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1.
Biochem Med (Zagreb) ; 32(2): 020706, 2022 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-35799985

RESUMO

Introduction: The aim of this study was to investigate lipoprotein particle distributions and the likelihood of achieving cholesterol homeostasis in the remission phase of nephrotic syndrome (NS) in paediatric patients. We hypothesized that lipoprotein particle distributions moved toward less atherogenic profile and that cholesterol homeostasis was achieved. Materials and methods: Thirty-three children, 2 to 9 years old with NS were recruited. Blood sampling took place both in the acute phase and during remission. Serum low-density lipoprotein particles (LDL) and high-density lipoprotein particles (HDL) were separated using non-denaturing polyacrylamide gradient gel (3-31%) electrophoresis. Serum non-cholesterols sterols (NCSs), desmosterol, lathosterol, 7-dehydrocholesterol (7-DHC), campesterol and ß-sitosterol were measured by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Results: All patients had desirable serum HDL cholesterol concentrations during remission. The dominant lipoprotein diameters and LDL subclass distribution did not change significantly during follow-up. In contrast, HDL lipoprotein particle distribution shifted towards larger particles. The absolute concentration of desmosterol was significantly lower during remission (P = 0.023). ß-sitosterol concentration markedly increased during remission (P = 0.005). Desmosterol/ß-sitosterol (P < 0.001) and 7-DHC/ß-sitosterol (P = 0.005) ratios significantly declined during disease remission. Conclusions: Favourable changes in the serum lipid profiles, HDL particle subclass distribution and cholesterol metabolism in paediatric patients with NS during remission took place. For the first time, we found that cholesterol homeostasis changed in favour of increased cholesterol absorption during disease remission. Nevertheless, complete cholesterol homeostasis was not achieved during disease remission.


Assuntos
Desmosterol , Síndrome Nefrótica , Criança , Pré-Escolar , HDL-Colesterol , Humanos , Lipoproteínas , Espectrometria de Massas em Tandem
2.
Dis Markers ; 2022: 2008556, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35493299

RESUMO

The cholesterol metabolism in humans can be indirectly reflected by measuring cholesterol metabolism marker levels. We aimed to investigate the association of cholesterol homeostasis markers on standard lipid profiling components in familial hypercholesteremia and hyperlipidemia patients. A total of 69 hyperlipidemia patients, 25 familial hypercholesteremia (FHC) patients, and 64 healthy controls were enrolled in this study. We performed routine testing of blood lipid water. Gas chromatography was used to determine the changes in the concentration of cholesterol synthesis (squalene, desmosterol, and lathosterol) and absorption markers (campesterol, sitosterol, and stigmasterol) in the blood. Baseline hyperlipidemia patients displayed significantly higher total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) levels in comparison to the control group, which was reflected in the increased levels of squalene, desmosterol, campesterol, and sitosterol observed (P < 0.05) in the hyperlipidemia patients. The desmosterol, lathosterol, campesterol, stigmasterol, and sitosterol were statistically different in the FHC group than the hyperlipidemic group (P < 0.05). The proportions of squalene/cholesterol, lathosterol/cholesterol, stigmasterol/cholesterol, and sitosterol/cholesterol in the FHC group were lower than those in the hyperlipidemic group; only desmosterol/cholesterol was higher than that in the hyperlipidemic group. Correlation studies between lipid metabolic factors showed that the proportion of moderate and strong correlations was much higher in the FHC group than in the other two groups (76.92% vs. 32.50% and 31.25%). Logistic regression analysis showed that the concentrations of glucose, LDL-C, lactosterol, and sitosterol were all independent risk factors for developing hyperlipidemia. This result was further confirmed by the ROC curve. These results indicated that the study of cholesterol synthesis and decomposition markers can serve as a reference index for related diseases caused by changes in its concentration.


Assuntos
Hipercolesterolemia , Hiperlipidemias , Hiperlipoproteinemia Tipo II , Colesterol , LDL-Colesterol , Desmosterol , Humanos , Hiperlipoproteinemia Tipo II/diagnóstico , Lipídeos , Sitosteroides , Esqualeno , Estigmasterol
3.
Transl Psychiatry ; 12(1): 139, 2022 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-35379782

RESUMO

Major depressive disorder (MDD) is a common, disabling, and heterogeneous condition that responds unpredictably to current treatments. We previously showed an association between depressive symptoms and plasma concentrations of two cholesterol precursors, desmosterol and 7-dehydrocholesterol (7DHC). Here, we measured total cholesterol and sterol concentrations with mass spectrometry in postmortem brain samples from depressed and control subjects. Mean (±SEM) desmosterol concentration was 8.9 ± 0.97 ng/mg in the depressed versus 10.7 ± 0.72 ng/mg in the control group. The mean of the posterior probability distribution for the difference in desmosterol concentration between the two groups was 2.36 (95% highest density interval [HDI] 0.59-4.17). Mean 7DHC concentrations, 12.5 ± 4.1 ng/mg in the depressed versus 5.4 ± 0.74 ng/mg in the control group, were unlikely to be different (95% HDI, [-1.37-0.34]). We found that presence of trazodone in the peri-mortem toxicology screen accounted for the observed difference in desmosterol concentrations. We also observed extremely high 7DHC levels in all 4 subjects who had taken trazodone. Trazodone has been recently found to inhibit 7-dehydrocholesterol reductase and alter sterol concentrations in rodents, cell culture, human fibroblasts, and blood. In this study, we demonstrate for the first time that trazodone alters human brain sterol composition. Given congenital deficiency of 7-dehydrocholesterol reductase results in Smith-Lemli-Opitz syndrome, our findings support the hypothesis that this commonly used medication may have previously unappreciated risks.


