Muscle stem cells differentiate into haematopoietic lineages but retain myogenic potential

Muscle-derived stem cells (MDSCs) can differentiate into multiple lineages, including haematopoietic lineages. However, it is unknown whether MDSCs preserve their myogenic potential after differentiation into other lineages. To address this issue, we isolated from dystrophic muscle a population of MDSCs that express stem-cell markers and can differentiate into various lineages. After systemic delivery of three MDSC clones into lethally irradiated mice, we found that differentiation of the donor cells into various lineages of the haematopoietic system resulted in repopulation of the recipients' bone marrow. Donor-derived bone-marrow cells, isolated from these recipients by fluorescence-activated cell sorting (FACS), also repopulated the bone marrow of secondary, lethally irradiated, recipients and differentiated into myogenic cells both in vitro and in vivo in normal mdx mice. These findings demonstrate that MDSC clones retain their myogenic potential after haematopoietic differentiation.     

Cell kinetic status of haematopoietic stem cells

The haematopoietic stem cell (HSC) population supports a tremendous cellular production over the course of an animal's lifetime, e.g. adult humans produce their body weight in red cells, white cells and platelets every 7 years, while the mouse produces about 60% of its body weight in the course of a 2 year lifespan. Understanding how the HSC population carries this out is of interest and importance, and a first step in that understanding involves the characterization of HSC kinetics. Using previously published continuous labelling data (of Bradford et al. 1997 and Cheshier et al. 1999 ) from mouse HSC and a standard G₀ model for the cell cycle, the steady state parameters characterizing these HSC populations are derived. It is calculated that in the mouse the differentiation rate ranges between about 0.01 and 0.02, the rate of cell re-entry from G₀ back into the proliferative phase is between 0.02 and 0.05, the rate of apoptosis from the proliferative phase is between 0.07 and 0.23 (all units are days⁻¹), and the duration of the proliferative phase is between 1.4 and 4.3 days. These values are compared with previously obtained values derived from the modelling by Abkowitz and colleagues of long-term haematopoietic reconstitution in the cat ( Abkowitz et al. 1996 ) and the mouse ( Abkowitz et al. 2000 ). It is further calculated using the estimates derived in this paper and other data on mice that between the HSC and the circulating blood cells there are between 17 and 19.5 effective cell divisions giving a net amplification of between ~170 000 and ~720 000.     

Expansion of haematopoietic stem cells in vitro: a challenge to stem cell biologists

Expansion of haematopoietic stem cells from various sources has gained importance so as to provide a clinically potential graft, which shows ideal growth kinetics, resulting in reduction of the period of neutropenia and thrombocytopenia in any autologous or allogenic transplant setting. Expansion also facilitates transduction of genes for gene therapy. This review examines the various means employed to achieve the expansion of stem cells, and the criteria used to score the extent of expansion based on how stem cells are identified. It tries to analyse the ideal manner in which expansion should be carried out, with emphasis that expansion should not be at the expense of loss of stemness. It also attempts to judge the roles played by the stromal elements and cytokines, which are both part of the complex microenvironment, which in vivo has a strict regulation on haematopoiesis.     

Clinical strategies for expansion of haematopoietic stem cells

Haematopoietic stem cells (HSCs) give rise to all blood and immune cells and are used in clinical transplantation protocols to treat a wide variety of diseases. The ability to increase the number of HSCs either in vivo or in vitro would provide new treatment options, but the amplification of HSCs has been difficult to achieve. Recent insights into the mechanisms of HSC self-renewal now make the amplification of HSCs a plausible clinical goal. This article reviews the molecular mechanisms that control HSC numbers and discusses how these can be modulated to increase the number of HSCs. Clinical applications of HSC expansion are then discussed for their potential to address the current limitations of HSC transplantation.     

