ABSTRACT
The role of the sympathetic nervous system (SNS) in hematopoietic stem and progenitor cell (HSPC) mobilization has been largely investigated. However, there is a critical need for the identification of the underlying contributing factors to improve HSPC yield for transplantation. It has been demonstrated that miR-886-3p targets stromal-derived factor-1 (SDF-1), the central mediator of mobilization, and therefore may play a part in this process. Besides, miR-886-3p expression can be epigenetically regulated through DNA methylation modifications inits gene promoter. Here, to assess the contribution of miR-886-3p and other epigenetic factors in HSPC mobilization, human bone marrow-derived mesenchymal stem cells (MSCs) were treated with the ?-adrenergic agonist of isoprenaline. The expression of miR-886-3p and SDF-1and the gene promoter methylation status of this miRNA were then respectively evaluated through the appropriate PCR techniques. As expected, despite a transient initial increase in SDF-1mRNA level, its expression reduced, and miR-886-3plevel remarkably increased 48 h following treatment. The gene promoter methylation pattern of miR-886-3p also changed from a full methylated state to a partially methylated one. Together, our findings suggest that miR-886-3p can be epigenetically regulated and through suppressing the expression of SDF-1 play an active role in the SNS-mediated HSPC mobilization.
ABSTRACT
Background The chronic injury of the hematopoietic system caused by ionizing radiation (IR) is often ignored. The essential cause of this injury is the damage of hematopoietic stem and progenitor cells (HSPCs). Objective To explore the long-term effects of IR at different radiation doses and at different radiation fractions of the same radiation dose on HSPCs in the bone marrow of mice, and to provide a scientific basis for reducing the chronic damage to the hematopoietic system caused by IR. Methods A total of 16 male C57BL/6 mice aged 8-10 weeks were randomly divided into four groups that received different doses or fractions of total body X-ray irradiation, including 1.5 Gy×4 irradiation group (n=5), 3 Gy irradiation group (n=4), 6 Gy irradiation group (n=4), and non-irradiation group (n=3). Two months after irradiation, bone marrow cells from each mouse were collected and counted. The clone forming ability of bone marrow cells was analyzed by cobblestone area-forming cell (CAFC) assay. The proportion of HSPCs was measured by flow cytometry. The cell cycle of HSPCs was assessed by antigen identified by monoclonal antibody Ki 67 (Ki-67) and 7-amino-actinomycin D (7-AAD) double staining. The reactive oxygen species (ROS) levels of HSPCs were estimated with a 2,7-dichlorodihydrofluorescein diacetate (DCFDA) probe. The cellular senescence of HSPCs was evaluated with a 5-dodecanoylaminofluorescein di-β-D-galactopyranoside (C12FDG) probe. The expression of senescence related genes such as P16, P19, P21, and P27 was measured by real-time fluorescence quantitative PCR. Results There was no significant change in the numbers of bone marrow cells 2 months after different doses and fractions of radiation (P>0.05). The clone forming ability of bone marrow cells was significantly decreased after 3 Gy and 6 Gy irradiation when compared to non-irradiated mice (P<0.01). HSPCs responded inconsistently to different doses and fractions of irradiation. Overall, there was no significant change in long-term hematopoietic stem cells (LT-HSCs) proportion after irradiation (P>0.05), the proportions of hematopoietic progenitor cells (HPCs), hematopoietic stem cells (HSCs), short-term hematopoietic stem cells (ST-HSCs), and multipotent progenitors 2 (MPP2) increased after irradiation (P<0.05), and the proportions of LSK, MPP1, MPP3, and MPP4 cells decreased after irradiation (P<0.05); except for HPCs and MPP2, the proportion of HSPCs in G0 phase was decreased (P<0.05). The ROS production in HSPCs was increased significantly after 6 Gy irradiation (P<0.05), while the ROS levels after 3 Gy and 1.5 Gy×4 irradiation were similar to that of the non-radiation group (P>0.05). The cellular senescent proportion of HPCs, LSK, and HSCs increased after irradiation (P<0.05). The expression levels of senescence related genes P16, P19, and P21 in HSCs were significantly increased (P<0.05). Conclusion The responses of HSPCs in bone marrow to IR vary depending on doses and fractions of irradiation. Increased ROS production and cellular senescence may be involved in the damage process of HSPCs under radiation settings.
ABSTRACT
AIM:To observe the influences of artesunate(AS)on murine bone marrow hematopoietic stem and progenitor cells.METHODS:Murine bone marrow hematopoietic stem and progenitor cells were cultured in vitro with different concentration of AS.The numbers of erythrocyte colony-forming unit(CFU-E)or burst-forming unit(BFU-E)in liquid or semisolid culture were counted through light microscope.Apoptosis of cultured cells and the changes of mitochondrial membrane potential were detected by flow cytometry(FCM)analysis,meanwhile,DNA ladder gel electrophoresis were conducted.RESULTS:AS could significantly inhibit the formation of CFU-E and BFU-E(P