cells. Treatment of H4IIE cells grown in medium supplemented with fetal bovine serum with tamoxifen for 24h resulted in a dose-dependent induction of SI mRNA, with a concomitant suppression of sterol regulatory element binding protein-1 mRNA. Interestingly, this effect was not seen in emopamil-treated cells. The invivo experiments also indicate that both mRNA expression and enzymic activity of SI in liver were induced approx. 3-fold in rats fed 5% (w/w) cholestyramine plus 0.1% (w/w) lovastatin in normal chow for 2 weeks. With this newly cloned rat SI cDNA, it becomes possible to gain molecular understanding of previously unknown and tamoxifen-mediated gene regulation of SI that is involved in cholesterol metabolism, ischaemia and genetic diseases.

scholarly journals The effect of amino-acid-supplemented wheat gluten on cholesterol metabolism in the rat

1985 ◽  
Vol 53 (1) ◽  
pp. 25-30 ◽  
Author(s):  
M. Bassat ◽  
S. Mokady
Keyword(s):  
Amino Acid ◽  
Cholesterol Metabolism ◽  
Cholesterol Biosynthesis ◽  
Wheat Gluten ◽  
Blood Cholesterol ◽  
Amino Acid Supplementation ◽  
Faecal Excretion ◽  
Weanling Rats ◽  
Low Levels

1. The effect of lysine- and threonine-supplemented wheat gluten on cholesterol metabolism was studied using male weanling rats. Animals were fed on cholesterol-free diets containing 100 or 200 g gluten/kg with or without amino acid supplementation, and compared with animals given 50, 100 and 200 g casein/kg diets, for 3 weeks.2. A hypocholesterolaemic effect observed with the wheat gluten-fed rats, compared with the animals given 100 and 200 g casein/kg diets, was accompanied by increased turnover of cholesterol as expressed by enhanced cholesterol biosynthesis and increased faecal excretion of cholesterol and bile acids. This effect was not abolished by lysine and threonine supplementation.3. Low levels of blood cholesterol were also observed in the rats fed on the 50 g casein/kg diet. However, a different mechanism, related to impairment of cholesterol transport from the liver, was most likely responsible for the hypocholesterolaemia found in these protein-malnourished animals.4. The effect on cholesterol metabolism produced by dietary wheat gluten was independent of the low quality of the protein and of its specific deficiency in lysine and threonine.

scholarly journals Activation of Bone Marrow-Derived Cells Angiotensin (Ang) II Type 1 Receptor by Ang II Promotes Atherosclerotic Plaque Vulnerability

2018 ◽  
Vol 19 (9) ◽  
pp. 2621
Author(s):  
Maxime Pellegrin ◽  
Karima Bouzourène ◽  
Jean-François Aubert ◽  
Aimable Nahimana ◽  
Michel Duchosal ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mrna Expression ◽  
Atherosclerotic Plaque ◽  
Cholesterol Metabolism ◽  
Mrna Levels ◽  
Ang Ii ◽  
Plaque Vulnerability ◽  
Plaque Development ◽  
Pro Inflammatory Cytokines

