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Endocrinology Vol. 143, No. 2 690-699
Copyright © 2002 by The Endocrine Society

GRH-SOMATOSTATIN-GH

Age-Associated Loss of Bone Marrow Hematopoietic Cells Is Reversed by GH and Accompanies Thymic Reconstitution

Richard A. French1, Suzanne R. Broussard, William A. Meier, Christian Minshall, Sean Arkins2, James F. Zachary, Robert Dantzer and Keith W. Kelley

Laboratory of Immunophysiology (R.A.F., S.R.B., C.M., S.A., K.W.K.), Department of Animal Sciences, and Department of Veterinary Pathobiology (W.A.M., J.F.Z.), University of Illinois at Urbana-Champaign, Urbana, Illinois 61801; and Institut National de la Recherche Agronomique-Institut National de la Santé et de la Recherche Médicale Unité 394, Unité de Recherches de Neurobiologie Intégrative (R.D.), Institute François Magendie, 33077 Bordeaux, France

Address all correspondence and requests for reprints to: Keith W. Kelley, Laboratory of Immunophysiology, University of Illinois, 207 Edward R. Madigan Laboratory, 1201 West Gregory Drive, Urbana, Illinois 61801. E-mail: kwkelley{at}uiuc.edu


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Deterioration of the thymus gland during aging is accompanied by a reduction in plasma GH. Here we report gross and microscopic results from 24-month-old Wistar-Furth rats treated with rat GH derived from syngeneic GH3 cells or with recombinant human GH. Histological evaluation of aged rats treated with either rat or human GH displayed clear morphologic evidence of thymic regeneration, reconstitution of hematopoietic cells in the bone marrow, and multiorgan extramedullary hematopoiesis. Quantitative evaluation of formalin-fixed, hematoxylin and eosin-stained sections of bone marrow from aged rats revealed at least a 50% reduction in the number hematopoietic bone marrow cells, compared with that of young 3-month-old rats. This age-associated decline in bone marrow leukocytes, as well as the increase in bone marrow adipocytes, was significantly reversed by in vivo treatment with GH. Restoration of bone marrow cellularity was caused primarily by erythrocytic and granulocytic cells, but all cell lineages were represented and their proportions were similar to those in aged control rats. On a per-cell basis, GH treatment in vivo significantly increased the number of in vitro myeloid colony forming units in both bone marrow and spleen. Morphological evidence of enhanced extramedullary hematopoiesis was observed in the spleen, liver, and adrenal glands from animals treated with GH. These results confirm that GH prevents thymic aging. Furthermore, these data significantly extend earlier findings by establishing that GH dramatically promotes reconstitution of another primary hematopoietic tissue by reversing the accumulation of bone marrow adipocytes and by restoring the number of bone marrow myeloid cells of both the erythrocytic and granulocytic lineages.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
PLASMA CONCENTRATIONS OF GH and its growth-promoting peptide, IGF-I, are now well accepted to decline during aging (1). As animals age, there is a progressive atrophy of the thymus gland because of a loss of immature cortical thymocytes. This involution is associated with a reduction in the ability to produce new T cells to novel pathogens later in life and is accompanied by a decline in frequency of helper and cytotoxic T lymphocytes (2). Indeed, computer tomographic scans of cancer patients revealed that the ability to recover naive CD45RA+CD4+ cells following intensive chemotherapy requires functional thymic tissue (3). Several laboratories have reported that GH is thymotropic in aged (4, 5, 6, 7), severe combined immunodeficient (8, 9), dwarf ( 10, 11, 12), hypophysectomized (13), and azidothymidine- or 2cyclosporine-injected animals (14, 15).

