Stem Cells, Vol. 16, No. 6, 361-374,
November 1998
© 1998 AlphaMed Press
The Role of Blood Stem Cells in Hematopoietic Cell Renewal
Theodor M. Fliedner
Department of Clinical Physiology, Occupational and Social Medicine, University of Ulm, Ulm, Germany; present affiliation: Medicine Research Group, Medical Center of the University of Ulm, Ulm, Germany.
Key Words. Blood stem cells • Hematopoiesis • Cell renewal • Stem cell migration streams • Hematopoietic recovery • Progenitor cells
Dr. Theodor M. Fliedner, Radiation Medicine Research Group, Medical Center of the University of Ulm, Helmholtzstrasse 20, D-89081 Ulm, Germany.
 |
Abstract
|
|---|
It has been the purpose of this keynote address to review available evidence for the notion that the stem and progenitor cells circulating in the peripheral blood play a decisive role in the homeostasis of blood cell formation distributed throughout dozens of bone marrow units in the skeleton. Furthermore, if this notion is correct, one could speculate that the quantity and quality of stem and progenitor cells in the blood should reflect the functional state of the hematopoietic stem cell system throughout the skeletal bone marrow and provide a new tool for the evaluation of alteration in blood cell production. On this basis, the following questions are considered: A) What do we know about the quality and quantity of blood stem cells in steady-state conditions? B) In what way do blood stem cells respond to perturbations of the "steady-state" of blood cell formation? C) Which role do blood stem cells play during hemopoietic development assuming that the establishment of bone marrow hemopoiesis requires the "seeding" of blood stem cells into an appropriate cellular environment? D) What is the role of blood stem cells in hemopoietic regeneration after partial body irradiation with a small volume of marrow (and hence stem cells) protected? and E) What are the mechanisms and/or kinetics of hemopoietic recovery if stem cells introduced into the circulation were collected from exogenous (autologous or allogeneic) sources? In this review presentation, experimental work of our group and of other members of the scientific community is summarized. It becomes obvious that blood stem and progenitor cells play a key role in hematopoietic homeostasis. Furthermore, their physiology and pathophysiology deserve rigorous experimental studies in order to develop a novel tool in the diagnosis and prognosis of neoplastic and non-neoplastic disorders of blood cell formation.
|
Introduction: Scope and Purpose
|
|---|
It is the purpose of this presentation to examine and discuss two statements with respect to their experimental evidence and to suggest that the assessment of the quantity and quality of blood stem cells may well be useful as indicators of health and health impairments in the mammalian organism and, in particular, its blood-cell-forming tissues.
- The hematopoietic stem cell pool is distributed through the sites of hematopoietic cell renewal in a large number of bone marrow units distributed throughout the skeleton. That the hematopoietically active bone marrow sites act and respond as one organ system is due to the stem cell migration streams which connect via the blood stream all sites and assure a local stem cell concentration sufficient to maintain a balance between cell production and cell removal.
- The quality and quantity of stem cells in the peripheral blood should reflect the quality and quantity of the extravascular stem cell pools and the balance between the sites of hematopoietic cell renewal. Hence, a systematic investigation of blood stem cell changes in health and disease may well contribute to the arsenal of methods guiding the work of clinical hematology.
Alexander Maximow, while working in St. Petersburg as a military doctor in 1909, was the first to suggest that there was a hematopoietic stem cell with the morphological appearance of a "lymphocyte" capable of migrating through the blood to microecological niches that would allow them to proliferate and differentiate along lineage specific pathways [1]. Since then, many hematologists have contributed to the debate about the origin and mechanisms of homeostasis in hematopoietic cell renewal, and many decades have passed during which the "monophyletic" versus the "polyphyletic" origin of blood cell formation was discussed [2]. Today, we have to acknowledge the fact that Maximow, in principle, was right to suggest the origin of hematopoiesis in one stem cell capable of migrating from one site of hematopoiesis to the other via the blood stream and settling in tissue sites in which the microenvironment is conducive to differentiation and proliferation of blood progenitor and precursor cells.
The experimental evidence for the significance of blood stem cells in research and clinical practice can be derived from the developmental dynamics of scientific publications that are devoted in one way or the other to "blood stem cells." The International Database DIMDI for 1996 lists 314 references under the key word "blood stem cells." A search for the use of this term for 1970 showed only one reference (Fig. 1
). The first scientists to use or imply this term in experimental hematology were Goodman and Hodgson in 1962 [3].
View larger version (16K):
[in this window]
[in a new window]
|
Figure 1. Numbers of scientific publications with the key word "blood stem cells" in the International DIMDI Database.
|
|
Thus, it may be useful to review the experimental evidence of the physiology and pathophysiology of blood stem cells and to ask—even without being able to give final answers—what role they might play in the study and evaluation of hemopoietic cell renewal in health and disease.
In this presentation, stem cell migration streams will be examined using five key examples:
- steady-state situation in the adult;
- "physiological" perturbations of the steady state;
- hemopoietic development during embryogenesis;
- hematopoietic reconstitution after partial-body irradiation or chemotherapy, and
- hematopoietic recovery introducing stem cells into blood from exogenous sources.
It will become apparent that the stem and progenitor cells migrating in the blood stream may have a much higher significance for hematopoietic cell renewal than was assumed until recently. They do not appear to be "waste products" of bone marrow cell production [4] and require extensive research to further elucidate their physiological role in hematopoietic cell renewal and their diagnostic and therapeutic potentials.
|
Stem Cell Migration Streams in the Adult
|
|---|
The bone marrow in the adult organism is distributed throughout the skeleton which contains, according to the textbooks of anatomy, up to 206 bones, many of which house active blood-cell-forming tissue, and most of them can be activated to blood cell production under pathophysiological situations (leukemia, myelofibrosis, etc.). The amount of active bone marrow is known to amount to about 2,600 g, with about 126 x 1010 marrow cells and a turnover of 18.8 x 107/kg/h [5].