Assuntos
Transtorno Depressivo Maior , Trazodona , Encéfalo , Desidrocolesteróis , Desmosterol , Humanos , Trazodona/farmacologia
4.
Int J Mol Sci ; 23(8)2022 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-35456988

RESUMO

Squalene is a natural bioactive triterpene and an important intermediate in the biosynthesis of sterols. To assess the effect of this compound on the hepatic transcriptome, RNA-sequencing was carried out in two groups of male New Zealand rabbits fed either a diet enriched with 1% sunflower oil or the same diet with 0.5% squalene for 4 weeks. Hepatic lipids, lipid droplet area, squalene, and sterols were also monitored. The Squalene administration downregulated 9 transcripts and upregulated 13 transcripts. The gene ontology of transcripts fitted into the following main categories: transporter of proteins and sterols, lipid metabolism, lipogenesis, anti-inflammatory and anti-cancer properties. When the results were confirmed by RT-qPCR, rabbits receiving squalene displayed significant hepatic expression changes of LOC100344884 (PNPLA3), GCK, TFCP2L1, ASCL1, ACSS2, OST4, FAM91A1, MYH6, LRRC39, LOC108176846, GLT1D1 and TREH. A squalene-enriched diet increased hepatic levels of squalene, lanosterol, dihydrolanosterol, lathosterol, zymostenol and desmosterol. Strong correlations were found among specific sterols and some squalene-changed transcripts. Incubation of the murine AML12 hepatic cell line in the presence of lanosterol, dihydrolanosterol, zymostenol and desmosterol reproduced the observed changes in the expressions of Acss2, Fam91a1 and Pnpla3. In conclusion, these findings indicate that the squalene and post-squalene metabolites play important roles in hepatic transcriptional changes required to protect the liver against malfunction.


Assuntos
Lanosterol , Esqualeno , Aciltransferases , Animais , Desmosterol/metabolismo , Desmosterol/farmacologia , Lanosterol/farmacologia , Fígado/metabolismo , Masculino , Camundongos , Fosfolipases A2 Independentes de Cálcio/metabolismo , Coelhos , Esqualeno/farmacologia , Esteróis/metabolismo , Transcriptoma
5.
Proc Natl Acad Sci U S A ; 118(47)2021 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-34782454

RESUMO

Cholesterol biosynthetic intermediates, such as lanosterol and desmosterol, are emergent immune regulators of macrophages in response to inflammatory stimuli or lipid overloading, respectively. However, the participation of these sterols in regulating macrophage functions in the physiological context of atherosclerosis, an inflammatory disease driven by the accumulation of cholesterol-laden macrophages in the artery wall, has remained elusive. Here, we report that desmosterol, the most abundant cholesterol biosynthetic intermediate in human coronary artery lesions, plays an essential role during atherogenesis, serving as a key molecule integrating cholesterol homeostasis and immune responses in macrophages. Depletion of desmosterol in myeloid cells by overexpression of 3ß-hydroxysterol Δ24-reductase (DHCR24), the enzyme that catalyzes conversion of desmosterol to cholesterol, promotes the progression of atherosclerosis. Single-cell transcriptomics in isolated CD45+CD11b+ cells from atherosclerotic plaques demonstrate that depletion of desmosterol increases interferon responses and attenuates the expression of antiinflammatory macrophage markers. Lipidomic and transcriptomic analysis of in vivo macrophage foam cells demonstrate that desmosterol is a major endogenous liver X receptor (LXR) ligand involved in LXR/retinoid X receptor (RXR) activation and thus macrophage foam cell formation. Decreased desmosterol accumulation in mitochondria promotes macrophage mitochondrial reactive oxygen species production and NLR family pyrin domain containing 3 (NLRP3)-dependent inflammasome activation. Deficiency of NLRP3 or apoptosis-associated speck-like protein containing a CARD (ASC) rescues the increased inflammasome activity and atherogenesis observed in desmosterol-depleted macrophages. Altogether, these findings underscore the critical function of desmosterol in the atherosclerotic plaque to dampen inflammation by integrating with macrophage cholesterol metabolism and inflammatory activation and protecting from disease progression.