The thymotropic effects of GH have been presumed to occur at the level of the thymus gland because implantation of GH3 cells into mice with a congenital absence of a thymus gland does not lead to the appearance of functional T cells (16). Similarly, CD4-CD8- cells accumulate in the thymus of aged rats (17) and several strains of aged mice (18), and this defect is reversed in rats by implantation of GH3 cells (17). This reversal could be because of the presence of receptors for both GH (19, 20) and IGF-I ( 21) on immature CD4-CD8- thymocytes or on thymic epithelial cells (22, 23). However, using an in vitro system consisting of sequential colonization of lymphoid-depleted fetal thymus stroma, Knyszynski et al. (6) established that GH increases thymocyte progenitors derived from the bone marrow. This is an important finding because these workers reported earlier that the frequency of thymic progenitors in the bone marrow decline with age (24). Hormones such as IGF-I may act by increasing the migration and colonization of these bone marrow-derived precursors to the thymus (25, 26). Injections of GH into young animals also increase DNA synthesis of bone marrow cells (27) and the number of bone marrow-derived hematopoietic progenitors (14, 28). Collectively, these data suggest that bone marrow cells are another important target for the action of GH (29, 30), even in aged animals (31).

The intrathymic role of GH and IGF-I in aging animals does not preclude the possibility that these hormones also act at the level of the bone marrow. Here we demonstrate that implantation of syngeneic GH3-secreting pituitary epithelial cells or injections of recombinant GH not only reverse morphological aspects of thymic aging but also dramatically reverse the hypocellularity of hematopoietic cells in the bone marrow in aged animals. Even when compared on a per-cell basis, the number of bone marrow-derived granulocyte-macrophage progenitor cells in aged rats is significantly increased by in vivo treatment with GH. The increase in bone marrow cellularity appears to be normal in that the distribution of hematopoietic cells of myelocytic, erythrocytic, lymphocytic, and megakaryocytic lineages is similar in aged rats and in aged rats treated with GH. Exogenous GH also causes an increase in extramedullary hematopoiesis (EMH) that can be visualized in the spleen, liver, and adrenal glands. These data establish that GH treatment in vivo leads to an impressive increase in hematopoietic potential of a number of tissues from aged animals.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals and reagents
Female Wistar-Furth rats were raised under specific-pathogen-free, barrier-reared conditions in microisolator cages for their entire life, as described previously (16). They were exposed to a 12-h light, 12-h dark cycle. The colony was screened every 3 months for all common infectious diseases of rats and mice. Young female control rats were 3 months of age. At 22 months, female rats were implanted sc into the lateral abdomen with 3 x 106 syngeneic GH3 cells (ATCC, Manassas, VA) in a volume of 0.5 ml of Ham’s nutrient mixture F10 (Sigma, St. Louis, MO), as previously described (5, 17, 32). At 23 months, female rats were injected sc (1 mg/kg) twice daily with recombinant human GH (gift from Genentech, Inc., San Francisco, CA) or control buffer for 28 d. At 24 months, rats in all treatment groups were killed by decapitation following CO2 asphyxiation. All procedures involving rats were performed according to guidelines established by the National Institutes of Health Guide for the Care and Use of Laboratory Animals and were approved by the University of Illinois Institutional Laboratory Animal Care Advisory Committee. The average life span of Wistar-Furth females raised under conventional conditions is 21 months (33).

Preparation of splenic and bone marrow single-cell suspensions
Both a femur and a tibia of Wistar-Furth rats were flushed through an 18-gauge needle with DMEM (Life Technologies, Inc., Gaithersburg, MD) that was supplemented with 3.7 g/liter sodium bicarbonate, 2 mM L-glutamine, 20 mM HEPES, 10 mM sodium pyruvate and nonessential amino acids, 100 U/ml penicillin, and 100 µg/ml streptomycin (Sigma). Single-cell suspensions were prepared (34) by passage through a 22-gauge needle followed by gravity sedimentation for 5 min, lysis of contaminating erythrocytes in ammonium chloride and three washes in DMEM. Single suspensions of spleen cells (1 x 106/ml) were prepared as previously described (34).