The question that comes up repeatedly is, "How is it possible that such a hematopoietic tissue which is ‘disseminated’ throughout so many skeletal bone cavities can act as one organ system?" If the clinician examines bone marrow aspirates taken from sternum, iliac crests, or other sites actively engaged in blood cell production, he can be certain to find a more or less identical cellular composition. It is the message of this presentation to suggest that it is due to stem cell migration via the peripheral blood that all bone marrow sites actively participating in blood cell production have and maintain a sufficient local concentration of hematopoietic stem cells as a prerequisite for humeral and nerval regulatory actions.
Let us review briefly what we know about stem and progenitor cells in the blood under steady-state conditions. The peripheral blood contains hematopoietic stem and progenitor cells that have been and are measurable as CD34+ cells, as colony-forming units (based on the pioneering work of Metcalf and his group [6]) or, as in the mouse, as CFU-S (colony-forming units in the mouse spleen) [7]. In the steady-state situation, it can now be assumed that they are in an equilibrium with the stem and progenitor pools in the bone marrow. As will be shown later, the stem and progenitor cells in the blood are not just random samples of the bone marrow stem and progenitor cells but subsets with surface properties that enable them to circulate and to seed in environments suitable for their replication and/or differentiation.
In the steady state, we can assume that the relationship is correctly described as
meaning that the rate of stem and progenitor cells entering the blood stream from the bone marrow sites is equal to the rate of their leaving the blood stream (Fig. 2).
What is the concentration of stem and progenitor cells in the circulating blood? Table 1 summarizes current knowledge. It is obvious from this table that several mammalian species, including man, show a definite presence of stem and progenitor cells in the peripheral blood, the concentration depending on the method of assessment that was used. However, the concentrations reported are in the range of several hundred to a few thousand/ml blood.
Studies have been performed to measure the concentration of stem and progenitor cells during several days and weeks in order to examine variations and oscillations. Micklem [8] noted in mice a twofold diurnal variation in circulating numbers of CFU-S in mice. In the five dogs examined during more than 11 weeks, Nothdurft et al. [9] could show that the concentration of colony-forming unit-culture (CFU-C) fluctuated around 181-341 CFU-C/ml. It was of interest to note that each dog appeared to have its personal CFU-C concentration similar to the common clinical knowledge that the "normal" level of blood cells can well be an "individual" characteristic. In human beings, Kreutzmann and Fliedner [10] could show in three normal volunteers, studied three times a week for 70 days, that the numbers of CFU-C were 255 ± 108, 125 ± 82, and 222 ± 87/ml, respectively, and that there was evidence for significant oscillation with periods of 23, 19, and 25 days (which is about twice the transit time from the granulocytic progenitor level to granulocyte release into the peripheral blood). Another 35-day study of our group revealed a mean concentration of CFU-C and of BFU-E of between 29 and 209/ml and 200 and 624/ml respectively [11] (Fig. 3).
The residence time of stem and progenitor cells in the blood stream is not very well known. However, some information is available for the mouse, using the CFU-S assay. Half-times between six min and 6-10 h were calculated [12, 13]. In the dog, Raghavachar et al. studied the disappearance rate of granulocytic progenitor cells (CFU-C) of normal recipient dogs and found a blood transit time of 11.8 to 13.0 min [14]. Nothdurft et al. used the CFU-C decline after dextran sulfate mobilization to calculate the blood transit time to be 1.4 ± 0.5 h and the T 1/2 to be 1.0 ± 0.4 h [15].
Thus, being well aware of all inherent methodological limitations, we conclude that the blood transit time of circulating progenitor cells is about one to two h, which is short in comparison to other "leukocytes."
The data obtained so far are not in contradiction to the following notions:
- Stem and progenitor cells are normal constituents of the "leukocyte population" in the peripheral blood of man as well as experimental animals studied.
- Their concentration, as measured by a considerable spectrum of methods, is characteristic of the method used but also of the individual organism.
- The tentative evidence of regular oscillations found for specific subpopulations is suggestive for a feedback regulation of their concentration.
- Their residence time in blood appears to be short, even shorter than that of granulocytes. Their fate after leaving the blood remains to be determined in detail; possibilities are intravascular apoptosis, seeding to appropriate environments (medullary and extramedullary), and changing "recognition" criteria.
|
Stem Cell Migration Streams After Perturbations
|
|---|
Let us now examine in what way the blood stem cell pool reacts to specific perturbations. In the next scheme (Case 2) (Fig. 4), a continuous-flow centrifugation (CFC) is depicted as it was performed in dogs [16] and in man [17]. From these experiments, two examples appear to be of characteristic importance for the topic of our presentation. A CFC was performed in a dog for 12.5 h (Fig. 5). During this time, the CFU-C concentration in the blood decreased to 30% of normal. At the same time, it was evident that 60 times the number of CFU-C normally present in the blood stream could be collected from the blood. This was explained by the assumption that these cells were drained from extravascular sites. It can also be seen that after the end of CFC, the blood concentration of CFU-C rose above normal levels by day 5 and returned to normal levels only after more than 20 days. This type of experiment was repeated three times with, in essence, the same results. These results suggested to us that the concentration of blood stem and progenitor cells is a feedback-controlled number and that there is in extravascular sites a "reserve pool" of progenitor cells prepared to be released into the circulation. This notion was further substantiated by studying the physical properties of the progenitor cells released into the blood after dextran sulfate mobilization and/or after CFC mobilization.
View larger version (21K):
[in this window]
[in a new window]
|
Figure 4. Case 2: Perturbation of steady state in stem cell migration streams by measures such as continuous-flow centrifugation.
|
|
View larger version (27K):
[in this window]
[in a new window]
|
Figure 5. 12.5 hours of leukocytapheresis in a dog. The CFU-C show a decrease toward the end of leukapheresis, an overshoot between day 1 and day 10, and a return to normal levels by day 23.
|
|
In our group, relevant canine studies were performed by Gerhartz et al. [18]. In these studies, the physical characteristics of circulating CFU-C were compared to those in the bone marrow. It was found that a four-to-eight-h CFC after dextran sulfate mobilization resulted in a selective release of CFU-C from the marrow into the blood with a velocity sedimentation profile of 4.65 ± 0.64 mm/h (range 3.6-5.5 mm/h), which is the same as that of unperturbed CFU-C blood population. However, when the CFC was prolonged to eight h or more, there were already more than eight h of evidence for a release of CFU-C with a sedimentation profile resembling more closely that of bone marrow CFU-C. This is similar to the sedimentation profile of CFU-C in the bone marrow of the same dogs, which was 5.54 ± 54 mm/h (range 3.8-6.2 mm/h). These studies were taken to indicate that the bone marrow releases a specific subpopulation of progenitor cells which is considerably smaller than the average of bone marrow progenitor cells. It was shown by Raghavachar [19] in our group that the bone marrow progenitor cells characterized by the slow sedimentation rate of <5.1 mm/h result in a more pronounced hematopoietic regeneration than equal numbers of bone marrow progenitor cells characterized by a faster sedimentation rate (>7.1 mm/h).
