| 1. |
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| 2. |
- Lundqvist, Johan, et al.
(författare)
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1α,25-Dihydroxyvitamin D3 exerts tissue-specific effects on estrogen and androgen metabolism
- 2011
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Ingår i: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids. - : Elsevier BV. - 1388-1981 .- 1879-2618. ; 1811:4, s. 263-270
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Tidskriftsartikel (refereegranskat)abstract
- It is well-known that 1α,25-dihydroxyvitamin D(3) and analogs exert anti-proliferative and pro-differentiating effects and these compounds have therefore been proposed to be of potential use as anti-cancer agents. Due to its effects on aromatase gene expression and enzyme activity, 1α,25-dihydroxyvitamin D(3) has been proposed as an interesting substance in breast cancer treatment and prevention. In the present study, we have examined the effects of 1α,25-dihydroxyvitamin D(3) on estrogen and androgen metabolism in adrenocortical NCI-H295R cells, breast cancer MCF-7 cells and prostate cancer LNCaP cells. The NCI-H295R cell line has been proposed as a screening tool to study endocrine disruptors. We therefore studied whether this cell line reacted to 1α,25-dihydroxyvitamin D(3) treatment in the same way as cells from important endocrine target tissues. 1α,25-Dihydroxyvitamin D(3) exerted cell line-specific effects on estrogen and androgen metabolism. In breast cancer MCF-7 cells, aromatase gene expression and estradiol production were decreased, while production of androgens was markedly increased. In NCI-H295R cells, 1α,25-dihydroxyvitamin D(3) stimulated aromatase expression and decreased dihydrotestosterone production. In prostate cancer LNCaP cells, aromatase expression increased after the same treatment, as did production of testosterone and dihydrotestosterone. In summary, our data show that 1α,25-dihydroxyvitamin D(3) exerts tissue-specific effects on estrogen and androgen production and metabolism. This is important knowledge about 1α,25-dihydroxyvitamin D(3) as an interesting substance for further research in the field of breast cancer prevention and treatment. Furthermore, the observed cell line-specific effects are of importance in the discussion about NCI-H295R cells as a model for effects on estrogen and androgen metabolism.
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| 3. |
- Lundqvist, Johan, 1979-
(författare)
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Enzymatic Regulation of Steroidogenesis and Nuclear Receptor Activation : Special Focus on Vitamin D and Sex Hormones
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Enzyme-catalyzed reactions are important to regulate steroidogenesis and nuclear receptor activation. The present investigation examines the role of steroid metabolism catalyzed by CYP7B1 for regulation of hormone receptor activation and the effects of vitamin D on enzymatic regulation of steroidogenesis. The study reports data indicating that CYP7B1 can regulate estrogenic signaling by converting estrogens into inactive or less active metabolites. Similar results were obtained for CYP7B1-mediated metabolism of some androgen receptor ligands, indicating that CYP7B1 can be involved also in the regulation of androgenic signaling. CYP7B1 substrates and metabolites were found to exert androgenic effects in a cell line-specific manner. Furthermore, cell line differences were observed in the expression pattern for androgen receptor comodulators. This thesis reports that 1α,25-dihydroxyvitamin D3 alters the gene expression and enzyme activity of CYP21A2 and CYP17A1 leading to suppressed production of aldosterone, dehydroepiandrosterone and androstenedione in adrenocortical cells. These are novel findings on vitamin D action. A mechanism is reported for the vitamin D-mediated regulation of the CYP21A2 gene. Data indicate that vitamin D receptor interacting repressor (VDIR) and Williams syndrome transcription factor (WSTF) are key comodulators in this novel vitamin D receptor (VDR)-mediated mechanism. Furthermore, the results indicate that altered expression levels of VDIR and WSTF can shift the suppressing effect of vitamin D to a stimulatory effect. Also, epigenetic components were found to be involved in the effects of vitamin D on CYP21A2 transcriptional rate. In addition, a functional vitamin D response element was identified in the CYP21A2 promoter. This study also reports that 1α,25-dihydroxyvitamin D3 affects sex hormone production in a tissue-specific way. Gene expression and enzyme activity of aromatase were found to be downregulated in cells derived from breast, but not in cells derived from prostate and adrenal cortex. The production of estradiol and dihydrotestosterone was altered in a tissue-selective manner following vitamin D treatment. These findings are of importance for the discussion on vitamin D as a potential anti-breast cancer agent.