Histological preparation of tissues
Sections of thymus, spleen, liver, adrenal gland, and kidney were fixed by immersion in 10% neutral-buffered formalin, as previously described (35, 36). One femur from each rat was opened in a longitudinal plane with a rongeur and fixed by immersion in formalin. The fixed marrow was removed from the marrow cavity and was decalcified, post fixation, for 12–24 h in EDTA-formalin (5.5 g EDTA disodium salt in 90 ml H2O and 10 ml formalin). All tissues were embedded in paraffin, sectioned at 5 µm, mounted on charged slides (Fisher Scientific, Pittsburgh, PA) prewarmed to 60 C, deparaffinized with xylene, rehydrated in graded ethanols, and stained with hematoxylin and eosin (H&E). Permount was used to coverslip the stained sections, which were examined with an Optiphot-2 microscope (Nikon Corp., Melville, NY). Results were recorded by photomicrography.

Bone marrow cellularity was determined on H&E-stained, formalin-fixed midshaft femoral bone marrow sections. The total number of bone marrow cells was determined from the average number of cells on 10 (1 mm2) graticule grids of each tissue section. Using an oil immersion objective (100x), a field of bone marrow cells was randomly chosen. Sequential adjacent fields of cells were counted by rastering across the slide in a single direction.

Myeloid progenitor cell colony-forming assay
Bone marrow cells were suspended (2 x 105/ml) in Iscove’s modified Dulbeccos’s medium (Life Technologies, Inc.) containing 10% heat-inactivated charcoal-stripped FBS (Sigma; less than 25 pg endotoxin/ml) (34) and 0.66% soft agar (37). The major supplements to this semisolid medium consisted of 250 U murine IL-3 (Biosource International, Amarillo, CA) and 1.25 ng/ml of murine granulocyte-macrophage-colony stimulating factor (GM-CSF; R\|[amp ]\|D Systems, Minneapolis, MN). Bone marrow (2 x 104) and spleen (1 x 105) cells were cultured in duplicate in 1-ml volumes in 6-well plates (Costar, Cambridge, MA) at 37 C, 95% relative humidity, 7% CO2 for 7 d, at which time distinct colonies were counted in a blind fashion.

Statistical analysis
Experiments were conducted in a completely randomized design, and all data were analyzed by ANOVA using software from Statistical Analysis System (Cary, NC) (38). When treatment effects were found as determined by an F test (P < 0.05), differences among individual treatments were detected using Duncan’s multiple range test. Data were expressed as the mean ± SEM, and differences at P < 0.05 were considered to be significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
GH restores thymic morphology in aged rats
As expected, there was a clear loss of cortical thymocytes in 24-month-old rats, compared with 3-month-old controls (Fig. 1CGo vs. Fig. 1AGo), an effect that was most obvious at higher magnification (Fig. 1DGo vs. Fig. 1BGo). Thymic tissue from young rats had well-delineated cortical and medullary zones with clearly defined lobular architecture. In contrast, cortical and medullary zones from aged rats were poorly defined and were markedly hypocellular. There was also a significant widening of the interlobular septa with increased adiposity in aged rats, compared with their young controls. Following treatment of aged rats with GH of either rat or human origin, there was a dramatic reappearance of cortical thymocytes, compared with tissue from aged control rats (Fig. 1CGo vs. Fig. 1Go, E and G). The thymus glands of GH-treated aged rats exhibited increased numbers of thymocytes with well-defined cortical and medullary zones, compared with aged control rats (Fig. 1DGo vs. Fig. 1Go, F and H), displaying a lobular architecture that was similar to young control rats (Fig. 1AGo). Restoration of thymic morphology in aged rats by GH3 cells was accompanied by a nearly 2-fold increase (P < 0.05) in weight of the thymus relative to that of the kidneys (data not shown). These data establish and confirm (5) that GH reverses thymic involution in aged rats.