Assuntos
Aterosclerose/tratamento farmacológico , Desmosterol/farmacologia , Inflamassomos/metabolismo , Inflamação/tratamento farmacológico , Ativação de Macrófagos/efeitos dos fármacos , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Colesterol/metabolismo , Vasos Coronários , Células Espumosas/metabolismo , Humanos , Inflamação/metabolismo , Metabolismo dos Lipídeos , Receptores X do Fígado/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Placa Aterosclerótica/metabolismo , Esteróis/metabolismo
6.
Nat Neurosci ; 24(1): 47-60, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33349711

RESUMO

The repair of inflamed, demyelinated lesions as in multiple sclerosis (MS) necessitates the clearance of cholesterol-rich myelin debris by microglia/macrophages and the switch from a pro-inflammatory to an anti-inflammatory lesion environment. Subsequently, oligodendrocytes increase cholesterol levels as a prerequisite for synthesizing new myelin membranes. We hypothesized that lesion resolution is regulated by the fate of cholesterol from damaged myelin and oligodendroglial sterol synthesis. By integrating gene expression profiling, genetics and comprehensive phenotyping, we found that, paradoxically, sterol synthesis in myelin-phagocytosing microglia/macrophages determines the repair of acutely demyelinated lesions. Rather than producing cholesterol, microglia/macrophages synthesized desmosterol, the immediate cholesterol precursor. Desmosterol activated liver X receptor (LXR) signaling to resolve inflammation, creating a permissive environment for oligodendrocyte differentiation. Moreover, LXR target gene products facilitated the efflux of lipid and cholesterol from lipid-laden microglia/macrophages to support remyelination by oligodendrocytes. Consequently, pharmacological stimulation of sterol synthesis boosted the repair of demyelinated lesions, suggesting novel therapeutic strategies for myelin repair in MS.


Assuntos
Doenças Desmielinizantes/patologia , Microglia/fisiologia , Esteróis/biossíntese , Animais , Colesterol/metabolismo , Desmosterol/metabolismo , Encefalomielite Autoimune Experimental , Feminino , Perfilação da Expressão Gênica , Humanos , Inflamação/metabolismo , Inflamação/patologia , Metabolismo dos Lipídeos , Receptores X do Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Esclerose Múltipla , Oligodendroglia/metabolismo , Fagocitose , Esqualeno/metabolismo
7.
J Dairy Sci ; 104(2): 1378-1383, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33189282

RESUMO

We investigated the main sterols, phytosterols, and the α- and γ-tocopherol content in donkey milk during the first 2 mo of lactation. Cholesterol was the main sterol in milk (mean ± standard deviation = 0.97 ± 0.443 g/100 g of fat). Lanosterol was the main minor sterol of animal origin, followed by desmosterol (0.003 ± 0.001 and 0.001 ± 0.001 g/100 g of fat, respectively). Of the phytosterols, ß-sitosterol was the main sterol of vegetal origin in donkey milk (0.005 ± 0.002 g/100 g of fat), but lower levels of campesterol, brassicasterol, and stigmasterol were also recorded. Mean levels of α- and γ-tocopherol were 0.01 ± 0.007 and 0.003 ± 0.001 g/100 g of fat, respectively. We observed no significant changes in sterol or tocopherol content during the first 2 mo of lactation. The presence of lanosterol in donkey milk is of particular interest, because lanosterol is a potential drug and has important physiological effects. The presence of phytosterols, which are considered nutraceutical molecules, enhances the nutritional quality of donkey milk fat for consumers.


Assuntos
Equidae , Leite/química , Fitosteróis/análise , Tocoferóis/análise , Animais , Colestadienóis , Colesterol/análogos & derivados , Colesterol/análise , Desmosterol/análise , Feminino , Lactação/metabolismo , Lanosterol/análise , Valor Nutritivo , Saponinas/metabolismo , Sitosteroides , Esteróis
8.
Molecules ; 25(18)2020 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-32916848

RESUMO

We developed a simple and robust liquid chromatographic/mass spectrometric method (LC-MS) for the quantitative analysis of 10 sterols from the late part of cholesterol synthesis (zymosterol, dehydrolathosterol, 7-dehydrodesmosterol, desmosterol, zymostenol, lathosterol, FFMAS, TMAS, lanosterol, and dihydrolanosterol) from cultured human hepatocytes in a single chromatographic run using a pentafluorophenyl (PFP) stationary phase. The method also avails on a minimized sample preparation procedure in order to obtain a relatively high sample throughput. The method was validated on 10 sterol standards that were detected in a single chromatographic LC-MS run without derivatization. Our developed method can be used in research or clinical applications for disease-related detection of accumulated cholesterol intermediates. Disorders in the late part of cholesterol synthesis lead to severe malformation in human patients. The developed method enables a simple, sensitive, and fast quantification of sterols, without the need of extended knowledge of the LC-MS technique, and represents a new analytical tool in the rising field of cholesterolomics.


