RESUMO
Plasmacytoid dendritic cells (pDCs) are an immune subset devoted to the production of high amounts of type 1 interferons in response to viral infections. Whereas conventional dendritic cells (cDCs) originate mostly from a common dendritic cell progenitor (CDP), pDCs have been shown to develop from both CDPs and common lymphoid progenitors. Here, we found that pDCs developed predominantly from IL-7R+ lymphoid progenitor cells. Expression of SiglecH and Ly6D defined pDC lineage commitment along the lymphoid branch. Transcriptional characterization of SiglecH+Ly6D+ precursors indicated that pDC development requires high expression of the transcription factor IRF8, whereas pDC identity relies on TCF4. RNA sequencing of IL-7R+ lymphoid and CDP-derived pDCs mirrored the heterogeneity of mature pDCs observed in single-cell analysis. Both mature pDC subsets are able to secrete type 1 interferons, but only myeloid-derived pDCs share with cDCs their ability to process and present antigen.
Assuntos
Células Dendríticas/imunologia , Células-Tronco/imunologia , Animais , Linfócitos B/citologia , Linhagem da Célula , Células Cultivadas , Células Dendríticas/citologia , Feminino , Proteínas Ativadoras de GTPase/metabolismo , Fatores Reguladores de Interferon/metabolismo , Lectinas/metabolismo , Masculino , Camundongos , Receptores de Superfície Celular/metabolismo , Receptores de Interleucina-7/metabolismo , Transativadores/metabolismo , Transcrição GênicaRESUMO
Clinical evidence in oncology argues for the advantages of performing molecular analysis of blood biomarkers to provide information about systemic changes and tumor heterogeneity. Whereas the diagnostic value of cell-free circulating DNA (fcDNA) has successfully been demonstrated in several studies, DNA enclosed in extracellular vesicles (EV) has only recently been described, and its potential diagnostic value is unclear. We established a protocol for separation of EV and fc fractions and tested for presence of mutant BRAFV600E mediating resistance to Vemurafenib and cKITD816V mediating resistance to Imatinib in blood of patients with melanoma and mastocytosis. Our results show that EV contain significantly higher amounts of total DNA as compared to the fc fraction. However, about ten-fold higher copy numbers of the wild type and mutant BRAF and cKIT were detected in the fcDNA fraction supporting its diagnostic value and pointing to differences in fc and EV DNA content.
Assuntos
Ácidos Nucleicos Livres/genética , Vesículas Extracelulares/genética , Mastocitose Sistêmica/diagnóstico , Melanoma/diagnóstico , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-kit/genética , Biomarcadores Tumorais/genética , Ácidos Nucleicos Livres/sangue , Variações do Número de Cópias de DNA , Humanos , Mastocitose Sistêmica/genética , Melanoma/genéticaRESUMO
In cancer, extracellular vesicles (EV) contribute to tumor progression by regulating local and systemic effects. Being released into body fluids, EV may be used in nanomedicine as a valuable source for diagnostic biomarkers. In this work, infrared and Raman spectroscopy were used for comprehensive comparative analysis of cancer versus non-cancer EV and patient screening. Two different EV fractions enriched in exosomes and microvesicles were isolated by differential centrifugation from serum and plasma of cancer and non-cancer patients and from serum and plasma of a healthy donor. The EV fractions were then subjected to drop-coating deposition and drying on calcium fluoride substrates. Reduction of alpha-helix-rich proteins and enhancement of beta-sheet-rich proteins as a cancer-specific blood EV signature was determined, and subsequently this feature was applied for a pilot study aiming to detect prostate cancer in a test cohort of patients with high-grade prostate carcinoma and benign hypoplasia.