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  		Figure 1. GH increases both thymic lymphoid and bone marrow myeloid cells in aged rats. The most left-hand vertical panel is tissue stained with H&E that was derived from 3-month-old rats, and the second vertical panel depicts tissue from control aged 24-month-old rats. The third vertical panel is from aged rats treated with syngeneic GH3 cells, whereas the fourth vertical panel represents tissue from aged rats treated with recombinant GH. Thymic tissue is displayed in horizontal panels A, C, E, and G and B, D, F, and H (Figs. A, C, E, and G represent low magnification). Bone marrow tissue is shown in horizontal panels I, K, M, and O and J, L, N, and P (Figs. I, K, M, and O depict low magnification). Note the hypocellularity of both thymus and bone marrow that occurs in aged rats, which is accompanied by an increase in adipose tissue. Treatment with either rat GH in the form of GH3 cells or with direct injections of recombinant human GH significantly reverses the age-associated decline in hematopoietic cells. In the bone marrow (Figs. J, L, N, and P), both myeloid progenitors (large arrowhead) and erythroid precursors (small arrowheads) are identified. All arrowheads are placed beneath specific cell types. (Bar, 250 µm for A, C, E, and G; 150 µm for B, D, F, H, I, K, M, and O; 100 µm for J, L, N, and P.)

 
GH reconstitutes hematopoietic cells in the bone marrow of aged rats
The formed elements of bone marrow include erythroid precursors, which are intermingled with myeloid precursors. However, erythrocytic forms can be identified as nests of small, densely staining cells. Myeloid precursors are distinguished as larger cells with euchromatic nuclei and the presence of band forms or nuclear segmentation. Using these criteria, we found that histological sections of bone marrow from control 3-month-old Wistar-Furth rats showed a high degree of hematopoietic cellularity with very little adipose tissue (Fig. 1IGo). These characteristics were more obvious at higher magnification (Fig. 1JGo). Hematopoietic elements in the marrow of these young rats contained erythroid, granulocytic, lymphoid, and megakaryocytic cells, although erythroid and granulocytic elements constituted approximately 95% of all bone marrow hematopoietic cells. In contrast, aged Wistar-Furth rats showed a marked decrease in cellularity with the replacement of hematopoietic cells by lipid-laden adipocytes (Fig. 1Go, K and L). All hematopoietic lineages were represented but with a diminished number of absolute cells. Treatment of aged rats with GH3 cells resulted in a dramatic increase in the number of hematopoietic cells in the bone marrow (Fig. 1Go, M and N), reaching cell numbers that approached those of young control animals (Fig. 1Go, I and J). Bone marrow from these aged, GH3-treated rats displayed all of the hematopoietic elements of the untreated aged control group (arrows denoting myeloid and erythroid precursors in Figs. 1Go, J and L, N and P). Similar to the response to GH3 cells, treatment of aged rats with recombinant GH increased the number of hematopoietic cells in the bone marrow (Fig. 1OGo), which was accompanied by a reduction in the number of adipocytes (Fig. 1PGo).

The absolute number of fat and hematopoietic cells of all lineages in the bone marrow was enumerated in these H&E-stained sections. Bone marrow cells were expressed per square millimeter of bone marrow tissue in young (n = 4), aged (n = 5), and aged GH3-implanted (n = 5) rats. There was an 80% decline (P < 0.01) in bone marrow hematopoietic cells of aged rats, compared with the young animals (Fig. 2AGo). Aged rats supplemented with rat GH in the form of syngeneic GH3 cells exhibited a 4-fold increase (P < 0.01) in the number of hematopoietic cells in bone marrow, resulting in 250 ± 14 cells/mm2. In contrast, there was substantial increase (P < 0.01) in the number of bone marrow adipocytes with aging (Fig. 2BGo). The number of bone marrow adipocytes in aged rats implanted with GH3 cells was not different (P > 0.10) from the number of adipocytes in young rats. In separate experiments, we enumerated the absolute number of bone marrow cells in young (n = 5), aged (n = 5), and aged recombinant GH-treated rats (n = 5). Similar to results of the previous experiment, there was a 58% decline (P < 0.01) in the number of hematopoietic cells of aged, compared with young, rats (Fig. 2CGo). Treatment with recombinant GH doubled (P < 0.01) the number of bone marrow hematopoietic cells in aged rats. However, the number of hematopoietic cells in aged rats given recombinant GH remained less (P < 0.01) than young controls. The number of fat cells in bone marrow increased (Fig. 2DGo; P < 0.01) with age, and this rise was totally reversed by recombinant GH (P < 0.01).