Assuntos
Colesterol/análise , Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Esteróis/análise , Colecalciferol/análogos & derivados , Colecalciferol/análise , Desmosterol/análise , Fluorbenzenos/química , Deleção de Genes , Células Hep G2 , Hepatócitos/metabolismo , Humanos , Lanosterol/análise , Fenóis/química , Reprodutibilidade dos Testes
9.
J Membr Biol ; 253(5): 445-457, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32949248

RESUMO

The role of membrane cholesterol in modulating G protein-coupled receptor (GPCR) structure and function has emerged as a powerful theme in contemporary biology. In this paper, we report the subtlety and stringency involved in the interaction of sterols with the serotonin1A receptor. For this, we utilized two immediate biosynthetic precursors of cholesterol, 7-dehydrocholesterol (7-DHC) and desmosterol, which differ with cholesterol merely in a double bond in their chemical structures in a position-dependent manner. We show that whereas 7-DHC could not support the ligand binding function of the serotonin1A receptor in live cells, desmosterol could partially support it. Importantly, depletion and enrichment of membrane cholesterol over basal level resulted in an increase and reduction of the basal receptor activity, respectively. These results demonstrate the relevance of optimal membrane cholesterol in maintaining the activity of the serotonin1A receptor, thereby elucidating the relevance of cellular cholesterol homeostasis.


Assuntos
Colesterol/química , Colesterol/metabolismo , Receptor 5-HT1A de Serotonina/química , Receptor 5-HT1A de Serotonina/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Linhagem Celular , Membrana Celular/metabolismo , Células Cultivadas , Desmosterol/metabolismo , Humanos , Ligantes , Lipídeos de Membrana/metabolismo , Redes e Vias Metabólicas , Ligação Proteica , Relação Estrutura-Atividade
10.
Biosci Rep ; 40(7)2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32579186

RESUMO

It is controversial whether atherosclerosis is linked to increased intestinal cholesterol absorption or synthesis in humans. The aim of the present study was to relate atherosclerosis to the measurements of plasma markers of cholesterol synthesis (desmosterol, lathosterol) and absorption (campesterol, sitosterol). In healthy male (n=344), non-obese, non-diabetics, belonging to the city of São Paulo branch of the Brazilian Longitudinal Study of Adult Health (ELSA-Brasil), we measured in plasma these non-cholesterol sterol markers, together with their anthropometric, dietary parameters, traditional atherosclerotic risk factors, and blood chemistry, coronary arterial calcium score (CAC), and ultrasonographically measured common carotid artery intima-media thickness (CCA-IMT). Cases with CAC>zero had the following parameters higher than cases with CAC = zero: age, waist circumference (WC), plasma total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C), and non-high density lipoprotein-cholesterol (non HDL-C). Plasma desmosterol and campesterol, duly corrected for TC, age, body mass index (BMI), waist circumference (WC), hypertension, smoking, and the homeostasis model assessment-insulin resistance (HOMA-IR) correlated with CAC, but not with CCA-IMT. The latter related to increased age, BMI, waist circumference (WC), and systolic blood pressure (SBP). Plasma HDL-C concentrations did not define CAC or CCA-IMT degrees, although in relation to the lower tertile of HDL-C in plasma the higher tertile of HDL-C had lower HOMA-IR and concentration of a cholesterol synthesis marker (desmosterol). Present work indicated that increased cholesterol synthesis and absorption represent primary causes of CAD, but not of the common carotid artery atherosclerosis.


Assuntos
Aterosclerose/diagnóstico , Cálcio/análise , Vasos Coronários/química , Adulto , Idoso , Aterosclerose/sangue , Biomarcadores/sangue , Biomarcadores/metabolismo , Índice de Massa Corporal , Brasil , Artéria Carótida Primitiva/diagnóstico por imagem , Espessura Intima-Media Carotídea , Colesterol/análogos & derivados , Colesterol/sangue , Colesterol/metabolismo , HDL-Colesterol/sangue , LDL-Colesterol/sangue , Vasos Coronários/diagnóstico por imagem , Estudos Transversais , Desmosterol/sangue , Desmosterol/metabolismo , Feminino , Humanos , Absorção Intestinal , Mucosa Intestinal/metabolismo , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Fitosteróis/sangue , Fitosteróis/metabolismo , Estudos Prospectivos , Sitosteroides/sangue , Sitosteroides/metabolismo , Tomografia Computadorizada por Raios X , Ultrassonografia
11.
J Intern Med ; 288(5): 560-569, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32415867

RESUMO

BACKGROUND: We have earlier reported that amiodarone, a potent and commonly used antiarrhythmic drug increases serum desmosterol, the last precursor of cholesterol, in 20 cardiac patients by an unknown mechanism. OBJECTIVE: Here, we extended our study to a large number of cardiac patients of heterogeneous diagnoses, evaluated the effects of combining amiodarone and statins (inhibitors of cholesterol synthesis at the rate-limiting step of hydroxy-methyl-glutaryl CoA reductase) on desmosterol levels and investigated the mechanism(s) by which amiodarone interferes with the metabolism of desmosterol using in vitro studies. METHODS AND RESULTS: We report in a clinical case-control setting of 236 cardiac patients (126 with and 110 without amiodarone treatment) that amiodarone medication is accompanied by a robust increase in serum desmosterol levels independently of gender, age, body mass index, cardiac and other diseases, and the use of statins. Lipid analyses in patient samples taken before and after initiation of amiodarone therapy showed a systematic increase of desmosterol upon drug administration, strongly arguing for a direct causal link between amiodarone and desmosterol accumulation. Mechanistically, we found that amiodarone resulted in desmosterol accumulation in cultured human cells and that the compound directly inhibited the 24-dehydrocholesterol reductase (DHCR24) enzyme activity. CONCLUSION: These novel findings demonstrate that amiodarone blocks the cholesterol synthesis pathway by inhibiting DHCR24, causing a robust accumulation of cellular desmosterol in cells and in the sera of amiodarone-treated patients. It is conceivable that the antiarrhythmic potential and side effects of amiodarone may in part result from inhibition of the cholesterol synthesis pathway.