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| 4. |
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| 5. |
- Norlin, Maria
(författare)
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Cytochrome P450 Enzymes in the Metabolism of Cholesterol and Cholesterol Derivatives
- 2000
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cholesterol is metabolized to a variety of important biological products in the body including bile acids and vitamin D. The present investigation is focused on enzymes that catalyze 7α-hydroxylation or 27-hydroxylation in the metabolism of cholesterol, oxysterols (side chain-hydroxylated derivatives of cholesterol) and vitamin D3. The enzymes studied belong to the cytochrome P450 enzyme families CYP7 and CYP27.The study describes purification of a cytochrome P450 enzyme fraction active in 7α-hydroxylation of 25-hydroxycholesterol, 27-hydroxycholesterol, dehydroepiandrosterone and pregnenolone from pig liver microsomes. Peptide sequence analysis indicated that this enzyme fraction contains an enzyme belonging to the CYP7B subfamily. The purified enzyme was not active towards cholesterol or testosterone. Purification and inhibition experiments suggested that hepatic microsomal 7α -hydroxylation of 27-hydroxycholesterol and dehydroepiandrosterone involves at least two enzymes, probably closely related.The study shows that recombinantly expressed human and rat cholesterol 7α -hydroxylase (CYP7A) and partially purified pig liver cholesterol 7α -hydroxylase are active towards 20(S)-, 24-, 25- and 27-hydroxycholesterol. CYP7A was previously considered specific for cholesterol and cholestanol. The 7α -hydroxylation of 20(S)-, 25-, and 27-hydroxycholesterol in rat liver was significantly increased by treatment with cholestyramine, an inducer of CYP7A. Cytochrome P450 of renal origin showed 7α -hydroxylase activity towards 25- and 27-hydroxycholesterol, dehydroepiaundrosterone and pregnenolone but not towards 20(S)-, 24-hydroxycholesterol or cholesterol. The results indicate a physiological role for CYP7A as an oxysterol 7α -hydroxylase, in addition to the previously known human oxysterol 7α -hydroxylase CYP7B.The role of renal sterol 27-hydroxylase (CYP27A) in the bioactivation of vitamin D3 was studied with cytochrome P450 fractions purified from pig kidney mitochondria. Purification and inhibition experiments and experiments with a monoclonal antibody against CYP27A indicated that CYP27A plays a role in renal 25-hydroxyvitamin D3 l α -hydroxylation.The expression of CYP7A, CYP7B and CYP27A during development was studied. The levels of CYP27A in livers of newborn and six months old pigs were similar whereas the levels of CYP7A increased. The expression of CYP7B varied depending on the tissue. The expression of CYP7B increased with age in the liver whereas the CYP7B levels in kidney showed a marked age-dependent decrease.
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| 6. |
- Norlin, Maria, et al.
(författare)
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Enzymes in the conversion of cholesterol into bile acids
- 2007
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Ingår i: Current molecular medicine. - : Bentham Science Publishers Ltd.. - 1566-5240 .- 1875-5666. ; 7:2, s. 199-218
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Forskningsöversikt (refereegranskat)abstract
- This article aims to give an overview on the characterization, properties and regulation of enzymes, particularly the cytochrome (CYP) P450 enzymes, in the formation of bile acids from cholesterol. Bile acids are biologically active molecules that promote absorption of dietary lipids in the intestine and stimulate biliary excretion of cholesterol. Bile acids and oxysterols, formed from cholesterol, act as ligands to nuclear receptors regulating the expression of important genes in cholesterol homeostasis. Thus, the bioactivation of cholesterol into bile acids is crucial for regulation of cholesterol homeostasis. The primary human bile acids, cholic acid and chenodeoxycholic acid, are formed from cholesterol via several pathways involving many different enzymes. Many of these enzymes are cytochrome P450 (CYP) enzymes, introducing a hydroxyl group in the molecule. The “classic” pathway of bile acid formation starts with a 7α-hydroxylation of cholesterol by CYP7A1 in the liver. The “acidic” pathway starts with a hepatic or extrahepatic 27-hydroxylation by CYP27A1. There also exist some quantitatively minor pathways which may be of importance under certain conditions. Formation of cholic acid requires insertion of a 12α-hydroxyl group performed by CYP8B1. Oxysterols are precursors to bile acids, participate in cholesterol transport and are known to affect the expression of several genes in cholesterol homeostasis. Enzymes with capacity to form and metabolize oxysterols are present in liver and extrahepatic tissues. The enzymes, nuclear receptors and transcription factors involved in bile acid biosynthesis are potential pharmaceutical targets for the development of new drugs to control hypercholesterolemia and to prevent atherosclerosis and other diseases related to disturbed cholesterol homeostasis. The review will also discuss some inborn errors of bile acid biosynthesis and the recently acquired knowledge on the genetic defects underlying these diseases.