Non-circadian rhythm in proliferation of haematopoietic stem cells

The proportion of haematopoietic stem cells (CFU-s) engaged in DNA synthesis was determined by means of the [3H]-thymidine [( 3H]TdR) suicide technique during recovery of bone marrow from the damage caused by a sublethal total body irradiation. In contrast with previous reports the [3H]TdR suicide rate was not permanently increased. It was observed that CFU-s passed through S phase in synchronous waves, following a dose of irradiation of 1.5 Gy. After a dose of 2.6 Gy, there was only one initial wave of increased CFU-s sensitivity to the action of [3H]TdR. Following the depression occurring 26 hr after the irradiation with 2.6 Gy, the proportion of CFU-s killed by the [3H]TdR was permanently increased until 5-6 days after irradiation. Thereafter large differences in the [3H]TdR suicide data were observed among individual mice. Evidence was obtained that individual mice, which had been irradiated by a dose of 2.6 Gy 8-9 days before, had identical values of the CFU-s [3H]TdR suicide rate in the bone marrow from different bones of the lower extremities. The recurrence of the synchronous waves in CFU-s passage through the cell cycle was recorded when the CFU-s population regenerated to only about 10% of its normal value. These waves were obviously not related to a particular time of the day and, consequently, they did not represent the circadian rhythm. It is concluded that the synchronous waves in which CFU-s proliferation occurred reflected the action of the control mechanism on CFU-s proliferation. This mechanism should be endowed with an important systemic component besides locally operating factors.     

Irradiated stem cells and ageing of the haematopoietic system

In the work presented here, changes in haematopoiesis of mice (B6129SF2/J) were studied 1 year after their whole-body exposure to a dose of 7 Gy (72% of mice survived). The irradiated mice were compared with non-irradiated younger (4 months of age) and older (16 months of age) mice. There was a significant increase in the relative abundance of primitive stem cells with long-term capability of the haematopoiesis recovery lin⁻/Sca-1⁺/CD117⁺/CD34⁻ in the bone marrow of mice aged 16 months (irradiated and non-irradiated) compared with those aged 4 months. In terms of the ability to respond to further whole-body irradiation at a dose of 1 Gy, the presence of γH2A.X foci was studied in lin⁻ bone marrow cells. There was a considerable number of persisting foci in lin⁻ stem cells isolated from the bone marrow of the older irradiated mice. In the blood count from the peripheral blood of the older mice (both non-irradiated and irradiated at 7 Gy), there was a significant increase in granulocytes. In the group exposed to 7 Gy, the numbers of thrombocytes significantly increased, and on the contrary, the numbers of erythrocytes, the amount of haemoglobin, and haematocrit significantly decreased.     

Persistent response of Fanconi anemia haematopoietic stem and progenitor cells to oxidative stress

Oxidative stress is considered as an important pathogenic factor in many human diseases including Fanconi anemia (FA), an inherited bone marrow failure syndrome with extremely high risk of leukemic transformation. Members of the FA protein family are involved in DNA damage and other cellular stress responses. Loss of FA proteins renders cells hypersensitive to oxidative stress and cancer transformation. However, how FA cells respond to oxidative DNA damage remains unclear. By using an in vivo stress-response mouse strain expressing the Gadd45β-luciferase transgene, we show here that haematopoietic stem and progenitor cells (HSPCs) from mice deficient for the FA gene Fanca or Fancc persistently responded to oxidative stress. Mechanistically, we demonstrated that accumulation of unrepaired DNA damage, particularly in oxidative damage-sensitive genes, was responsible for the long-lasting response in FA HSPCs. Furthermore, genetic correction of Fanca deficiency almost completely abolished the persistent oxidative stress-induced G₂/M arrest and DNA damage response in vivo. Our study suggests that FA pathway is an integral part of a versatile cellular mechanism by which HSPCs respond to oxidative stress.     