In human beings, it was shown by our group in initial studies in 1980 that one CFC results (without mobilization) in the collection of some 8.7 ± 4.3 x 105 CFU-C (mean of 35 leukaphereses), which is 20 times the number of CFU-C in the circulation. Repeated CFC resulted in a collection of 1.5 times the number of CFU-C in the circulation [17, 20]. More recent studies using CD34+ cells as an indicator for stem and progenitor cell properties indicated that the administration of recombinant colony stimulating factors (CSF) increases the concentration of CD34+ cells from 3.8 ± 0.8 x 103 to 61.9 ± 11.3 x 103/ml blood [21]. A subsequent leukocytapheresis is then capable of collecting about 5 x 108 CD34+ cells, which is about 26 times the number of CD34+ cells normally in the blood stream. There is also evidence that such "perturbations" of the pool of circulating stem and progenitor cells can only be explained by a selective release of such cells from the bone marrow into the blood. It is evident that the perturbance is temporary and that after several days the system returns to its steady-state situation [22].
|
Stem Cell Migration Streams During Bone Marrow Development
|
|---|
Evidence for the important role of stem and progenitor cells in the peripheral blood for hematopoietic cell renewal comes from the studies on embryonic and fetal development of hematopoiesis. It is now accepted, in contrast with the classical views of hematopoietic development [23], that hematopoiesis in the bone marrow cavity is the result of the seeding of hematopoietic stem cells onto a matrix characterized by a very specific innervated vascular and cellular structure within a firm bony capsule [24]. This process has been studied in detail in rats [25, 26], in dogs [27], and in man [28]. In these mammalian organisms, there is a characteristic development of hematopoiesis in each bone cavity. The cartilage in the skeletal part becomes necrobiotic, leaving a cavity in which the mesenchymal elements of the perichondrium penetrate followed by blood vessels and nerves. All these elements form the stroma or matrix of the marrow in which blood-borne stem cells find the adequate microenvironment to divide, replicate, and/or differentiate [29, 30].
In dogs, it can be shown that there is in each skeletal bone an identical sequence: from cartilage (C) to a prehemopoietic stroma (S) to hemopoiesis (H) (Table 2 ). The time for the first bones in the dog to develop a prehematopoietic stroma is day 37 to 38 of gestation, and it takes the entire fetal development to colonize all bone cavities. In man, the stromal matrix becomes established in the clavicle as early as six to eight weeks of gestation. Fifteen out of 38 weeks of human development are needed to establish "the" bone marrow distributed throughout the entire skeleton [24, 28].
It is now of interest to examine the hematopoietic development in the bone marrow from the viewpoint of blood stem cell physiology. The concept can be described as follows (Fig. 6): it is possible to conceive the establishment of hematopoiesis in the marrow as a consequence of the introduction of hemopoietic stem and progenitor cells into the blood entering and leaving the vascular system of the marrow matrix. These embryonic stem and progenitor cells in the blood can be measured. Nothdurft et al. [27] were able to show that the blood of a dog on its 35th day of gestation, when the liver contains already some 50 x 105 GM-CFC, shows a concentration of some 31 x 103 GM-CFC per ml (Fig. 7). Thereafter, the numbers decline to reach about 4 x 103 pre-partum (adult dog about 200-300/ml). In other words, the blood stem and progenitor cell concentration during hemopoietic development is at a factor of 30 or more higher than in the adult steady-state situation. Hence it is likely that the stem cell egress from the blood into the marrow matrix exceeds by far the egress of stem cells from the marrow into the blood until the marrow sites have established a suitable stem cell concentration.
View larger version (24K):
[in this window]
[in a new window]
|
Figure 6. Case 3: Hemopoietic development of stem cell migration during embryogenesis. There is evidence for the migration of stem and progenitor cells through the blood stream into the bone marrow.
|
|
View larger version (27K):
[in this window]
[in a new window]
|
Figure 7. Absolute numbers of GM-CFC in the yolk sac and liver and their concentration per ml blood in canine fetuses between days 20 and 57 to 59 of gestation. Values are also shown for two pups on day 4 post partum and two collectives of adult dogs [27].
|
|
In the human being, the pattern of hemopoietic development in relation to blood stem cell migration is identical in principle. During weeks 10 and 11 of gestation, the blood contains about 134 CFU-C/ml. At this time, only a few bones are ready to accept immigrating stem cells. During weeks 16 to 18 of gestation, when bones in all parts of the skeleton are ready to allow seeding of stem cells, the concentration of stem and progenitor cells reaches a peak value of as many as 65,000 CFU-C/ml. At the time of birth when it can be assumed that all suitable bone marrow sites have been seeded with stem and progenitor cells, their concentration is down to about 10,000 CFU-GM/ml [24].
It is, therefore, concluded that the embryonic and fetal development of hematopoiesis in the bone marrow can be compared to the seeding of stem cells to a suitable, well-prepared, and biochemically characterized bone matrix environment showing at the time of the colonization the microscopic picture of an "aplastic" marrow. In this morphological and functional sense, it may be speculated that a blood stem cell transfusion results in essentially the same pattern of hemopoietic development as seen during embryogenesis.
|
Stem Cell Migration Streams After Partial-Body Irradiation
|
|---|
A classical example of stem cell migration streams comes from radiobiological research. Evidence for the fact that intact stem cells can migrate from one part of the bone marrow to the other was derived from studies in mice in which radiation was given in the lethal range but in a way that one part of the body was shielded during irradiation. This resulted in a significant decrease of the (LD) 50/30 days. Hartweg [31], as early as 1954, showed the protective effect of shielding a femur in an otherwise lethally irradiated rat. In shielding experiments, several authors have demonstrated that a lethally irradiated organism could be saved if the cell migration streams would be established from the nonirradiated to the irradiated part of the body [32-34]. In human beings, it is well known that bone marrow sites receiving therapeutic exposure doses up to several 1,000 cGy can recover by endogenous stem cell migration [35].
