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1.
Sci Rep ; 13(1): 1899, 2023 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-36732570

RESUMO

High-density lipoprotein (HDL) cholesterol efflux capacity (CEC), which is a conventional metric of HDL function, has been associated with coronary heart disease risk. However, the CEC assay requires cultured cells and takes several days to perform. We previously established a cell-free assay to evaluate cholesterol uptake capacity (CUC) as a novel measure of HDL functionality and demonstrated its utility in coronary risk stratification. To apply this concept clinically, we developed a rapid and sensitive assay system based on a chemiluminescent magnetic particle immunoassay. The system is fully automated, providing high reproducibility. Measurement of CUC in serum is completed within 20 min per sample without HDL isolation, a notably higher throughput than that of the conventional CEC assay. CUC decreased with myeloperoxidase-mediated oxidation of HDL or in the presence of N-ethylmaleimide, an inhibitor of lecithin: cholesterol acyltransferase (LCAT), whereas CUC was enhanced by the addition of recombinant LCAT. Furthermore, CUC correlated with CEC even after being normalized by ApoA1 concentration and was significantly associated with the requirement for revascularization due to the recurrence of coronary lesions. Therefore, our new assay system shows potential for the accurate measurement of CUC in serum and permits assessing cardiovascular health.


Assuntos
Doenças Cardiovasculares , Lipoproteínas HDL , Humanos , Doenças Cardiovasculares/diagnóstico , Reprodutibilidade dos Testes , HDL-Colesterol , Imunoensaio
2.
Clin Chim Acta ; 503: 136-144, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31972150

RESUMO

BACKGROUND: Cholesterol efflux from atherosclerotic lesion is a key function of high-density lipoprotein (HDL). Recently, we established a simple, high-throughput, cell-free assay to evaluate the capacity of HDL to accept additional cholesterol, which is herein referred to as "cholesterol uptake capacity (CUC)". OBJECTIVE: To clarify the cross-sectional relationship between CUC and coronary plaque properties. METHODS: We enrolled 135 patients to measure CUC and assess the morphological features of angiographic stenosis by optical coherence tomography (OCT). We estimated the extent of the lipid-rich plaque by multiplying the mean lipid arc by lipid length (lipid index). The extent of the OCT-detected macrophage accumulation in the target plaque was semi-quantitatively estimated using a grading system. RESULTS: Lipid-rich plaque lesions were identified in 125 patients (92.6%). CUC was inversely associated with the lipid index (R = -0.348, P < 0.0001). In addition, CUC was also inversely associated with macrophage score (R = -0.327, P < 0.0001). Conversely, neither circulating levels of HDL cholesterol nor apoA1 showed a similar relationship. CONCLUSIONS: We demonstrated that CUC was inversely related to lipid-rich plaque burden and the extent of macrophage accumulation, suggesting that CUC could be useful for cardiovascular risk stratification.


Assuntos
Colesterol/farmacocinética , Doença da Artéria Coronariana/patologia , Lipoproteínas HDL/fisiologia , Placa Aterosclerótica/patologia , Idoso , Apolipoproteína A-I , HDL-Colesterol , Doença da Artéria Coronariana/metabolismo , Feminino , Humanos , Lipídeos/análise , Macrófagos/metabolismo , Masculino , Pessoa de Meia-Idade , Placa Aterosclerótica/metabolismo , Tomografia de Coerência Óptica/métodos
3.
J Am Heart Assoc ; 8(9): e011975, 2019 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-30995875

RESUMO

Background We evaluated the importance of high-density lipoprotein (HDL) functionality for target-lesion revascularization in patients treated with coronary stents using a rapid cell-free assay system to evaluate the functional capacity of HDL to accept additional cholesterol (cholesterol-uptake capacity; CUC). Methods and Results From an optical coherence tomography (OCT) registry of patients treated with coronary stents, 207 patients were enrolled and their HDL was functionally evaluated by measuring the CUC. Follow-up OCT was performed (median duration, 24.5 months after stenting) to evaluate the presence of neoatherosclerosis. Clinical follow-up was performed to assess target-lesion revascularization for a median duration of 42.3 months after stent implantation. Neoatherosclerosis was identified in 37 patients (17.9%). Multivariate logistic regression analysis revealed that a decreased CUC was independently associated with neoatherosclerosis (odds ratio, 0.799; P<0.001). The CUC showed a significant inverse correlation with incidence of target-lesion revascularization (odds ratio, 0.887; P=0.003) and with lipid accumulation inside stents, suggesting that neoatherosclerosis contributes to the association between CUC and target-lesion revascularization. Conclusions Impaired HDL functionality, detected as decreased CUC, might lead to future stent failure by provoking atherogenic changes of the neointima within stents. Both quantitative and qualitative assessments of HDL might enable the improved prediction of clinical outcomes after stent implantation.