Differentiation of monkey embryonic stem cells into neural lineages

Embryonic stem (ES) cells are self-renewing, pluripotent, and capable of differentiating into all of the cell types found in the adult body. Therefore, they have the potential to replace degenerated or damaged cells, including those in the central nervous system. For ES cell-based therapy to become a clinical reality, translational research involving nonhuman primates is essential. Here, we report monkey ES cell differentiation into embryoid bodies (EBs), neural progenitor cells (NPCs), and committed neural phenotypes. The ES cells were aggregated in hanging drops to form EBs. The EBs were then plated onto adhesive surfaces in a serum-free medium to form NPCs and expanded in serum-free medium containing fibroblast growth factor (FGF)-2 before neural differentiation was induced. Cells were characterized at each step by immunocytochemistry for the presence of specific markers. The majority of cells in complex/cystic EBs expressed antigens (alpha-fetal protein, cardiac troponin I, and vimentin) representative of all three embryonic germ layers. Greater than 70% of the expanded cell populations expressed antigenic markers (nestin and musashi1) for NPCs. After removal of FGF-2, approximately 70% of the NPCs differentiated into neuronal phenotypes expressing either microtubule-associated protein-2C (MAP2C) or neuronal nuclear antigen (NeuN), and approximately 28% differentiated into glial cell types expressing glial fibrillary acidic protein. Small populations of MAP2C/NeuN-positive cells also expressed tyrosine hydroxylase (approximately 4%) or choline acetyltransferase (approximately 13%). These results suggest that monkey ES cells spontaneously differentiate into cells of all three germ layers, can be induced and maintained as NPCs, and can be further differentiated into committed neural lineages, including putative neurons and glial cells.     

Uncertainty in the niches that maintain haematopoietic stem cells

Haematopoietic stem cell (HSC) niches are specialized microenvironments that contain stem cells and regulate their maintenance. Cells at the interface of bone and the bone marrow (the endosteum) contribute to the creation of HSC niches. It remains uncertain whether this interface itself is a niche, or whether endosteal cells secrete factors that diffuse to nearby niches. Vascular and/or perivascular cells may also create niches as many HSCs are observed around sinusoidal blood vessels, and perivascular cells secrete factors that regulate HSC maintenance. Do endosteal and perivascular cells create distinct niches, or do they contribute to a common niche? We discuss a range of niche models consistent with recent evidence.     

Diffusion chamber and spleen colony assay of murine haematopoietic stem cells

Mouse bone marrow cells have been cultured in diffusion chambers and their capacity to form spleen colonies in irradiated mice investigated after different culture periods. The number of spleen colony-forming units (CFU) in the chambers decreased during the first day of culture. The number then increased rapidly to a level significantly above the original chamber value on the third to fifth day of culture. By that time large numbers of granulocytes and macrophages had also appeared. Histological examination of spleen colonies showed that prior culturing did not alter the ratio between the different types of colonies. Cultured bone marrow cells which were transferred to new chambers retained granulopoietic capacity. This capacity increased between the first and second day of primary culturing. At this time hydroxyurea injections to chamber hosts revealed that the progenitor cells were proliferating. The results show that the granulopoietic progenitor cells of the chambers are stem cells, and that one progenitor cell type is identical with the CFU.     

Dynamic niches in the origination and differentiation of haematopoietic stem cells

Haematopoietic stem cells (HSCs) are multipotent, self-renewing progenitors that generate all mature blood cells. HSC function is tightly controlled to maintain haematopoietic homeostasis, and this regulation relies on specialized cells and factors that constitute the haematopoietic 'niche', or microenvironment. Recent discoveries, aided in part by technological advances in in vivo imaging, have engendered a new appreciation for the dynamic nature of the niche, identifying novel cellular and acellular niche components and uncovering fluctuations in the relative importance of these components over time. These new insights significantly improve our understanding of haematopoiesis and raise fundamental questions about what truly constitutes a stem cell niche.     

The ATM-BID pathway regulates quiescence and survival of haematopoietic stem cells

BID, a BH3-only BCL2 family member, functions in apoptosis as well as the DNA-damage response. Our previous data demonstrated that BID is an ATM effector acting to induce cell-cycle arrest and inhibition of apoptosis following DNA damage. Here we show that ATM-mediated BID phosphorylation plays an unexpected role in maintaining the quiescence of haematopoietic stem cells (HSCs). Loss of BID phosphorylation leads to escape from quiescence of HSCs, resulting in exhaustion of the HSC pool and a marked reduction of HSC repopulating potential in vivo. We also demonstrate that BID phosphorylation plays a role in protecting HSCs from irradiation, and that regulating both quiescence and survival of HSCs depends on BID's ability to regulate oxidative stress. Moreover, loss of BID phosphorylation, ATM knockout or exposing mice to irradiation leads to an increase in mitochondrial BID, which correlates with an increase in mitochondrial oxidative stress. These results show that the ATM-BID pathway serves as a critical checkpoint for coupling HSC homeostasis and the DNA-damage stress response to enable long-term regenerative capacity.     