Thus we can, in principle, depict the following scheme (Case 4, Fig. 8). If a fraction of bone marrow is shielded (upper part), then one would postulate that a migration of stem cells would commence from this part of the bone marrow to bone marrow parts irradiated highly enough [36, 37]. One would then expect an emigration from stem and progenitor cells from blood into irradiated bone marrow sites until a new steady state occurs.
View larger version (22K):
[in this window]
[in a new window]
|
Figure 8. Case 4: Hemopoietic reconstitution after partial-body irradiation. It is assumed that in the nonirradiated part of the body, there is an influx of stem and progenitor cells from the blood derived from nonirradiated parts of the bone marrow.
|
|
In our own group, we have studied associated problems both in rats [38] and in dogs [39, 40]. In the rat, we were interested in finding out whether "resting bone marrow mononuclear cells" that had been shown in transfusion experiments to be associated with hemopoietic restoration potential and that were labeled by tritiated thymidine using the "complete thymidine labeling method" [41] would start to proliferate when distant bone marrow parts were irradiated. This indeed was the case; the resting mononuclear cells of the protected bone marrow sites were recruited to proliferate and differentiate at the time when stem and progenitor cells were needed to repopulate the irradiated bone marrow sites. In a smilar way, Micklem and Ford were able to demonstrate in mice, using chromosomal markers, under what circumstances stem and progenitor cells can be recruited to migrate from a protected marrow site into an irradiated marrow area [42].
These problems of stem cell migration were studied most extensively by Nothdurft et al. [39, 40]. It is sufficient here to point out that a myeloablative dose to 70% of the bone marrow while 30% was shielded results in a perturbation of the GM-CFU fraction of the shielded marrow. In the irradiated parts, obviously, virtually all GM-CFU were destroyed within one day after irradiation (Fig. 9A). The blood pool of GM-CFU showed a decrease after partial-body irradiation and "overshoot values" between days 10 and 35 after irradiation—which means during the time of hemopoietic reconstitution in the irradiated marrow parts.
If 30% of the total marrow mass were irradiated and 70% protected [40], one could again observe an initial decrease of circulating GM-CFC immediately after partial-body exposure, but then during the re-establishment of hemopoiesis in the irradiated bone marrow sites an overshooting reappearance of circulating GM-CFC associated with a gradual return of hematopoiesis in the irradiated sites (Fig. 9B
).
In summary, the observations are in agreement with the assumption that blood stem cell migration plays a crucial role in re-establishing a sufficient level of stem cells in bone marrow units that were rendered aplastic by irradiation or other means, for instance, mechanical depletion as shown by Meyer-Hamme et al. [43].
|
Stem Cell Migration to Reconstitute Hematopoiesis
|
|---|
The role of blood stem and progenitor cells can also be considered and appreciated in all those cases in which bone marrow hematopoiesis is damaged or eradicated by total body radiation exposure or after myeloablative chemotherapy. In all these cases, the following scheme might help to elucidate the problems on hand.
In a situation schematically depicted in Figure 10 (Case 5), hematopoietic stem and progenitor cells are introduced into the blood pool derived from exogenous stem cell sources.
View larger version (22K):
[in this window]
[in a new window]
|
Figure 10. Case 5: Hemopoietic reconstitution after myeloablative irradiation or chemotherapy. The objective of blood stem cell transplantation is the hemopoietic reconstitution which can only occur by stem cell migration.
|
|
In our group, back in the days of collaboration between the Brookhaven Laboratory and the Mary Imogene Basset Hospital in Cooperstown, we studied the disappearance rate of transfused bone marrow precursor cells in dogs labeled in vitro in their DNA with tritiated thymidine [44]. The bulk of labeled cells disappeared from the circulating blood within two to three h but could not be traced to bone marrow sites. Today we know that the bulk of pluripotent stem cells is not in DNA synthesis but at rest [45], and it was only in studies using the reverse approach—activation of thymidine-labeled resting mononuclear cells by partial-body irradiation and/or by their transfusion into lethally irradiated recipient animals—that we came to the appropriate assumption that the pluripotent stem cell is a resting cell capable of migration through the blood [46], but not recircling through the lymph [47], that can home in hematopoietic stroma prepared to accept this seeding process in medullary but also in appropriate extramedullary sites [48].
This notion is in accordance with studies of other groups in mice [49], dogs [50], and monkeys [51], supporting the concept that it is the role of blood stem cells to replenish the stem cell pool of hematopoietic tissue in case of a stem cell concentration deficit.
This was substantiated by our previous studies in dogs. In the first experiments studying blood stem cell transplantation using CFU-C as an indicator for the presence of progenitor cells in the transfusate, it was quite evident that the number of CFU-C in the transfusate determined the quality and quantity of bone marrow restoration. The histology of dog bone marrow 10 days after a lethal whole-body exposure and transfusion of blood mononuclear cells containing 7.5 x 105 CFU-C showed only a few spongiosa niches with full hemopoietic recovery while adjacent spongiosa niches were completely aplastic. If 15 x 105 CFU-C were in the transfusate, more niches were filled but others were still completely empty. It was only after administration of some 30 x 105 CFU-C that all niches were completely recovered within 10 days [52-54].
This prompted us to examine the quality and quantity of stem cells collected from different sources, including blood, in terms of their regenerative potential. Comparing transfusates of mononuclear cells from fetal liver, bone marrow, and peripheral blood containing equal numbers of CFU-C, it was found that the recovery of blood granulocytes was very quick when fetal liver cells and blood mononuclear cells were used in comparison to bone-marrow-derived cells. However, the complete recovery of blood granulocytes after blood stem cell transplantation was delayed in comparison to fetal liver transplantation [55, 56] (Fig. 11).