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| 7. |
- Pettersson, Hanna, 1968-
(författare)
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Steroid-Metabolizing Cytochrome P450 (CYP) Enzymes in the Maintenance of Cholesterol and Sex Hormone Levels
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The enzymes CYP27A1 and CYP7B1 are widely expressed in various human tissues and perform catalytic reactions in cholesterol homeostasis and endocrine signaling. We have investigated the metabolism of a synthetic oxysterol. In this study, we show that CYP27A1 is the enzyme responsible for a 28-hydroxylation of this oxysterol and that the rate of CYP27A1-mediated metabolism is relatively slow. This may give an explanation for the prolonged inhibitory effects on cholesterol biosynthesis that have been shown for this oxysterol. The current study contributes to the knowledge of synthetically produced oxysterols and their potential use as cholesterol lowering drugs. In two studies we investigated CYP7B1-mediated metabolism of different sex hormones. Our data indicate that CYP7B1 may carry out a previously unknown catalytic reaction involving an androgen. Taken together the data suggest that varying steroid concentrations in cells and tissues may be important for CYP7B1-dependent metabolism of sex hormones and sex hormone precursors. CYP7B1-mediated hydroxylation of sex hormones may influence the cellular levels of these steroids and may be a potential pathway for elimination of the steroids from the cell. Some known CYP7B1 substrates are agonists for ERα and ERβ but the reported role(s) of CYP7B1 for ER action are not fully understood. In the last study we investigated the role(s) of CYP7B1-mediated metabolism for ER-mediated action. Our data indicate that CYP7B1-mediated conversion of steroids that affect ER-mediated response into their 7α-hydroxymetabolites will result in loss of action. This indicates that CYP7B1 may have an important role for regulation of ER-mediated processes in the body. In summary, results from this thesis contribute to the knowledge on the metabolism of synthetic oxysterols of potential use as cholesterol lowering drugs and the role(s) of CYP7B1-mediated metabolism for processes related to the functions of sex hormones.
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| 8. |
- Svenningsen, Åsa Fex, et al.
(författare)
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Effects on DHEA levels by estrogen in rat astrocytes and CNS co-cultures via the regulation of CYP7B1-mediated metabolism
- 2011
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Ingår i: Neurochemistry International. - : Elsevier BV. - 0197-0186 .- 1872-9754. ; 58:6, s. 620-624
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Tidskriftsartikel (refereegranskat)abstract
- The neurosteroid dehydroepiandrosterone (DHEA) is formed locally in the CNS and has been implicated in several processes essential for CNS function, including control of neuronal survival. An important metabolic pathway for DHEA in the CNS involves the steroid hydroxylase CYP7B1. In previous studies, CYP7B1 was identified as a target for estrogen regulation in cells of kidney and liver. In the current study, we examined effects of estrogens on CYP7B1-mediated metabolism of DHEA in primary cultures of rat astrocytes and co-cultures of rat CNS cells. Astrocytes, which interact with neurons in several ways, are important for brain neurosteroidogenesis. We found that estradiol significantly suppressed CYP7B1-mediated DHEA hydroxylation in primary mixed CNS cultures from fetal and newborn rats. Also, CYP7B1-mediated DHEA hydroxylation and CYP7B1 mRNA were markedly suppressed by estrogen in primary cultures of rat astrocytes. Interestingly, diarylpropionitrile, a well-known agonist of estrogen receptor β, also suppressed CYP7B1-mediated hydroxylation of DHEA. Several previous studies have reported neuroprotective effects of estrogens. The current data indicate that one of the mechanisms whereby estrogen can exert protective effects in the CNS may involve increase of the levels of DHEA by suppression of its metabolism.