Angiotensin (Ang) II triggers vulnerable atherosclerotic plaque development. Bone marrow (BM)-derived cells are key players in atherogenesis but whether Ang II induces plaque vulnerability directly through Ang II type 1 receptor (AT1R) activation on these cells remains to be clarified. In the present study, we investigated whether a lack of AT1R on BM-derived cells might affect Ang II-mediated vulnerable plaque development. The 2-kidney, 1-clip (2K1C) model (Ang II-dependent mouse model of advanced atherosclerosis and vulnerable plaques) was generated in ApoE−/− mice transplanted with AT1aR−/− or AT1aR+/+ BM. Plasma cholesterol as well as hepatic mRNA expression levels of genes involved in cholesterol metabolism were significantly lower in 2K1C mice transplanted with AT1aR−/− BM than in controls. Atherosclerotic lesions were significantly smaller in AT1aR−/− BM 2K1C mice (−79% in the aortic sinus and −71% in whole aorta compared to controls). Plaques from AT1aR−/− BM 2K1C mice exhibited reduced lipid core/fibrous cap and macrophage/smooth muscle cells ratios (−82% and −88%, respectively), and increased collagen content (+70%), indicating a more stable phenotype. Moreover, aortic mRNA levels of pro-inflammatory cytokines IL-12p35, IL-1β, and TNF-α were significantly reduced in AT1aR−/− BM 2K1C mice. No significant differences in either the number of circulating Ly6Chigh inflammatory monocytes and Ly6Clow resident anti-inflammatory monocyte subsets, or in mRNA levels of aortic M1 or M2 macrophage markers were observed between the two groups. No significant differences were observed in splenic mRNA levels of T cell subsets (Th1, Th2, Th17 and Treg) markers between the two groups. In conclusion, direct AT1R activation by Ang II on BM-derived cells promotes hepatic mRNA expression of cholesterol-metabolism-related genes and vascular mRNA expression of pro-inflammatory cytokines that may lead to plaque instability.

scholarly journals Identification of a distinct low-affinity receptor for human interleukin-4 on pre-B cells

Blood ◽  
1993 ◽  
Vol 81 (11) ◽  
pp. 2998-3005 ◽  
Author(s):  
WC Fanslow ◽  
MK Spriggs ◽  
CT Rauch ◽  
KN Clifford ◽  
BM Macduff ◽  
...  
Keyword(s):  
Amino Acid ◽  
B Cells ◽  
Binding Protein ◽  
Polyclonal Antiserum ◽  
Biologically Active ◽  
Interleukin 4 ◽  
Mrna Levels ◽  
Inhibition Of Proliferation ◽  
Affinity Receptor ◽  
B Lines

Abstract Biotinylated interleukin-4 (IL-4) was used to examine IL-4 receptor (IL- 4R) expression on a range of human B-cell lines by flow cytometry. Using high concentrations of biotinylated IL-4, we have identified a novel low-affinity IL-4 receptor expressed at high levels on pre-B lines. Expression of this low-affinity receptor did not correlate with detected mRNA levels for the previously cloned receptor or with reactivity of two anti-human IL-4R monoclonal antibodies (MoAb). Radiolabeled IL-4 cross-linking studies using pre-B lines showed a doublet of 65 to 75 Kd in contrast to the 110- to 130-Kd molecule detected on cells expressing the cloned IL-4R. A soluble IL-4 binding protein (IL-4bp) was purified from the supernatants of three pre-B lines expressing the low-affinity receptor on their surface. IL-4bp could block both IL-4-mediated CD23 induction on tonsil B cells and IL- 4-induced inhibition of proliferation of the pre-B line JM1. Partial N- terminal amino acid sequence was obtained from purified IL-4bp that confirmed this protein to be novel. A 12 amino acid peptide based on the IL-4bp sequence was used to produce a polyclonal antiserum that was reactive with purified IL-4bp, and also bound to the surface of pre-B cells but not to murine CTLL cells transfected with the human IL-4R. Blocking MoAb against the previously characterized high-affinity receptor inhibited IL-4-mediated proliferation of hIL-4R+ CTLL cells but had no effect on IL-4-induced inhibition of JM1 cell proliferation, and only partially inhibited IL-4-mediated CD23 and sIgM induction and proliferation of tonsil B cells. The data presented here provide evidence for a novel cell-surface expressed low-affinity IL-4R that also exists as a biologically active soluble IL-4 binding protein.

scholarly journals The Prognostic Values of the Insulin-Like Growth Factor Binding Protein Family in Ovarian Cancer

2020 ◽  
Vol 2020 ◽  
pp. 1-15
Author(s):  
Ruoyi Zheng ◽  
Wenming Chen ◽  
Weiting Xia ◽  
Jingyu Zheng ◽  
Qing Zhou
Keyword(s):  
Ovarian Cancer ◽  
Growth Factor ◽  
Protein Expression ◽  
Mrna Expression ◽  
Binding Protein ◽  
Progression Free Survival ◽  
Prognostic Impact ◽  
Mrna Levels ◽  
Insulin Like Growth Factor ◽  
Factor Binding