View larger version (26K):
[in this window]
[in a new window]
|
Figure 11. The immigration of stem cells from blood, from bone marrow, and from fetal liver in dogs was analyzed on the basis of the recovery of blood granulocytes after total-body irradiation and stem cell transfusion. Blood-derived stem cells have a similar initial course of granulocyte recovery in comparison to fetal liver cells. However, fetal liver cell transfusion allows the granulocyte concentration to return to normal and even to overshoot within 20 days.
|
|
We developed and used a biomathematical model [57] to try to understand these differences and found that the fetal cells must have a replicative power that exceeds that of bone-marrow- and blood-derived stem cells. While the computer model could fit the recovery data of bone-marrow- and blood-derived stem cells utilizing a replication probability value of 0.63 [58], it was necessary to assume a replication probability of 0.95 for fetal stem cells [57].
The same biomathematical model was used to study the relationship between the number of blood stem cells in a transfusate measured as CD34+ cells and hematopoietic recovery in patients with multiple myeloma treated with cyclophosphamide, busulfan, and thiotepa at the M.D. Anderson Hospital under the leadership of Dr. M. Körbling and his colleagues [59-61]. The simulation model developed first by Steinbach [62] and extended by Hofer and Tibken [63] consists of eight cellular and two regulatory compartments [64] (Figs. 12A, 12B). It assumes a homeostatic equilibrium which has to guarantee that for each granulocyte leaving the circulation by senescence or emigration one granulocyte enters the circulation from the bone marrow (compartment F). For each cell leaving the extravascular bone marrow sites there has to be a net gain of one granulocyte through cell division (compartments S, CBM, and P). The life spans of the cells involved and their proliferation rates as well as cell cycle characteristics are largely known [65]. It can also be assumed that there are feedback regulatory mechanisms mediated through regulatory molecules such as cytokines [66, 67]. For this model, two regulatory compartments are assumed, and each of them would contain a balance of stimulatory and inhibitory effects (Reg. I and II). The control scheme of granulocytopoiesis is given in Fig. 12B (and the essential differential equations are described in mathematical detail elsewhere) [68, 69]. It is sufficient here to indicate that the model consists, in terms of regulation technology, of bilinear subsystems which are connected with each other by nonlinear static transfer linkages. This type of bilinear system is very useful in order to describe the cell proliferation and fluxes in the different cell compartments. Changes in the input values of a bilinear system are able to describe increases as well as decreases of cell numbers. The modeling of cell regulatory processes becomes a very natural way of using bilinear systems but requires no fewer than 37 differential equations.
View larger version (40K):
[in this window]
[in a new window]
|
Figure 12. (A) Model of granulocytopoiesis used as the basis for a biomathematical model to assess radiation effects. (B) Control scheme of granulocytopoiesis (the details of the differential equations used are given in [63, 64, 68, 69]).
|
|
This model was used to simulate the blood granulocyte recovery in five multiple myeloma patients given autologous blood stem cell transfusates containing between 0.24 and 17.54 x 106 CD 34+ cells per kg body weight [61]. It was possible to simulate the recovery pattern and calculate the number of "biomathematical stem cell units" that must have been in the transfusate to achieve this type of recovery (Patients No. 20 and No. 11) (Figs. 13A, 13B). It was gratifying to see that there was a linear relationship between the number of CD 34+ blood cells in the transfusate and the number of calculated (mathematical) stem cells. This indicated a new approach to relate the quality as well as the quantity of blood stem cell suspensions to the pattern of hemopoietic recovery and hence also to its quality and quantity (Fig. 14).
View larger version (31K):
[in this window]
[in a new window]
|
Figures 13A, 13B. Computer simulation curves of the granulocyte recovery in two patients (No. 20 and No. 11) with multiple myeloma treated with high-dose chemotherapy and showing rapid granulocyte recovery.
|
|
View larger version (11K):
[in this window]
[in a new window]
|
Figure 14. The correlation is shown between the number of CD positive cells/kg body weightx106as used in the multiple myeloma treatment scheme [6] in relation to the number of stem and progenitor cells calculated from the biomathematical model [59] by means of simulating the granulocyte recovery curve.
|
|
From such concepts and experimental studies, it appears justified to conclude that blood-derived stem cells must have qualitative properties that enable them to circulate in the peripheral blood without necessarily being "trapped" but capable of migrating to sites of tissue within which they can "home." Apparently, this property of entering and remaining in the blood stream has to do with adhesion receptors, especially of the integrin family [70-72]. Their homing within tissues (medullary and extramedullary matrix such as in liver and/or spleen) is likely to depend on "local tissue" conditions (as described by a number of authors [69]) and also from stem cell surface properties as described by others [73-75]. Thus, it will be of great importance to study in detail the physiological and pathophysiological properties of blood stem cells that were introduced into the circulation and to follow their fate and function and their transforming potentials.
|
The Role of Blood Stem Cells in Hematopoietic Cell Renewal
|
|---|
This keynote presentation for the International Workshop on "Pathophysiology, Diagnostic and Therapeutic Implications of Blood Stem Cells" served the purpose of reviewing the present concepts regarding the role of blood stem cells in hematopoietic cell renewal.
A number of conclusions might be appropriate:
- In the steady-state situation of blood cell production and removal, there are stem and progenitor cells present in the circulation. They can be identified by their surface properties, clonogenic potentials, and biophysical properties, while their morphology is indistinguishable from the bulk of cells called "lymphocytes." Additional work is necessary before one begins to understand their life cycle, their migratory properties, and their fate after immigrating into the circulation.
- Blood stem cells are apparently in an equilibrium with extravascular sites; leukocytapheretic procedures indicate that there is an extravascular reserve of an easily mobilizable pool of stem and progenitor cells which can even be further expanded by the administration of specific stimulatory molecules. Under steady-state and conditions "early" after apheresis, the physical properties of blood stem cells are similar to those of the easily mobilizable cells. It remains to be determined in what way the quality of blood stem cells changes as a function of time after recombinant cytokine stimulation. The type of perturbations after CFC indicates that blood stem cells are in a feedback regulated equilibrium with bone marrow stem cells.
- Blood stem cells are of decisive importance for the establishment of hematopoiesis in all skeletal bones. Apparently, the hematogenous seeding of fetal stem cells results in a "filling-up" of the local (semiautonomous) hematopoietic sites in the bone marrow. If there is additional need for hematopoiesis, such as in certain disease states (polycythemia, thalassemia, etc.) then "fatty marrow" can become hematopoietic, most likely as a result of stem cell seeding.