Assuntos
HDL-Colesterol/sangue , Doença da Artéria Coronariana/terapia , Vasos Coronários/metabolismo , Macrófagos/metabolismo , Intervenção Coronária Percutânea/instrumentação , Placa Aterosclerótica , Stents , Idoso , Idoso de 80 Anos ou mais , Doença da Artéria Coronariana/sangue , Doença da Artéria Coronariana/diagnóstico por imagem , Vasos Coronários/diagnóstico por imagem , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Intervenção Coronária Percutânea/efeitos adversos , Sistema de Registros , Fatores de Tempo , Tomografia de Coerência Óptica , Resultado do Tratamento
4.
J Appl Lab Med ; 2(2): 186-200, 2017 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32630971

RESUMO

BACKGROUND: Recent studies have shown that the cholesterol efflux capacity of HDL is a better predictor of cardiovascular disease (CVD) than HDL cholesterol. However, the standard procedures used for measuring cholesterol efflux capacity involve radioisotope-labeled cholesterol and cultured macrophages. Thus, a simpler method to measure HDL functionality is needed for clinical application. METHODS: We established a cell-free assay system to evaluate the capacity of HDL to accept additional cholesterol, which we named cholesterol "uptake capacity," using fluorescently labeled cholesterol and an anti-apolipoprotein A1 antibody. We quantified cholesterol uptake capacity of apolipoprotein B (apoB)-depleted serum samples from patients with coronary artery disease who had previously undergone revascularization. RESULTS: This assay system exhibited high reproducibility (CV <10%) and a short processing time (<6 h). The myeloperoxidase-mediated oxidation of apoB-depleted serum impaired cholesterol uptake capacity. Cholesterol uptake capacity correlated significantly with cholesterol efflux capacity (r2 = 0.47, n = 30). Furthermore, cholesterol uptake capacity correlated inversely with the requirement for revascularization because of recurrence of coronary lesions in patients with optimal control of LDL cholesterol (P < 0.01, n = 156). A multivariate analysis adjusted for traditional coronary risk factors showed that only cholesterol uptake capacity remained significant (odds ratio, 0.48; 95% CI, 0.29-0.80; P = 0.0048). CONCLUSIONS: Cholesterol uptake capacity assay evaluates the functionality of HDL in a sensitive and high-throughput manner without using radioisotope label and cells. This assay system could be used for the assessment of CVD risk in the clinical settings.

5.
Mol Cell Biol ; 29(17): 4729-41, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19546234

RESUMO

FLASH has been shown to be required for S phase progression and to interact with a nuclear protein, ataxia-telangiectasia locus (NPAT), a component of Cajal bodies in the nucleus and an activator of histone transcription. We investigated the role of human FLASH by using an inducible FLASH knockdown system in the presence or absence of various mutant forms of mouse FLASH. While carboxyl-terminal deletion mutants of FLASH, which do not interact with NPAT, can support S phase progression, its amino-terminal deletion mutants, which are unable to self associate, cannot support S phase progression, replication-dependent histone transcription, or the formation of Cajal bodies. Furthermore, FLASH was shown to be associated with arsenite resistance protein 2 (ARS2) through its central region, which is composed of only 13 amino acids. The expression of ARS2 and the interaction between FLASH and ARS2 are required for S phase progression. Taking these results together, FLASH functions in S phase progression through interaction with ARS2.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Ciclo Celular/fisiologia , Proteínas Nucleares/metabolismo , Fase S/fisiologia , Sequência de Aminoácidos , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ciclo Celular/genética , Linhagem Celular , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , Dados de Sequência Molecular , Proteínas Nucleares/genética , RNA/genética , RNA/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Transcrição Gênica
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