Purpose. To assess the expression of insulin-like growth factor binding protein (IGFBP) family and its prognostic impact in ovarian cancer (OC) patients. Materials and Methods. The mRNA expression and protein expression of individual IGFBPs in healthy ovarian samples and OC tissues were explored through Oncomine, Gene Expression Profiling Interactive Analysis, and Human Protein Atlas database. Additionally, the prognostic values of the six IGFBP members in patients with OC were evaluated by Kaplan-Meier plotter. Results. IGFBP2 and IGFBP4 mRNA expression were remarkably upregulated in patients with OC. To be specific, the mRNA expression of IGFBP2 was upregulated in patients with serous ovarian cancer (SOC), while IGFBP1/3/4/5/6 mRNA levels were downregulated. In addition, the IGFBP4 protein expression was upregulated in SOC, and the IGFBP6 protein expression was upregulated in both of SOC and endometrioid ovarian cancer (EOC) tissues. High IGFBP1 mRNA levels showed favorable overall survival (OS) and progression-free survival (PFS) in all OC. Meanwhile, increased IGFBP5/6 mRNA levels revealed worsen OS and PFS in all OC patients. IGFBP4/6 mRNA levels predicted unfavorable OS and PFS only in SOC patients. Moreover, the aberrant mRNA expression of IGFBP1/2/4/5/6 was correlated with significantly prognosis in patients receiving different chemotherapeutic regimens. Conclusion. This study indicates that the IGFBP family reveals distinct prognosis in patients with OC. IGFBP1/2/4/5/6 are useful prognostic predictors for chemotherapeutic effect in OC patients, and IGFBP2/4 are potential tumor markers for the diagnosis of OC.

scholarly journals mRNA expression profiles for corticotrophin-releasing hormone, urocortin, CRH-binding protein and CRH receptors in human term gestational tissues determined by real-time quantitative RT-PCR

2004 ◽  
Vol 32 (2) ◽  
pp. 339-348 ◽  
Author(s):  
B Sehringer ◽  
HP Zahradnik ◽  
M Simon ◽  
R Ziegler ◽  
C Noethling ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Real Time ◽  
Binding Protein ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Releasing Hormone ◽  
Gestational Tissues ◽  
Crh Receptors

Increasing maternal plasma levels of corticotrophin-releasing hormone (CRH) during the last weeks of pregnancy suggest that this stress hormone plays an important role in the control of human parturition. Little is known about the quantitative contribution of gestational tissues (other than placenta) to intrauterine formation of CRH, urocortin and CRH-binding protein (CRH-BP), or about the distribution of CRH receptors within the uterus. We have investigated the mRNA expression of CRH, urocortin, CRH-BP and CRH receptors 1 and 2 (CRH-R1 and -R2) in gestational tissues by real-time RT-PCR. Placenta, myometrium and choriodecidua were collected after uncomplicated pregnancies at term, before the onset of labour. Distribution of CRH-R1 and CRH-R2 protein was also investigated by immunostaining with receptor subtype-specific antibodies. The placenta was identified as the main site of CRH and CRH-BP mRNA expression, displaying mRNA levels >1000 and >20 times higher than those found in the myometrium and choriodecidua respectively (P<0.05 in each case). mRNA expression of urocortin was low in all tissues investigated. Myometrium and choriodecidua expressed relevant amounts of both receptor subtypes, whereas the CRH receptor population in placenta consisted mainly of CRH-R2. The high expression of CRH in placenta and the substantial expression of CRH receptors in choriodecidua and myometrium suggested that CRH derived from placenta exerts direct or indirect actions on these tissues. Neither CRH produced by myometrium or choriodecidua nor urocortin from other intrauterine sources seem to play a major role in the control of labour.