- The blood stem cell migration streams become most obvious in "enforced" migration, as observed in partial body irradiation, but also in the process of establishing extramedullary hematopoiesis.
- If hematopoietic stem cells are introduced into the circulating blood, it is obvious that "blood-derived" stem cells have migratory potentials most suitable for "homing" in appropriate hematopoietic microenvironments. More work is needed to characterize the migration properties and streams in the steady-state situation of hematopoiesis and in diseased conditions, especially "stem cell disorders."
|
Acknowledgments
|
|---|
Throughout the years, the following scientists (among others) have contributed in particular to the experimental work of our group: W. Calvo, F. Carbonell, H.D. Flad, H.H. Gerhartz, G. Grilli, R.J. Haas, E.B. Harriss, E. Herbst, D. Hoelzer, S. Issaragrisil, M. Körbling, P. Kovacs, H. Kreutzmann, W. Nothdurft, H. Pflieger, O. Prümmer, A. Raghavachar, W.M. Ross, H.J. Seidel, K.H. Steinbach, and B.L. Ziegler.
The research work was supported by the European Commission, Brussels, the Ministries of the Federal Government of Germany, and the State of Baden-Wuerttemberg.
From CHARACTERISTICS AND POTENTIALS OF BLOOD STEM CELLS. STEM CELLS 1998;16(suppl 1):13-29. Contact AlphaMed Press for information about this supplement.
|
References
|
|---|
-
Maximow A. Der Lymphozyt als gemeinsame Stammzelle der verschiedenen Blutelemente in der embryonalen Entwicklung und im postfetalen Leben der Säugetiere. Folia Haematologica VIII 1909;8:125-134.
-
Lee GR et al. Wintrobe's Clinical Hematology. Philadelphia, London: Lea and Febiger, 1993.
-
Goodman JW, Hodgson G. Evidence for stem cells in the peripheral blood of mice. Blood 1962;19:702-714.[Abstract/Free Full Text]
-
Micklem HS, Anderson N, Ross E. Limited potential of circulating haemopoietic stem cells. Nature 1975;256:41-43.[Medline]
-
Fliedner TM, Steinbach KH, Hoelzer D. Adaptation to environmental changes: the role of cell-renewal systems. In: Finckh, ed. The Effects of Environment on Cells and Tissue. Amsterdam/Oxford: Excerpta Medica, 1976:20-38.
-
Metcalf D, Moore MAS. Haemopoietic Cells. Amsterdam: North-Holland Publishing Co, 1971.
-
Till JE, McCulloch EA. A direct measurement of the radiation sensitivity of normal mouse bone marrow cells. Radiat Res 1961;14:213-222.[Medline]
-
Micklem HS. Effect of phytohemagglutinin-M (PHA) on the spleen-colony-forming capacity of mouse lymph node and blood cells. Transplantation 1966;4:732-741.[Medline]
-
Nothdurft W, Fliedner TM. The response of the granulocytic progenitor cells (CFU-C) of blood and bone marrow in dogs exposed to low doses of x-irradiation. Radiat Res 1992;89:38-52.
-
Kreutzmann H, Fliedner TM. Studies on the presence and possible oscillation of granulocytic progenitor cells (CFU-C) in human blood. Scand J Haematol 1979;23:360-366.[Medline]
-
Grilli G, Carbonell F, Fliedner TM. Variations in erythroid and myeloid progenitor cell numbers in normal human peripheral blood. Br J Haematol 1980;44:679-681.[Medline]
-
Hodgson G, Guzman E, Herrera C. Characterization of the stem cell population of phenyl hydrazine treated rodents. In: Doyle E, ed. Effects of Radiation on Cellular Proliferation and Differentiation. Vienna: IAEA, 1968:163-170.
-
Moloney MA, Patt HM. Marrow stem cell release in the autorepopulation assay. Exp Hematol 1978;6:227-232.[Medline]
-
Raghavachar A, Steinbach KH, Prümmer O et al. Survival of transfused cryopreserved granulocytic progenitor cells (CFU-C) in recipient circulation. Cell Tissue Kinet 1983;16:303-311.[Medline]
-
Nothdurft W, Steinbach KH, Ross WM et al. Quantitative aspects of granulocytic progenitor cell (CFUc) mobilization from extravascular sites in dogs using dextran sulfate. Cell Tissue Kinet 1982;15:331-340.[Medline]
-
Fliedner TM, Calvo W, Körbling M et al. Hematopoietic stem cells in blood: characteristics and potentials. In: Golde DW, Cline MJ, Metcalf D et al., eds. Hematopoietic Cell Differentiation. ICN-UCLA Symposia on Molecular Biology. New York: Academic Press 1978:193-212.
-
Körbling M, Fliedner TM, Pflieger H. Collection of large quantities of granulocyte/macrophage progenitor cells (CFUS) in man by continuous flow leukapheresis. Scand J Haematol 1980;24:22-28.[Medline]
-
Gerhartz HH, Fliedner TM. Velocity sedimentation and cell cycle characteristics of granulopoietic progenitor cells (CFU-C) in canine blood and bone marrow: influence of mobilization and CFU-C depletion. Exp Hematol 1980;8(suppl 2):209-218.[Medline]
-
Raghavachar A, Prümmer O, Calvo W et al. Repopulating potential of canine bone marrow cells: differences between large and small cells separated by velocity sedimentation. Br J Haematol 1985;60:33-40.[Medline]
-
Fliedner TM, Körbling M, Arnold R et al. Collection and cryopreservation of mononuclear blood leukocytes and of CFU-C in man. Exp Hematol 1979;7(suppl 5):398-408.
-
Körbling M, Anderlini P, Durett A et al. Delayed effects of rhG-CSF mobilization treatment and apheresis on circulating CD34+ and CH34+Thy-ldim CD38+ progenitor cells, and lymphoid subsets in normal stem cell donors for allogeneic transplantation. Bone Marrow Transplant 1996;18:1073-1079.[Medline]
-
Nothdurft W, Fliedner TM. Stem cell migration after irradiation. In: Okada S et al., eds. Radiation Research. Tokyo: Toppan, 1979:657-663.