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  		Figure 2. GH-induced reconstitution of bone marrow myeloid cells is accompanied by a reduction in adipocytes. The number of bone marrow hematopoietic cells is lower (A, C; P < 0.01) in aged rats, which is accompanied by an increase (B, D; P < 0.01) in bone marrow adiposity. Treatment of aged rats with either syngeneic GH3 cells (A) or recombinant GH (C) reversed (P < 0.01) the age-associated loss of bone marrow myeloid cells and inhibited the increase in bone marrow adiposity (B, D). The asterisks in all graphs indicate a significant difference (P < 0.01) between aged rats and aged rats treated with either GH3 cells or recombinant GH (n = 4 to 5 rats per treatment, see text).

 
Bone marrow differential counts in hormone-supplemented aged rats
To learn whether restoration of hematopoietic cells in the bone marrow altered the lineage of hematopoietic cells, differential cell counts were determined on H&E-stained tissue from young (n = 9), aged (n = 7), and aged rats implanted with GH3 cells (n = 9). The proportion of erythrocytic, granulocytic, lymphocytic, and megakaryocytic cells in the bone marrow was determined. There was an inversion of the erythroid to granulocyte ratio in young rats, compared with aged rats (Fig. 3AGo). Although there was a significant increase in the number of hematopoietic cells in aged rats implanted with GH3 cells, the proportion of erythrocytic and granulocytic cells remained unchanged, compared with aged control rats. Identical results were observed in independent experiments that examined bone marrow differential counts in young (n = 5), aged (n = 5), and aged rats injected with recombinant GH (n = 5; Fig. 3BGo). In both sets of experiments, lymphocytes and megakaryocytes accounted for less than 5% of the total population. The distribution of bone marrow lymphocytes was unaffected by aging, GH3 cells, or recombinant GH (data not shown). However, the proportion of megakaryocytes was elevated (P < 0.05) in aged rats treated with GH3 cells, compared with either young or aged rats (young = 0.9 ± 0.2%; aged = 1.3 ± 0.3%; aged + GH3 = 3.0 ± 0.7%). This increase in bone marrow megakaryocytes also occurred in aged rats treated with human GH, but the increase was not significant (P > 0.10) (young = 2.2 ± 0.4%; aged = 1.8 ± 0.4%; aged + GH = 3.6 ± 1.1%).



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  		Figure 3. The distribution of hematopoietic cells in bone marrow of aged rats is not affected by treatment with GH3 cells (A) or recombinant GH (B). More than 95% of the morphologically identifiable hematopoietic cells in bone marrow are of either the erythroid or granulocytic lineage. Both aged and hormone-treated aged rats had a lower (P < 0.01) proportion of erythrocytic cells and a higher (P < 0.01) proportion of granulocytic cells. Lymphoid and megakaryocytic cells comprise less than 5% of bone marrow cells, and neither cell population was affected by aging or hormone treatment (data not shown). For each lineage of cells, asterisks indicate that the means are significantly different (P < 0.01) from the proportion of cells in young rats (n = 5 to 9 rats per treatment; see text).