scholarly journals Characterization of a full-length cDNA encoding human liver S-adenosylmethionine synthetase: tissue-specific gene expression and mRNA levels in hepatopathies

1993 ◽  
Vol 293 (2) ◽  
pp. 481-486 ◽  
Author(s):  
L Alvarez ◽  
F Corrales ◽  
A Martín-Duce ◽  
J M Mato
Keyword(s):  
Amino Acid ◽  
Human Liver ◽  
Structural Features ◽  
Full Length ◽  
Northern Analysis ◽  
Specific Gene ◽  
Mrna Levels ◽  
Calculated Molecular Mass ◽  
Full Length Cdna ◽  
Adenosylmethionine Synthetase

The sequence of a full-length cDNA coding for human liver S-adenosylmethionine synthetase has been determined. It spans 3217 nucleotides and encodes a protein of 395 amino acid residues, with a calculated molecular mass of 43,647 Da. The structural features deduced from the amino acid sequence show a close similarity to those of the rat liver enzyme. The liver-specific S-adenosylmethionine synthetase gene appears to be present as a single copy in the genome, as revealed by Southern analysis. The occurrence of a single mRNA species for this enzyme has been determined by primer extension and Northern analysis. Among several human tissues examined, this gene is expressed only in the liver. Similar S-adenosylmethionine synthetase mRNA levels have been detected in biopsies from normal human liver and from patients with alcoholic cirrhosis and hepatocellular carcinoma. Based on these results, a possible mechanism of regulation of human liver S-adenosylmethionine synthetase is discussed.

scholarly journals Modulation of Cholesterol Metabolism Improves Response to Enzalutamide Treatment in Prostate Cancer

2021 ◽  
Vol 5 (Supplement_2) ◽  
pp. 269-269
Author(s):  
Sora Kim ◽  
Kee-Hong Kim
Keyword(s):  
Prostate Cancer ◽  
Mrna Expression ◽  
Cholesterol Metabolism ◽  
Mrna Level ◽  
Mrna Levels ◽  
Index Method ◽  
Cell Counts ◽  
Potential Biomarker ◽  
Ester Formation ◽  
Short Period

Abstract Objectives Prostate cancer (PCa) growth is mediated by androgens via activation of androgen receptor (AR). Accordingly, androgen deprivation therapy (ADT) is the gold standard for the treatment of advanced PCa, but progression to castration-resistant PCa (CRPC) follows. Enzalutamide (ENZ) is an AR antagonist used for the management of CRPC. However, patients acquire resistance to the drug in a short period. As cholesterol metabolism is dysregulated in PCa and lipogenesis is upregulated by AR signaling, we hypothesized that inhibition of cholesteryl ester formation and suppression of lipogenesis by blockage of sterol-O-acyltransferase 1 (SOAT1) could enhance the response to ENZ. Methods The mRNA expression of SOAT1 in PCa patients was analyzed using Oncomine and TCGA database. Survival analysis of TCGA PCa patients data was executed using cBioPortal. 22RV1 cells inherently resistant to ENZ, were challenged to avasimibe (AVA), a pharmacological inhibitor of SOAT1, with or without ENZ. Results We found SOAT1 mRNA is significantly overexpressed in prostate carcinoma compared to the normal tissue in separate datasets (Taylor 3; Grasso; Liu). Analysis of the TCGA PCa dataset among patients who had undergone ADT showed longer disease-free status (P = 0.039) and a trend toward better overall survival status (P = 0.066) in low SOAT1 mRNA expressing patients. Moreover, SOAT1 mRNA expression positively correlated with AR mRNA expression with Pearson coefficient of 0.5 (P = 7.475e-6). We also found that combination of 20 μM ENZ and 2 μM AVA reduced 52% of cell counts after 5 days and 53% of colony formation after 2 weeks, whereas AVA resulted in 20% and 28% reduction, respectively, with no effect from ENZ. Synergism of these two drugs was observed based on MTT assay with Chou-Talalay's combination index method. 4 μM AVA treatment for 24h downregulated AR downstream nkx3.1 mRNA level with no difference in AR or ARV7 mRNA levels. AVA treatment also downregulated lipogenic SCD1 mRNA level, inhibition of which was previously reported to enhance response to ENZ. Conclusions SOAT1 is a potential biomarker predictive of the success of ENZ treatment in PCa according to TCGA data. Our in vitro study further supports that SOAT1-regulated cholesterol metabolism is an important factor for the ENZ response. Funding Sources Purdue Research Foundation, Ralph W. and Grace M. Showalter Research Trust.

