RESUMO
Data-independent acquisition (DIA) allows comprehensive proteome coverage, while it also potentially works as a unified protocol to determine a multitude of proteins found in blood. Because of its high specificity, mass spectrometry may greatly reduce the interference observed in other assays to evaluate blood markers. Here, we combined DIA with volumetric absorptive microsampling (VAMS) and automated proteomics sample processing in a platform to assess clinical markers. As a proof of concept, we evaluated two hemoglobin-related biomarkers: the glycated hemoglobin (HbA1c) and hemoglobin (Hb) variants. HbA1c by DIA showed good correlation with the reference method, but method imprecision did not meet the quality requirement for this biomarker. We developed a strategy to identify Hb variants based on a customized database combined with a workflow for DIA data extraction and rigorous peptide evaluation. Data are available via ProteomeXchange with identifier PXD029918.
Assuntos
Coleta de Amostras Sanguíneas , Teste em Amostras de Sangue Seco , Biomarcadores , Coleta de Amostras Sanguíneas/métodos , Teste em Amostras de Sangue Seco/métodos , Hemoglobinas Glicadas , Espectrometria de Massas/métodosRESUMO
BACKGROUND: The most primitive leukemic precursor in acute myeloid leukemia (AML) is thought to be the leukemic stem cell (LSC), which retains the properties of self-renewal and high proliferative capacity and quiescence of the hematopoietic stem cell. LSC seems to be immunophenotypically distinct and more resistant to chemotherapy than the more committed blasts. Considering that the multidrug resistance (MDR) constitutive expression may be a barrier to therapy in AML, we have investigated whether various MDR transporters were differentially expressed at the protein level by different leukemic subsets. METHODS: The relative expression of the drug-efflux pumps P-gp, MRP, LRP, and BCRP was evaluated by mean fluorescence index (MFI) and the Kolmogorov-Smirnov analysis (D values) in five leukemic subpopulations: CD34+CD38-CD123+ (LSCs), CD34+CD38+CD123-, CD34+CD38+CD123+, CD34+CD38+CD123-, and CD34- mature cells in 26 bone marrow samples of CD34+ AML cases. RESULTS: : The comparison between the two more immature subsets (LSC versus CD34+CD38-CD123- cells) revealed a higher P-gp, MRP, and LRP expression in LSCs. The comparative analysis between LSCs and subsets of intermediate maturation (CD34+CD38+) demonstrated the higher BCRP expression in the LSCs. In addition, P-gp expression was also significantly higher in the LSC compared to CD34+CD38+CD123- subpopulation. Finally, the comparative analysis between LSC and the most mature subset (CD34-) revealed higher MRP and LRP and lower P-gp expression in the LSCs. CONCLUSIONS: Considering the cellular heterogeneity of AML, the higher MDR transporters expression at the most immature, self-renewable, and quiescent LSC population reinforces that MDR is one of the mechanisms responsible for treatment failure.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Proteínas de Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Biomarcadores Tumorais/metabolismo , Estudos de Casos e Controles , Diferenciação Celular , Resistencia a Medicamentos Antineoplásicos , Citometria de Fluxo/métodos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Células-Tronco Neoplásicas/classificação , Células-Tronco Neoplásicas/patologiaRESUMO
Patologias do sistema hemostático são causas freqüentes de atendimento médico de urgência. Este texto revisa aspectos relevantes no que diz respeito a diagnóstico e tratamento de hemofilias e coagulação intravascular disseminada (CIVD)