Assuntos
Amiodarona/efeitos adversos , Antiarrítmicos/efeitos adversos , Arritmias Cardíacas/sangue , Arritmias Cardíacas/tratamento farmacológico , Colesterol/biossíntese , Desmosterol/sangue , Proteínas do Tecido Nervoso/antagonistas & inibidores , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Estudos de Casos e Controles , Células Cultivadas , Colesterol/sangue , Feminino , Humanos , Lipídeos/sangue , Masculino , Pessoa de Meia-Idade
12.
Biochim Biophys Acta Biomembr ; 1862(9): 183357, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32416195

RESUMO

Ram sperm are particularly sensitive to freeze-thawing mainly due to their lipid composition, limiting their use in artificial insemination programs. We evaluated the extent of cholesterol and desmosterol incorporation into ram sperm through incubation with increasing concentrations of methyl-ß-cyclodextrin (MßCD)-sterol complexes, and its effect on membrane biophysical properties, membrane lateral organization and cryopreservation outcome. Sterols were effectively incorporated into the sperm membrane at 10 and 25 mM MßCD-sterols, similarly increasing membrane lipid order at physiological temperature and during temperature decrease. Differential ordering effect of sterols in ternary-mixture model membranes revealed a reduced tendency of desmosterol of segregating into ordered domains. Live cell imaging of fluorescent cholesterol showed sterol incorporation and evidenced the presence of sperm sub-populations compatible with different sterol contents and a high concentration of sterol rich-ordered domains mainly at the acrosome plasma membrane. Lateral organization of the plasma membrane, assessed by identification of GM1-related rafts, was preserved after sterol incorporation except when high levels of sterols (25 mM MßCD-desmosterol) were incorporated. Ram sperm incubation with 10 mM MßCD-sterols prior to cryopreservation in a cholesterol-free extender improved sperm quality parameters after cooling and freezing. While treatment with 10 mM MßCD-cholesterol increased sperm motility, membrane integrity and tolerance to osmotic stress after thawing, incorporation of desmosterol increased the ability of ram sperm to overcome osmotic stress. Our research provides evidence on the effective incorporation and biophysical behavior of cholesterol and desmosterol in ram sperm membranes and on their consequences in improving functional parameters of sperm after temperature decrease and freezing.


Assuntos
Membrana Celular/metabolismo , Criopreservação , Desmosterol/farmacologia , Motilidade Espermática/efeitos dos fármacos , Espermatozoides/metabolismo , Animais , Masculino , Ovinos , beta-Ciclodextrinas/farmacologia
13.
ACS Chem Neurosci ; 11(10): 1413-1423, 2020 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-32286791

RESUMO

Amiodarone is prescribed for the treatment and prevention of irregular heartbeats. Although effective in clinical practice, the long-term use of amiodarone has many unwanted side effects, including cardiac, pulmonary, hepatic, and neurological toxicities. Our objective was to elucidate effects of amiodarone exposure on the cholesterol metabolism in cultured neuronal and non-neuronal cells and in individuals taking amiodarone. We observed that amiodarone increases distinct cholesterol precursors in different cell types in a dose-dependent manner. In liver and kidney cell lines, amiodarone causes increase in desmosterol levels, and in primary cortical neurons and astrocytes, amiodarone increases zymosterol, zymostenol, and 8-dehydrocholesterol (8-DHC). We conclude that amiodarone inhibits two enzymes in the pathway, emopamil binding protein (EBP) and dehydrocholesterol reductase 24 (DHCR24). Cortical neurons and astrocytes are more sensitive to amiodarone than liver and kidney cell lines. We confirmed the inhibition of EBP enzyme by analyzing the sterol intermediates in EBP-deficient Neuro2a cells versus amiodarone-treated control Neuro2a cells. To determine if the cell culture experiments have clinical relevance, we analyzed serum samples from amiodarone users. We found that in patient serum samples containing detectable amount of amiodarone there are elevated levels of the sterol precursors zymosterol, 8-DHC, and desmosterol. This study illustrates the need for close monitoring of blood biochemistry during prolonged amiodarone use to minimize the risk of side effects.


