Supression of DNMTs Accelerates the In Vitro Erythropoietic Differentiation of Human Cd34+ Progenitor Cells

다국어 초록 (Multilingual Abstract)

Epigenetic modification dependent DNA methyltransferases (DNMTs) play an important role in tissue- and stage-specific gene regulation and normal mammalian development. In this study, we show that DNMTs are expressed at different levels during hematopoietic stem cell (HSC) differentiation to proerythrocytes. DNMT1, DNMT3A, and DNMT3B were highly expressed at day 7 after differentiation. We used specific siRNA as a tool to probe the relationship between the expression of DNMTs and erythropoietic differentiation. When introduced siRNA of DMNT1 and DMNT3b in human CD34+ cells, these more differentiated into erythrocytes. This was confirmed by glycophorin A (GPA) positive cell analysis and globin gene expression. GPA+ cells increased up to 20～30%, and γ- and ε-globin genes increased in siRNA transfected cells. Therefore, our data suggest that suppression of DNA methylation can affect positively differentiation of HSC and may contribute to expression of erythrocyte lineage genes including GPA and globins.

목차 (Table of Contents)

• ABSTRACT
• INTRODUCTION
• Isolation and Culture Conditions of Human CD34+ Cells
• Cell line and Erthropoiesis Induction
• Construction of siRNA Vectors for DNMT1, DNMT3b and Transfection in K562 and Human CD34+ Cells
• Fluorescence Activated Cell Sorter(FACS) Analysis of Antigen Expression
• Reverse-Transvcription (RT) PCR and Quantitative Real-Time PCR
• Western Blotting
• Statistical analysis
• RESULTS
• Differentiation of Human CD34+ Cells Originationg from Human UCB into Proerythrocytes
• Expression of DNMTs during HSC Differentiation
• Increment of Erythropoiesis in K562 and Human CD34+ Cells that Suppressed DNMTs
• DISCUSSION
• ACKNOWLEDGMENT
• REFERENCES