RESUMO
BACKGROUND INFORMATION: Megakaryocytes (MKs) follow a unique cell cycle duplication process, called endomitosis, resulting in polyploidisation of cells. It is hypothesised that polyploidy, as well as an increment in cytoplasm volume, allow more efficient platelets generation from MKs. Although polyploidy leads to an increase in the DNA amount, which impacts gene expression, little is known about ribosomal biogenesis in these polylobulated polyploid cells. RESULTS: The nucleolus acts as a hub for ribosomal biogenesis, which in turn governs the protein synthesis rate of the cells. We therefore estimated the size and activity of the nucleolus in polyploid cells during megakaryopoiesis in vitro. Polyploid megakaryocytic cell lines and in vitro cultured MKs, which were obtained from human cord blood-derived CD 34+ cells, revealed that miRNA 146b regulated the activity of nucleolar and coiled-body phosphoprotein 1, which plays an integral role in determining nucleolar size and activity. Additionally, miRNA-146b was up-regulated during endomitosis and was found to promote megakaryopoiesis. CONCLUSION: We propose that miRNA 146b regulates not only nucleolar size and activity, but also megakaryopoiesis. SIGNIFICANCE: This study highlights the importance of nucleolar activity and miRNA in the progression of megakaryopoiesis and thrombopoiesis.
Assuntos
Nucléolo Celular/metabolismo , Megacariócitos/citologia , Megacariócitos/metabolismo , MicroRNAs/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Sangue Fetal/citologia , Humanos , Células K562 , Proteínas Nucleares/metabolismo , Tamanho das Organelas , Fosfoproteínas/metabolismo , PoliploidiaRESUMO
During chronic hepatitis B (CHB), CD8+ T cells down-regulate CD28, the primary co-stimulation molecule for T-cell activation. Diverse functional attributes of CD8+CD28- T cells are suggested in various disease contexts. The present study aimed to characterize CD8+CD28- T cells in different phases of chronic Hepatitis B virus (HBV) infection (CHI)- Immune-tolerance (IT), Hepatitis B e-antigen-positive CHB (EP-CHB), Inactive carriers (IC) and Hepatitis B e-antigen-negative CHB (EN-CHB), to appraise their contribution in HBV-related disease pathophysiology. Flow cytometry analysis of T cells in peripheral blood of study subjects revealed enhanced CD8+CD28- T-cell accumulation in EP-/EN-CHB, compared with IT/IC and they expanded equivalently in HBV-specific and non-specific CD8+ T-cell compartments. Profound increase in CD8+CD28- T cells expressing perforin/granzyme-B/CD57/IFN-γ/TNF-α and markers of terminal differentiation were observed exclusively in EP-/EN-CHB. Further, activation with anti-NKG2D resulted in heightened IFN-γ/TNF-α production selectively from CD8+CD28- T cells, suggesting NKG2D-mediated alternative co-stimulation. CD8+CD28- T cells sorted from CHB patients induced enhanced apoptosis of peripheral blood mononuclear cells (PBMC), including CD4+ T cells. However, NKG2D-ligand (major histocompatibility complex class I chain-related molecule A/B (MICA/B)) was preferentially expressed by HBV-specific CD4+ T cells of CHB patients, making these cells a potential target to NKG2D-dependent CD8+CD28- T-cell killing. Both CD28+ and CD28- T cells in CHB expressed CXCR3 at similar levels and thus capable of homing to the liver. A positive correlation was seen between CD8+CD28- T-cell frequency and serum-alanine transaminase (ALT) levels and CHB-derived CD8+CD28- T cells caused pronounced cell death in HBV-transfected Huh7 cells. Immunofluorescence staining identified greater intrahepatic incidence of CD8+CD28- T cells but decline in CD4+ T cells in CHB than IC. Collectively, CD8+CD28- T cells demonstrated differential distribution and phenotypic/functional skewing in different CHI phases and contribute to disease progression by Perforin-Granzyme- or IFN-γ-TNF-α-mediated cytotoxicity while restraining antiviral immunity through NKG2D-dependent HBV-specific CD4+ T-cell depletion.
Assuntos
Antígenos CD28/imunologia , Linfócitos T CD8-Positivos/imunologia , Hepatite B Crônica/imunologia , Adolescente , Adulto , Carcinoma Hepatocelular/imunologia , Linhagem Celular Tumoral , Criança , Técnicas de Cocultura , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Imunofluorescência , Hepatite B Crônica/etiologia , Humanos , Neoplasias Hepáticas/imunologia , Masculino , Pessoa de Meia-Idade , Subpopulações de Linfócitos T , Adulto JovemRESUMO
In recent years, an explosion of interest for genome methylation in hematopoietic stem cells (HSCs) differentiation and transplantation has been observed. The differentiation and timely proliferation of HSCs require a stringent regulation of gene expression; this can't be maintained without the regulation of genome methylation. Evidences of changes in genome methylation patterns in healthy and cancer cells have opened a new paradigm for diagnostic and therapeutic strategies. Genome methylation or DNA methylation is achieved by DNA methyltransferases (DNMTs) through the transfer of methyl group leading to the regulation of gene's expression. Till now five DNMTs have been found in mammals, known as DNMT1, DNMT2, DNMT3A, DNMT3B and DNMT3L. In this review, we have carefully evaluated the spacio-temporal landscape expression and role of DNMTs during hematopoiesis, hematopoietic malignancy and HSCs transplantation. Focusing on the connection of DNA methylation with gene expression in association with cellular signalling, histone modifications, it is proposed for further improvement of strategies to use DNA methylation as a marker in clinical diagnosis as well as in therapeutic interventions by applying DNMTs inhibitors.
Assuntos
Metilação de DNA , Neoplasias Hematológicas , Animais , Metilação de DNA/genética , Genoma , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/terapia , Processamento de Proteína Pós-TraducionalRESUMO
We report the influence of Fe3 O4 nanoparticles (NPs) on porphyrins in the development of photosensitizers (PSs) for efficient photodynamic therapy (PDT) and possible post-PDT responses for inflicting cancer cell death. Except for Au, most metal-based nanomaterials are unsuitable for clinical applications. The US Food and Drug Administration and other agencies have approved Feraheme and a few other iron oxide NPs for clinical use, paving the way for novel biocompatible immunoprotective superparamagnetic iron oxide nanohybrids to be developed as nanotherapeutics. A water-soluble nanohybrid, referred to here as E-NP, comprising superparamagnetic Fe3 O4 NPs functionalised with tripyridyl porphyrin PS was introduced through a rigid 4-carboxyphenyl linker. As a PDT agent, the efficacy of E-NP toward the AGS cancer cell line showed enhanced photosensitising ability as determined through inâ vitro photobiological assays. The cellular uptake of E-NPs by AGS cells led to apoptosis by upregulating ROS through cell-cycle arrest and loss of mitochondrial membrane potential. The subcellular localisation of the PSs in mitochondria stimulated apoptosis through upregulation of p21, a proliferation inhibitor capable of preventing tumour development. Under both PDT and non-PDT conditions, this nanohybrid can act as an anti-inflammatory agent by decreasing the production of NO and superoxide ions in murine macrophages, thus minimising collateral damage to healthy cells.