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| 9. |
- Tang, Wanjin, et al.
(författare)
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Regulation of steroid hydroxylase CYP7B1 by androgens and estrogens in prostate cancer LNCaP cells
- 2006
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 344:2, s. 540-546
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Tidskriftsartikel (refereegranskat)abstract
- The present study reports effects of androgens and estrogens on human CYP7B1 transcription in prostate cancer LNCaP cells. Studies with rodents have suggested a role for the CYP7B1 enzyme in balancing cellular hormone levels important for prostate growth. Little is, however, known about the regulation of human CYP7B1. The current study showed strong suppression of a human CYP7B1 luciferase reporter gene by dihydrotestosterone (DHT) in prostate cancer LNCaP cells. Also, DHT and overexpression of androgen receptor (AR) suppressed CYP7B1 promoter activity and CYP7B1-mediated catalysis in kidney-derived HEK293 cells. Effects on CYP7B1 transcription were observed also by estrogen receptors (ER). The effects appeared different for different estrogens. CYP7B1 was stimulated by synthetic ER agonists but suppressed by 17beta-estradiol and 5alpha-androstane-3beta,17beta-diol in LNCaP cells. Our data indicate an important role for CYP7B1 in balancing prostate hormone levels in human cells. In particular, the data suggest that androgens may control intraprostatic levels of estrogen via regulation of CYP7B1-mediated metabolism.
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| 10. |
- Ellfolk, Maria, 1976-
(författare)
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Regulation of Vitamin D 25-hydroxylases : Effects of Vitamin D Metabolites and Pharmaceutical Compounds on the Bioactivation of Vitamin D
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- A 700bp portion of the promoter of CYP2D25, the porcine microsomal vitamin D 25-hydroxylase was isolated and sequenced. The computer analysis of the sequence revealed the existence of a putative VDRE at 220 bp upstream of the transcription start site. A CYP2D25 promoter-luciferase reporter plasmid was constructed in order to study the transcriptional regulation of the gene. Treatment with the vitamin D metabolites calcidiol and calcitriol suppressed the promoter, provided that the nuclear receptors VDR and RXR were overexpressed. Phenobarbital was also capable of suppressing the promoter if the nuclear receptors PXR or CAR were overexpressed.The 25-hydroxylases are not expressed solely in liver but in a wide array of other organs as well. It is therefore possible at least in theory to study the vitamin D 25-hydroxylation in human subjects using cells from extrahepatic organs, from which biopsy retrieval is easier than from the liver. Dermal fibroblasts are frequently used to study different pathological conditions in human subjects and they are easy to come by. Dermal fibroblasts were shown to express two vitamin D 25-hydroxylases: CYP27A1 and CYP2R1. The expression pattern of CYP2R1 displayed considerable interindividual variation. The fibroblasts were also capable of measurable vitamin D 25-hydroxylation, which makes dermal fibroblasts a possible tool in studying vitamin D 25-hydroxylation in human subjects.Little is known about the regulation of expression and activity of the human vitamin D 25-hydroxylases. Therefore dermal fibroblasts – expressing CYP2R1 and CYP27A1 – and human prostate cancer LNCaP cells, that express CYP2R1 and CYP2J2, were treated with calcitriol and phenobarbital and efavirenz, two drugs that give rise to vitamin D deficiency. Treatment decreased the mRNA levels of CYP2R1 and CYP2J2 provided that the treated cells also expressed the necessary nuclear receptors. CYP27A1 did not respond to any of the treatments. The treatments also managed to decrease the 25-hydroxylating activity of the cells.The results show that vitamin D 25-hydroxylases can be regulated by both endogenous and xenobiotic compounds.
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