 
Extramedullary hematopoiesis in GH-supplemented aged rats
The remarkable increase in erythroid and myeloid cells in the bone marrow of hormone-treated aged rats suggested that other tissues that support hematopoiesis might also be affected. A typical example of the spleen of an aged rat stained with H&E is shown in Fig. 4AGo. This tissue displayed a sharp, well-delineated splenic margin with a clear demarcation of red and white pulp. There were prominent hemosiderin-laden macrophages in the red pulp, and erythroid elements were detected easily at higher magnification (Fig. 4BGo). In aged rats implanted with GH3 cells, the spleen was markedly enlarged with rounded margins (Fig. 4CGo). The zones between red and white pulp were poorly delineated, and there was a reduction in the number of macrophages with abundant hemosiderin (Fig. 4DGo). Marked EMH consisting of erythroid, granulocytic, and megakaryocytic lineages was observed, which expanded the red pulp (Fig. 4DGo). Aged rats implanted with GH3 cells also displayed EMH in other sinusoidal organs, including the liver (compare aged control [Fig. 4EGo] with GH3-implanted aged rat [Fig. 4FGo]) and adrenal gland (aged control [Fig. 4GGo] vs. GH3-implanted aged rat [Fig. 4HGo]). All blood cell lineages were represented in these sites of EMH. Interestingly, aged rats implanted with GH3 cells displayed a marked metarubricytosis, in which the nucleated red blood cells were characterized by small pyknotic nuclei. This effect was particularly evident in hepatic venous blood in which the nucleated red blood cells displayed small pyknotic nuclei (compare liver of control aged rat [Fig. 4IGo] with liver of GH3-treated aged rat [Fig. 4JGo]). Mild metarubricytosis could be detected in blood, but the complete count of blood cells in control and aged rats implanted with GH3 cells did not show significant differences in either absolute or differential counts (data not shown). The fact that no immature band cells were released into the circulation indicates that the release of blood cells from the marrow and EMH continue to be regulated in aged rats treated with GH.



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  		Figure 4. GH increases the number of EMH cells in the spleen, liver, and adrenal gland. Splenic tissue from control aged rats (A is low magnification and B is higher magnification) shows that treatment of aged rats with GH3 cells leads to splenic enlargement (C is low magnification and D is higher magnification). Similar results occur in the liver ( E and F) and adrenal gland (G and H). Panels E and G represent tissue from a control aged rat, whereas the respective tissue from aged rats treated with GH3 cells is shown in panels F and H. Most of the increase in EMH caused by GH3 cells consists of erythroid cells, but myeloid and megakaryocytic lineages are represented as well (arrowheads in F and H). The release of metarubricytes and reticulocytes in the hepatic sinusoids can be easily detected in aged rats treated with GH3 cells (J, arrowheads) but not in their aged controls (I). Identical results were observed in aged rats treated with recombinant GH. All arrowheads are placed beneath specific cell types. (Bar, 500 µm for A and C; 100 µm for B and D; 150 µm for E, F, G, and H; 75 µm for I and J.)

 
Myeloid colony-forming units (CFUs) in bone marrow and spleen is increased in aged rats implanted with GH3 cells
In young mice, treatment with GH in vivo increases the number of myeloid progenitor cells that can be generated on a constant basis of input cells (11, 28). To determine whether similar results occur in hormone-treated aged rats, we incubated a fixed number (2 x 104) of bone marrow cells from young, aged, and GH3-implanted aged female Wistar-Furth rats (n = 10) with IL-3 and GM-CSF and counted the resulting number of colonies that were generated in vitro in soft agar. There was a reduction (P < 0.05) in the number of myeloid CFUs in aged animals, compared with young controls (Fig. 5AGo). However, the number of CFUs in aged rats implanted with GH3 cells was greater (P < 0.05) than control aged rats and identical (P > 0.10) to the number of CFUs from young rats. Similar experiments were conducted with spleen cells (1 x 105 input cells) to determine whether myeloid progenitors could be detected in GH3-implanted aged rats. Although the number of splenic myeloid CFUs in the spleen was barely detectable in either young or aged rats, spleen cells from aged rats implanted with GH3 cells generated substantially more myeloid CFUs (P < 0.05; Fig. 5BGo). These results are consistent with the morphological finding of EMH in the spleen of hormone-treated aged rats.