-
Doan CA. On the origin and developmental potentialities of blood cells. Acad Med 1939;15:668-697.
-
Fliedner TM, Calvo W. Hematopoietic stem cell seeding of a cellular matrix: a principle of initiation and regeneration of hematopoiesis. In: Clarkson B et al., eds. Cold Spring Harbor Conferences on Cell Proliferation. Cold Spring Harbor aboratory, 1978;5:757-773.
-
Calvo W, Haas RJ. Die Histogenese des Knochenmarkes der Ratte. Z Zellforsch 1969;95:377-395.[Medline]
-
Haas RJ, Hoelzer D, Kurrle E et al. Experimental analysis of developing haematopoiesis in fetal bone marrow. Pediatr Res 1976;10:164-168.[Medline]
-
Nothdurft W, Braasch E, Calvo W et al. Ontogeny of the granulocyte/macrophage progenitor cell (GM-CFV) pools in the beagle. J Embryol Exp Morph 1984;80:87-103.[Medline]
-
Keleman E, Calvo W, Fliedner TM. Atlas of Human Hemopoietic Development. Berlin: Springer-Verlag, 1979:
-
Tavassoli M. Embryonic and fetal hemopoiesis: an overview. Blood Cells 1991;1:269-281.
-
Zon LI. Developmental biology of hematopoiesis. Blood 1995;86:2876-2891.[Abstract/Free Full Text]
-
Hartweg H. Die Wirkung geschützten homologen Knochenmarks auf die Regeneration des haemopoetischen Systems nach dem Strahleninsalt. Strahlentherapie 1954;95:594-604.[Medline]
-
Swift MN, Takta ST, Bond VP. Regionally fractionated x-irradiation equivalent in dose to total body exposure. Radiat Res 1954;1:241-252.[Medline]
-
Hanks GE. In vivo migration of colony forming units from shielded bone marrow in the irradiated mouse. Nature 1964;203:1393-1395.
-
Croizat H, Frissdal E, Tubiana M. The effects of partial body irradiation on haemopoietic stem cell migration. Cell Tissue Kinet 1980;13:319-325.[Medline]
-
Chone B. Knochenmarkveränderungen im Rahmen der Strahlentherapie in morphologischer und elektronenoptischer Sicht. Nucl Med 1963;2:425-433.
-
Micklem HS, Ford CE, Evans EP et al. Compartments and cell flows within the mouse haemopoietic system. I. Restricted interchange between haemopoietic sites. Cell Tissue Kinet 1975;8:219-232.[Medline]
-
Micklem HS, Ogden DA, Evans EP et al. Compartments and cell flows within the mouse haemopoietic system. II. Estimated rates of interchange. Cell Tissue Kinet 1975;8:233-248.[Medline]
-
Haas R, Bone F, Fliedner TM. Cytokinetic analysis of slowly proliferating bone marrow cells during recovery from radiation injury. Cell Tissue Kinet 1971;4:31-45.[Medline]
-
Nothdurft W, Calvo W, Klinnert V et al. Acute and long-term alterations in the granulocyte/macrophage progenitor cell (GM-CFC) compartment of dogs after partial body irradiation. Irradiation of the upper body with a single myeloablative dose. Int J Radiat Oncol 1986;12:949-957.[Medline]
-
Baltschukat K, Fliedner TM, Nothdurft W. Hematological effects in dogs of the lower part of the body with a single myeloablative dose. Radiother Oncol 1989;14:239-246.[Medline]
-
Fliedner TM, Haas RJ, Stehle H et al. Complete labeling of all cell nuclei in new-born rats. A tool for the evaluation of rapidly and slowly proliferating cell systems. Lab Invest 1968;18:249.[Medline]
-
Micklem HS, Ford CD, Evans EP et al. Interrelationships of myeloid and lymphoid cells: studies with chromosome-marked cells transfused into lethally irradiated mice. Proc Roy Soc 1966;165:78-102.
-
Meyer-Hamme K, Haas RJ, Fliedner TM. Cytokinetics of bone marrow stroma cells after stimulation by partial depletion of the medullary cavity. Acta Haematol 1971;46:349-361.[Medline]
-
Fliedner TM, Thomas ED, Meyer LM et al. The fate of transfused H3-thymidine labeled bone marrow cells in irradiated recipients. Ann NY Acad Sci 1964;114:510-526.
-
To LB, Haylock DN, Döwse T et al. A comparative study of the phenotype and proliferative capacity of peripheral blood CD34+ cells mobilized by four different protocols and those of steady-phase PB and bone marrow CD34+ cells. Blood 1994;84:2930.[Abstract/Free Full Text]
-
Haas RJ, Flad HD, Fliedner TM et al. Correlation between cytokinetically resting lymphocytes and bone marrow restoration: experiments using a discontinuous albumin gradient. Blood 1973;42:209-218.[Abstract/Free Full Text]
-
Storb R, Epstein RB, Thomas ED. Marrow repopulating ability of peripheral blood cells compared to thoracic duct cells. Blood 1968;32:662-667.[Abstract/Free Full Text]
-
Fliedner TM, Wandl UB, Calvo W. Medulläre und extramedulläre Haemopoese im Hund nach Ganzkörperbestrahlung und Transfusion von aus dem Blut gewonnenen Stammzellen. Schweizer Medizinische Wochenschrift 1982;112:1423-1429.
-
Barnes DW, Loutit JF. Effects of irradiation and antigenic stimulation on circulating haemopoietic stem cells of the mouse. Nature 1967;213:1142-1143.[Medline]
-
Abrams RA, McCormack K, Bowles C et al. Cyclophosphamide treatment expands the circulating hematopoietic stem cell pool in dogs. J Clin Invest 1981:67:1392-1399.
-
Storb R, Graham TC, Epstein RB et al. Demonstration of hemopoietic stem cells in the peripheral blood of baboons by cross circulation. Blood 1977;5:537-542.