Assuntos
Amiodarona , Oxirredutases atuantes sobre Doadores de Grupo CH-CH , Amiodarona/farmacologia , Proteínas de Transporte , Colesterol , Desidrocolesteróis , Desmosterol , Humanos , Proteínas do Tecido Nervoso/metabolismo , Oxirredutases , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Verapamil/análogos & derivados
14.
Nutrients ; 11(7)2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31247945

RESUMO

Human studies have shown diurnal rhythms of cholesterol and bile acid synthesis, but a better understanding of the role of the circadian system in cholesterol homeostasis is needed for the development of targeted interventions to improve metabolic health. Therefore, we performed a systematic literature search on the diurnal rhythms of cholesterol synthesis and absorption markers and of bile acid synthesis markers. We also examined the diurnal rhythms of the cholesterol synthesis markers lathosterol and desmosterol, and of the cholesterol absorption markers cholestanol, campesterol, and sitosterol in serum samples from the Bispebjerg study. These samples were collected every three hours over a 24-hour period in healthy males (n = 24) who consumed low-fat meals. The systematic search identified sixteen papers that had examined the diurnal rhythms of the cholesterol synthesis markers lathosterol (n = 3), mevalonate (n = 9), squalene (n = 2), or the bile acid synthesis marker 7α-hydroxy-4-cholesten-3-one (C4) (n = 4). Results showed that lathosterol, mevalonate, and squalene had a diurnal rhythm with nocturnal peaks, while C4 had a diurnal rhythm with daytime peaks. Furthermore, cosinor analyses of the serum samples showed a significant diurnal rhythm for lathosterol (cosinor p < 0.001), but not for desmosterol, campesterol, sitosterol, and cholestanol (cosinor p > 0.05). In conclusion, cholesterol synthesis and bile acid synthesis have a diurnal rhythm, though no evidence for a diurnal rhythm of cholesterol absorption was found under highly standardised conditions. More work is needed to further explore the influence of external factors on the diurnal rhythms regulating cholesterol homeostasis.


Assuntos
Ácidos e Sais Biliares/biossíntese , Colesterol/biossíntese , Colesterol/sangue , Ritmo Circadiano , Absorção Intestinal , Adolescente , Adulto , Biomarcadores/sangue , Colestanol/sangue , Colesterol/análogos & derivados , Desmosterol/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fitosteróis/sangue , Sitosteroides/sangue , Fatores de Tempo , Adulto Jovem
15.
J Inherit Metab Dis ; 42(5): 934-943, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-30891795

RESUMO

Cholesterol serves as a building material for cellular membranes and plays an important role in cellular metabolism. The brain relies on its own cholesterol biosynthesis, which starts during embryonic development. Cholesterol is synthesized from two immediate precursors, desmosterol and 7-dehydrocholesterol (7-DHC). Mutations in the DHCR24 enzyme, which converts desmosterol into cholesterol, lead to desmosterolosis, an autosomal recessive developmental disorder. In this study, we assessed the brain content of desmosterol, 7-DHC, and cholesterol from development to adulthood, and analyzed the biochemical, molecular, and anatomical consequences of Dhcr24 mutations on the sterol profile in a mouse model of desmosterolosis and heterozygous Dhcr24+/- carriers. Our HPLC-MS/MS studies revealed that by P0 desmosterol almost entirely replaced cholesterol in the Dhcr24-KO brain. The greatly elevated desmosterol levels were also present in the Dhcr24-Het brains irrespective of maternal genotype, persisting into adulthood. Furthermore, Dhcr24-KO mice brains showed complex changes in expression of lipid and sterol transcripts, nuclear receptors, and synaptic plasticity transcripts. Cultured Dhcr24-KO neurons showed increased arborization, which was also present in the Dhcr24-KO mouse brains. Finally, we observed a shared pathophysiological mechanism between the mouse models of desmosterolosis and Smith-Lemli-Opitz syndrome (a genetic disorder of conversion of 7-DHC to cholesterol).


Assuntos
Anormalidades Múltiplas/metabolismo , Encéfalo/metabolismo , Colesterol/biossíntese , Erros Inatos do Metabolismo Lipídico/metabolismo , Anormalidades Múltiplas/genética , Animais , Membrana Celular/metabolismo , Desidrocolesteróis/metabolismo , Desmosterol/metabolismo , Feminino , Homeostase , Erros Inatos do Metabolismo Lipídico/genética , Masculino , Camundongos , Mutação , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/genética , Síndrome de Smith-Lemli-Opitz , Esteróis/metabolismo , Espectrometria de Massas em Tandem
16.
Sci Rep ; 9(1): 1550, 2019 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-30733456