Stability and Hopf bifurcation in an approachable haematopoietic stem cells model

We consider the haematopoietic stem cells model (HSC) with one delay introduced by Mackey [M.C. Mackey, Unified hypothesis for the origin of aplastic anemia and periodic hematopoiesis, blood 51 (1978) 5; M.C. Mackey, Mathematical models of haematopoietic cell replication and control, in: The Art of Mathematical Modelling: Case Studies in Ecology, Physiology and Biofluids, H.G. Othmer, F.R. Adler, M.A. Lewis, J.C. Dallon (Eds), Prentice-Hall, New York, 1997, p. 149] and Andersen and Mackey [L.K. Andersen, M.C. Mackey, Resonance in periodic chemotherapy: a case study of acute myelogenous leukemia, J. theor. Biol. 209 (2001) 113]. There are two possible stationary states in the model. One of them is trivial and the second E( *)(tau) depending on the delay is non-trivial . This paper investigates the stability of the non-trivial state and occurrence of the Hopf bifurcation depending on time delay. We prove the existence and uniqueness of a critical values tau(0) and tau of the delay such that E( *)(tau) is asymptotically stable for tau相似文献   

Enforced expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo

Hoxa5 is preferentially expressed in haematopoietic stem cells (HSCs) and multipotent progenitor cells (MPPs), and is more highly expressed in expanding HSCs. To date, little is known regarding the role of Hoxa5 in HSCs and downstream progenitor cells in vivo. In this study, we show that increased expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo. Hoxa5 differentially modifies the cell cycle of HSCs and lineage committed progenitor cells, depending on the cellular context. Hoxa5 drives HSCs, but not MPPs, through the cell cycle and arrests erythroid progenitor cells in G0 phase. Although the HSC pool shrinks after overexpression of Hoxa5, HSCs sustain the abilities of self-renewal and multipotency. In vivo, Hoxa5 has two effects on erythropoiesis: it causes a predominance of mature erythroid lineage cells and the partial apoptosis of erythroid progenitors. RNA-seq indicates that multiple biological processes, including erythrocyte homeostasis, cell metabolism, and apoptosis, are modified by Hoxa5. The results of this study indicate that Hoxa5 is a key regulator of the HSC cell cycle, and the inappropriate expression of Hoxa5 in lineage-committed progenitor cells leads to aberrant erythropoiesis.     

Enforced expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo

Hoxa5 is preferentially expressed in haematopoietic stem cells (HSCs) and multipotent progenitor cells (MPPs), and is more highly expressed in expanding HSCs. To date, little is known regarding the role of Hoxa5 in HSCs and downstream progenitor cells in vivo. In this study, we show that increased expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo. Hoxa5 differentially modifies the cell cycle of HSCs and lineage committed progenitor cells, depending on the cellular context. Hoxa5 drives HSCs, but not MPPs, through the cell cycle and arrests erythroid progenitor cells in G0 phase. Although the HSC pool shrinks after overexpression of Hoxa5, HSCs sustain the abilities of self-renewal and multipotency. In vivo, Hoxa5 has two effects on erythropoiesis: it causes a predominance of mature erythroid lineage cells and the partial apoptosis of erythroid progenitors. RNA-seq indicates that multiple biological processes, including erythrocyte homeostasis, cell metabolism, and apoptosis, are modified by Hoxa5. The results of this study indicate that Hoxa5 is a key regulator of the HSC cell cycle, and the inappropriate expression of Hoxa5 in lineage-committed progenitor cells leads to aberrant erythropoiesis.