scholarly journals Effects of alfalfa saponin extract on the performance and cholesterol metabolism of laying hens

2015 ◽  
Author(s):  
Senhao Zhang ◽  
Yinghua Shi ◽  
Minggen Liang ◽  
Jia Li ◽  
Chengzhang Wang
Keyword(s):  
Bile Acid ◽  
Mrna Expression ◽  
Laying Hens ◽  
Cholesterol Metabolism ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Mrna Levels ◽  
Density Lipoprotein Receptor ◽  
Yolk Cholesterol

The experiment was performed to determine the effects of alfalfa saponin extract (ASE) on the performance and cholesterol metabolism of laying hens. A total of 150 Hy-Line Brown hens with 28 weeks old, were randomly divided into five treatment groups (five replicates per treatment with six hens per replicate). Diets containing 0, 60, 120, 240, and 480 mg ASE/kg were fed to hens for 77 days. The shell thickness had a trend to increase. The yolk cholesterol and liver bile acid decreased significantly (ASE 60 and 480 mg/kg groups for yolk cholesterol, and ASE 60 and 240 mg/kg groups for liver bile acid). Fecal bile acid has an elevation trend as ASE increased. The expression of very low density apolipoprotein-Ⅱ (apoVLDL-Ⅱ) gene was not affected by adding ASE. However, the mRNA expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase gene and cholesterol 7α-hydroxylase (CYP7A1) gene were significantly up-regulated. The mRNA expression of very-low-density-lipoprotein receptor(VLDLR) gene was suppressed due to adding ASE supplementation in the diet. These findings indicated that dietary ASE could regulate cholesterol levels in hens by up-regulating the mRNA levels of HMG-CoA and CYP7A1 and suppressing the expression of VLDLR.

scholarly journals Effects of alfalfa saponin extract on the performance and cholesterol metabolism of laying hens

2015 ◽  
Author(s):  
Senhao Zhang ◽  
Yinghua Shi ◽  
Minggen Liang ◽  
Jia Li ◽  
Chengzhang Wang
Keyword(s):  
Bile Acid ◽  
Mrna Expression ◽  
Laying Hens ◽  
Cholesterol Metabolism ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Mrna Levels ◽  
Density Lipoprotein Receptor ◽  
Yolk Cholesterol

The experiment was performed to determine the effects of alfalfa saponin extract (ASE) on the performance and cholesterol metabolism of laying hens. A total of 150 Hy-Line Brown hens with 28 weeks old, were randomly divided into five treatment groups (five replicates per treatment with six hens per replicate). Diets containing 0, 60, 120, 240, and 480 mg ASE/kg were fed to hens for 77 days. The shell thickness had a trend to increase. The yolk cholesterol and liver bile acid decreased significantly (ASE 60 and 480 mg/kg groups for yolk cholesterol, and ASE 60 and 240 mg/kg groups for liver bile acid). Fecal bile acid has an elevation trend as ASE increased. The expression of very low density apolipoprotein-Ⅱ (apoVLDL-Ⅱ) gene was not affected by adding ASE. However, the mRNA expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase gene and cholesterol 7α-hydroxylase (CYP7A1) gene were significantly up-regulated. The mRNA expression of very-low-density-lipoprotein receptor(VLDLR) gene was suppressed due to adding ASE supplementation in the diet. These findings indicated that dietary ASE could regulate cholesterol levels in hens by up-regulating the mRNA levels of HMG-CoA and CYP7A1 and suppressing the expression of VLDLR.

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