RESUMO

Preeclampsia is one of the most serious complications during pregnancy, defined as development of hypertension during late pregnancy affecting other organ systems (proteinuria, thrombocytopenia, renal insufficiency, liver involvement, cerebral symptoms or pulmonary edema). Preeclampsia is known to be associated with significant dyslipidemia, but the cause or mechanism of this metabolic aberration is not clear. Quantitative analysis of cholesterol precursors and metabolites can reveal metabolic signatures of cholesterol, and provide insight into cholesterol biosynthetic and degradation pathways. We undertook this study to compare the metabolic signatures of cholesterol in serum and amniotic fluid collected from women who delivered in the late preterm period. Matching serum and amniotic fluid samples were collected from women who delivered in the late preterm period (34-0/7-36-6/7 weeks), had undergone amniocentesis within 3 days of delivery, had no evidence of rupture of membranes or intra-amniotic infection/inflammation, and who had not received antenatal corticosteroid prior to amniocentesis. Patients were classified into 3 groups according to the etiology of their preterm birth: Group 1, preeclampsia; Group 2, spontaneous preterm labor; Group 3, other maternal medical indications for iatrogenic preterm birth. Quantitative metabolite profiling of cholesterols was performed using gas chromatography-mass spectrometry. A total of 39 women were included in the analysis (n = 14 in Group 1, n = 16 in Group 2, n = 9 in Group 3). In maternal blood, patients in Group 1 had significantly higher ratios of cholesterol/desmosterol and cholesterol/7-dehydrocholesterol (which represent 24- and 7-reductase enzyme activity, respectively) than those in Group 3 (p < 0.05 for each), which suggests increased cholesterol biosynthesis. In contrast, patients in Group 1 had significantly decreased ratios of individual cholesterol esters/cholesterol and total cholesterol esters/cholesterol than those in Groups 3 (p < 0.01 for each), suggesting increased reverse cholesterol transport. No differences in cholesterol ratios were found in amniotic fluid among the 3 groups. In conclusion, the metabolic signatures of cholesterol suggest increased cholesterol biosynthesis and accumulation in the maternal blood (but not amniotic fluid) of women with preeclampsia.


Assuntos
Líquido Amniótico/metabolismo , Colesterol/sangue , Pré-Eclâmpsia/patologia , Adulto , Colesterol/análise , Desidrocolesteróis/análise , Desidrocolesteróis/sangue , Desmosterol/análise , Desmosterol/sangue , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Pré-Eclâmpsia/sangue , Gravidez , Nascimento Prematuro
17.
Nutrients ; 11(1)2019 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-30634478

RESUMO

Non-cholesterol sterols are validated biomarkers for intestinal cholesterol absorption and endogenous cholesterol synthesis. However, their use in metabolic disturbances has not been systematically explored. Therefore, we conducted a systematic review to provide an overview of non-cholesterol sterols as markers for cholesterol metabolism in different metabolic disorders. Potentially relevant studies were retrieved by a systematic search of three databases in July 2018 and ninety-four human studies were included. Cholesterol-standardized levels of campesterol, sitosterol and cholestanol were collected to reflect cholesterol absorption and those of lathosterol and desmosterol to reflect cholesterol synthesis. Their use as biomarkers was examined in the following metabolic disorders: overweight/obesity (n = 16), diabetes mellitus (n = 15), metabolic syndrome (n = 5), hyperlipidemia (n = 11), cardiovascular disease (n = 17), and diseases related to intestine (n = 16), liver (n = 22) or kidney (n = 2). In general, markers for cholesterol absorption and synthesis displayed reciprocal patterns, showing that cholesterol metabolism is tightly regulated by the interplay of intestinal absorption and endogenous synthesis. Distinctive patterns for cholesterol absorption or cholesterol synthesis could be identified, suggesting that metabolic disorders can be classified as 'cholesterol absorbers or cholesterol synthesizers'. Future studies should be performed to confirm or refute these findings and to examine whether this information can be used for targeted (dietary) interventions.


Assuntos
Biomarcadores/sangue , Colesterol/metabolismo , Doenças Metabólicas/sangue , Esteróis/sangue , Doenças Cardiovasculares/sangue , Colesterol/análogos & derivados , Colesterol/sangue , Desmosterol/sangue , Diabetes Mellitus/sangue , Humanos , Absorção Intestinal , Enteropatias/sangue , Nefropatias/sangue , Hepatopatias/sangue , Obesidade/sangue , Sobrepeso/sangue , Fitosteróis/sangue , Sitosteroides/sangue
18.
JCI Insight ; 3(24)2018 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-30568031

RESUMO

Drug-induced kidney injury, largely caused by proximal tubular intoxicants, limits development and clinical use of new and approved drugs. Assessing preclinical nephrotoxicity relies on animal models that are frequently insensitive; thus, potentially novel techniques - including human microphysiological systems, or "organs on chips" - are proposed to accelerate drug development and predict safety. Polymyxins are potent antibiotics against multidrug-resistant microorganisms; however, clinical use remains restricted because of high risk of nephrotoxicity and limited understanding of toxicological mechanisms. To mitigate risks, structural analogs of polymyxins (NAB739 and NAB741) are currently in clinical development. Using a microphysiological system to model human kidney proximal tubule, we exposed cells to polymyxin B (PMB) and observed significant increases of injury signals, including kidney injury molecule-1 KIM-1and a panel of injury-associated miRNAs (each P < 0.001). Surprisingly, transcriptional profiling identified cholesterol biosynthesis as the primary cellular pathway induced by PMB (P = 1.22 ×10-16), and effluent cholesterol concentrations were significantly increased after exposure (P < 0.01). Additionally, we observed no upregulation of the nuclear factor (erythroid derived-2)-like 2 pathway, despite this being a common pathway upregulated in response to proximal tubule toxicants. In contrast with PMB exposure, minimal changes in gene expression, injury biomarkers, and cholesterol concentrations were observed in response to NAB739 and NAB741. Our findings demonstrate the preclinical safety of NAB739 and NAB741 and reveal cholesterol biosynthesis as a potentially novel pathway for PMB-induced injury. To our knowledge, this is the first demonstration of a human-on-chip platform used for simultaneous safety testing of new chemical entities and defining unique toxicological pathway responses of an FDA-approved molecule.


