ABSTRACT
The bromodomain and extra terminal (BET) protein family member BRD4 is a transcriptional regulator, critical for cell cycle progression and cellular viability. Here, we show that BRD4 plays an important role in embryonic stem cell (ESC) regulation. During differentiation of ESCs, BRD4 expression is upregulated and its gene promoter becomes demethylated. Disruption of BRD4 expression in ESCs did not induce spontaneous differentiation but severely diminished hematoendothelial potential. Although BRD4 regulates c-Myc expression, our data show that the role of BRD4 in hematopoietic commitment is not exclusively mediated by c-Myc. Our results indicate that BRD4 is epigenetically regulated during hematopoietic differentiation ESCs in the context of a still unknown signaling pathway.
Subject(s)
Embryonic Stem Cells/cytology , Hematopoietic Stem Cells/cytology , Nuclear Proteins/metabolism , Transcription Factors/metabolism , Cell Cycle Proteins , Cell Differentiation , Cell Line , DNA Methylation , Embryonic Stem Cells/metabolism , Epigenesis, Genetic , Fetal Blood/cytology , Hematopoiesis , Hematopoietic Stem Cells/metabolism , Humans , Infant, Newborn , Nuclear Proteins/genetics , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Signal Transduction , Transcription Factors/geneticsABSTRACT
Epigenetic deregulation is considered a common hallmark of cancer. Nevertheless, recent publications have demonstrated its association with a large array of human diseases. Here, we explore the DNA methylation dynamics in blood samples during hematopoietic cell transplant and how they are affected by pathophysiological events during transplant evolution. We analyzed global DNA methylation in a cohort of 47 patients with allogenic transplant up to 12 months post-transplant. Recipients stably maintained the donor's global methylation levels after transplant. Nonetheless, global methylation is affected by chimerism status. Methylation analysis of promoters revealed that methylation in more than 200 genes is altered 1 month post-transplant when compared with non-pathological methylation levels in the donor. This number decreased by 6 months post-transplant. Finally, we analyzed methylation in IFN-γ, FASL, IL-10, and PRF1 and found association with the severity of the acute graft-versus-host disease. Our results provide strong evidence that methylation changes in blood are linked to underlying physiological events and demonstrate that DNA methylation analysis is a viable strategy for the study of transplantation and for development of biomarkers.
Subject(s)
DNA Methylation/genetics , Hematopoietic Stem Cell Transplantation , Adolescent , Adult , Child , Child, Preschool , Fas Ligand Protein/blood , Female , Humans , Infant , Infant, Newborn , Interferon-gamma/blood , Interleukin-10/blood , Male , Middle Aged , Perforin , Pore Forming Cytotoxic Proteins/blood , Young AdultABSTRACT
To assess possible effects on subcellular organization after cryopreservation, we compared vitrified and slowly frozen oocytes in terms of their post-warm/thaw morphology, meiotic spindle configuration, and DNA integrity. DNA integrity of cryopreserved oocytes was not altered after the procedures, but vitrification was more effective than slow cooling, as shown by higher survival rate and spindle assessment despite a higher misalignment between meiotic spindle and polar body.