-
Calvo W, Fliedner TM, Herbst E et al. Regeneration of blood-forming organs after autologous leukocyte transfusion in lethally irradiated dogs. II. Distribution and cellularity of the marrow in irradiated and transfused animals. Blood 1967;47(suppl 4):593-601.[Abstract/Free Full Text]
-
Fliedner TM, Flad HD, Bruch CH et al. Treatment of aplastic anemia by blood stem cell transfusion: a canine model. Haematologica 1976;61:141-156.[Medline]
-
Nothdurft W, Bruch CH, Fliedner TM et al. Studies on the regeneration of the CFUc-population in blood and bone marrow of lethally irradiated dogs after autologous transfusion of cryopreserved mononuclear blood cells. Scand J Haematol 1977;19:470-481.[Medline]
-
Prümmer O, Raghavachar A, Werner C et al. Fetal liver transplantation in the dog. I. Restoration of haemopoiesis with cryopreserved fetal liver cells from DLA-identical siblings. Transplantation 1985:39;349-355.
-
Prümmer O, Werner C, Raghavachar A et al. Fetal liver transplantation in the dog. II. Repopulation of the granulocyte-macrophage progenitor cell compartment by fetal liver cells from DLA-identical siblings. Transplantation 1985;40:498-503.[Medline]
-
Fliedner TM, Steinbach KH. Simulationsmodelle von Perturbationen des Granulozytären Zellerneuerungssystems. In: Doerr W, Schipperges H, eds. Modelle der Pathologischen Physiologie. Berlin, Heidelberg, New York: Springer-Verlag, 1987:89-106.
-
Vogel H, Niewisch H, Matioli G. The self renewal probability of hemopoietic stem cells. J Cell Physiol 1968;72:221-228.[Medline]
-
Paul W. Die Analyse der Regeneration der Granulopoese nach Stammzelltransplantationen mit Hilfe einer Regelungstechnischen Implementation eines Biomathematischen Modells. Inaugural-Dissertation Universität Ulm, 1997.
-
Körbling M, Huh YO, Durrett A et al. Allogeneic blood stem cell transplantation: peripheralization and yield of donor-derived primitive hematopoietic progenitor cells (CD34+Thy-1dim) and lymphoid subsets, and possible predictors of engraftment and GVHD. Blood 1995;86:2842-2848.[Abstract/Free Full Text]
-
Alexanian R, Dimopoulos MA, Hester J et al. Early myeloablative therapy for multiple myeloma. Blood 1994;84:4278-4282.[Abstract/Free Full Text]
-
Steinbach KH, Raffler H, Pabst G. A mathematical model of canine granulocytopoiesis. J Math Biol 1980:10:1-12.[Medline]
-
Hofer EP, Tibken B, Fliedner TM. Modern control theory as a tool to describe the biomathematical model of granulopoiesis. In: Möller DPF, Richter O, eds. Analyse Dynamischer Systeme in Medizin, Biologie und Ökologie. Informatik-Fachberichte. Berlin: Springer Verlag, 1991;275:33-39.
-
Fliedner TM, Tibken B, Hofer EP et al. Stem cell responses after radiation exposure: a key to the evaluation and prediction of its effects. Health Phys 1996:70;787-797.[Medline]
-
Cronkite EP, Fliedner TM. Granulopoiesis. New Engl J Med 1964;270:1347, 1403.
-
Golde DW. Hematopoietic growth factors - An Overview. In: Murphy MJ, Rizzoli V, eds. Blood Cell Growth Factors: Their Biology and Clinical Application. Int J Cell Cloning 1990;8:4-10.
-
Broxmeyer HE. Biomolecule-cell interaction and the regulation of myelopoiesis: an update. In: Murphy MJ, ed. Concise Reviews in Clinical and Experimental Hematology. Dayton: AlphaMed Press, 1992:119-147.
-
Tibken B, Hofer EP. A biomathematical model of granulocytopoiesis for estimation of stem cell numbers. STEM CELLS 1995;13(suppl 1):282-289.
-
Tibken B, Hofer P. Constrained optimization algorithms and automatic differentiation for parameter estimation with applications to granulocytic models. In: Dolezal J, Fidler J, eds. System Modelling and Optimation. Proc 17th IFIP TC7 Conf on System Modelling and Optimization. Prag. London: Chapman and Hall, 1996:115-119.
-
To LB, Haylock DN, Simmons PJ et al. The biology and clinical uses of blood stem cells. Blood 1997;89:2223-2258.[Free Full Text]
-
Papayannopoulou T, Nakamoto B. Peripheralization of hemopoietic progenitors in primates treated with anti-VLA4 integrin. Proc Natl Acad Sci USA 1993;90:9374-9378.[Abstract/Free Full Text]
-
Tavassoli M, Hardy CL. Molecular basis of homing of intravenously transplanted stem cells. Blood 1990;76:1059-1070.[Free Full Text]
-
Turner ML. Regulation of hematopoietic progenitor cell migration, mobilization and homing. STEM CELLS 1994:12:227-229.[Medline]
-
Levesque JP, Haylock DN, Simmons PJ. Cytokine regulation of proliferation and cell adhesion are correlated events in human CD34+ hemopoietic progenitors. Blood 1996;88:1168-1176.[Abstract/Free Full Text]
-
Long MW. Blood cell cytoadhesion molecules. Exp Hematol 1992;20:288-301.[Medline]
This article has been cited by other articles:
|
|
|
|
 
T. Papayannopoulou and D. T. Scadden
Stem-cell ecology and stem cells in motion
Blood,
April 15, 2008;
111(8):
3923 - 3930.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
I. Shuryak, R. K. Sachs, L. Hlatky, M. P. Little, P. Hahnfeldt, and D. J. Brenner
Radiation-Induced Leukemia at Doses Relevant to Radiation Therapy: Modeling Mechanisms and Estimating Risks
J Natl Cancer Inst,
December 20, 2006;
98(24):
1794 - 1806.
[Abstract]
[Full Text]
[PDF]
|
|
|
Top
Abstract
Introduction: Scope and Purpose
= donor 1; • = donor 2; 
= donor 3. For donor 1: the mean values ± standard deviation were for BFU-E 501 ± 98, for CFU-C/ml blood 209 ± 95 (n=17). For donor 2: BFU-E/ml blood 200 ± 81, CFU-C/ml blood 29 ± 12 (n=17). For donor 3: BFU-E/ml blood 624 ± 202 and CFU-C/ml blood 102 ± 40 (n=19).