Assuntos
Injúria Renal Aguda/induzido quimicamente , Rim/efeitos dos fármacos , Polimixinas/toxicidade , Animais , Antibacterianos/toxicidade , Biomarcadores , Desidrocolesteróis , Desmosterol , Modelos Animais de Doenças , Expressão Gênica , Heme Oxigenase-1 , Receptor Celular 1 do Vírus da Hepatite A , Humanos , Rim/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Lanosterol , Fator 2 Relacionado a NF-E2/metabolismo , Polimixina B/farmacologia , Polimixinas/farmacologia
19.
J Lipid Res ; 59(10): 1916-1926, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30087204

RESUMO

Regulating blood cholesterol (Chol) levels by pharmacotherapy has successfully improved cardiovascular health. There is growing interest in the role of Chol precursors in the treatment of diseases. One sterol precursor, desmosterol (Des), is a potential pharmacological target for inflammatory and neurodegenerative disorders. However, elevating levels of the precursor 7-dehydrocholesterol (7-DHC) by inhibiting the enzyme 7-dehydrocholesterol reductase is linked to teratogenic outcomes. Thus, altering the sterol profile may either increase risk toward an adverse outcome or confer therapeutic benefit depending on the metabolite affected by the pharmacophore. In order to characterize any unknown activity of drugs on Chol biosynthesis, a chemical library of Food and Drug Administration-approved drugs was screened for the potential to modulate 7-DHC or Des levels in a neural cell line. Over 20% of the collection was shown to impact Chol biosynthesis, including 75 compounds that alter 7-DHC levels and 49 that modulate Des levels. Evidence is provided that three tyrosine kinase inhibitors, imatinib, ponatinib, and masitinib, elevate Des levels as well as other substrates of 24-dehydrocholesterol reductase, the enzyme responsible for converting Des to Chol. Additionally, the mechanism of action for ponatinib and masitinib was explored, demonstrating that protein levels are decreased as a result of treatment with these drugs.


Assuntos
Desidrocolesteróis/metabolismo , Desmosterol/metabolismo , Medicamentos sob Prescrição , Benzamidas , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Regulação da Expressão Gênica/efeitos dos fármacos , Ensaios de Triagem em Larga Escala , Humanos , Imidazóis/farmacologia , Proteínas do Tecido Nervoso/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Piperidinas , Piridazinas/farmacologia , Piridinas , Tiazóis/farmacologia , Estados Unidos , United States Food and Drug Administration
20.
Proc Natl Acad Sci U S A ; 115(20): E4680-E4689, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29632203

RESUMO

Activation of liver X receptors (LXRs) with synthetic agonists promotes reverse cholesterol transport and protects against atherosclerosis in mouse models. Most synthetic LXR agonists also cause marked hypertriglyceridemia by inducing the expression of sterol regulatory element-binding protein (SREBP)1c and downstream genes that drive fatty acid biosynthesis. Recent studies demonstrated that desmosterol, an intermediate in the cholesterol biosynthetic pathway that suppresses SREBP processing by binding to SCAP, also binds and activates LXRs and is the most abundant LXR ligand in macrophage foam cells. Here we explore the potential of increasing endogenous desmosterol production or mimicking its activity as a means of inducing LXR activity while simultaneously suppressing SREBP1c-induced hypertriglyceridemia. Unexpectedly, while desmosterol strongly activated LXR target genes and suppressed SREBP pathways in mouse and human macrophages, it had almost no activity in mouse or human hepatocytes in vitro. We further demonstrate that sterol-based selective modulators of LXRs have biochemical and transcriptional properties predicted of desmosterol mimetics and selectively regulate LXR function in macrophages in vitro and in vivo. These studies thereby reveal cell-specific discrimination of endogenous and synthetic regulators of LXRs and SREBPs, providing a molecular basis for dissociation of LXR functions in macrophages from those in the liver that lead to hypertriglyceridemia.


Assuntos
Biomimética , Desmosterol/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hepatócitos/metabolismo , Receptores X do Fígado/metabolismo , Macrófagos/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Animais , Células Hep G2 , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Humanos , Receptores X do Fígado/genética , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Regiões Promotoras Genéticas , Proteína de Ligação a Elemento Regulador de Esterol 1/genética
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