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  		Figure 5. The number of myeloid CFUs per cell increases in aged rats following treatment with GH3 cells. Dispersed suspensions of bone marrow (A) or spleen (B) cells were prepared from young, aged rats and aged rats treated with GH3 cells. Cells were incubated in semisolid medium with IL-3 and GM-CSF for 7 d. Aged rats had fewer (P < 0.05) bone marrow-derived myeloid CFUs than aged animals. However, this decline in the number of CFUs per unit of input cells was completely reversed by treatment with GH3 cells (*, P < 0.05). In the spleen, treatment with GH3 cells increased (*, P < 0.05) myeloid CFUs, compared with both young and aged rats (n = 10).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Somatopause (GH/IGF-I), menopause (estrogen), andropause (T), and adrenopause (dehydroepiandrosterone) are physiological consequences of normal aging, and hormone replacement therapies are currently being tested to ameliorate these conditions (1). Replacement therapy with GH in elderly patients is well tolerated and leads to increases in plasma IGF-I, lean body mass, and bone metabolism (39). However, the benefits and risks of hormone replacement for the somatopause of aging remain unknown (40). Here we used standard histological techniques to examine the effect of GH supplementation on the age-associated loss of hematopoietic cells in the bone marrow, a primary hematopoietic tissue. These results demonstrate a remarkable ability of in vivo treatment with GH to replenish hematopoietic cells in the bone marrow (Figs. 1Go and 2Go). The ability of GH to significantly reverse the bone marrow hypocellularity of aging is caused almost entirely by an increase in the number of erythrocytic and granulocytic cells without affecting their relative proportions to one another (Fig. 3Go). When cells were cultured in vitro for assays that adjusted for unequal numbers of cells caused by GH treatment, GH statistically increased the number of myeloid CFUs in both the bone marrow and spleen (Fig. 5Go). Finally, we obtained morphological evidence that administration of GH to aged animals stimulates EMH in other sinusoidal organs, such as the liver and adrenal gland (Fig. 4Go). Collectively, these data establish that the reduction in hematopoietic cells that occurs in the bone marrow of aged animals is a physiological process that is significantly reversed by GH.

A unique aspect of the present report is the use of histological techniques to visualize the effects of in vivo administration of GH on primary hematopoietic tissue. The standard approach is to flush cells from the thymic capsule or bone marrow plug and count the number of output cells that can be maintained as a single-cell suspension. This approach can be an inefficient process that may not accurately reflect the number of hematopoietic cells in situ, particularly in aged animals. Furthermore, this standard technique does not provide an evaluation of the potential effect of hormone therapy on the architecture of the primary hematopoietic tissue, such as the cortex and medulla of the thymus or the erythropoietic and granulocytic zones of hematopoiesis in the bone marrow (Fig. 1Go). This simplistic approach of preparing single-cell suspensions from whole tissue and counting the resulting cells has led us (17) and others ( 26, 41) to conclude that GH does not fully reverse the age-associated decline in hematopoietic cells in primary hematopoietic tissue to the levels found in young animals. However, by specifically counting the number hematopoietic cells in situ as described in this report, we found an impressive recovery of hematopoietic cells in the bone marrow. Although the number of bone marrow hematopoietic cells in GH-treated aged rats remained statistically less (P < 0.05) than young controls (Fig. 2Go), aged rats implanted with either GH3 cells or human GH recovered a full 80% of the hematopoietic cells found in young controls. This is the most complete morphological restoration of hematopoietic cells, based on quantitative morphological criteria, that has yet been reported. More importantly, treatment of aged rats with either form of GH completely reversed the accumulation of adipocytes in the bone marrow, in which the number of adipocytes in the bone marrow of young rats was not different (P > 0.10) from aged rats implanted with GH3 cells or treated with human GH. These quantitative histological data are consistent with the idea that it is possible to simultaneously reverse the loss of hematopoietic cells and the accumulation of adipocytes that occurs in the bone marrow of aged animals.

Treatment of aged rats with GH in vivo did not affect the lineage of cells that developed from progenitors. There was a significant inversion of the erythrocyte to myeloid ratio in aged rats. Even though GH dramatically increased the total number of bone marrow hematopoietic cells in aged rats, the kinds of cells within the total bone marrow population were unaffected. This finding suggests that factors other than GH are responsible for the age-associated inversion of the erythroid/granulocyte ratio. To determine whether GH affected the number of myeloid progenitor cells ex vivo, we prepared single-cell suspensions, incubated equal numbers of these cells for 7 d in IL-3 and GM-CSF and counted the resulting number of CFUs. Even when expressed on the basis of a constant number of input cells, GH increased the number of myeloid progenitor cells in the bone marrow (Fig. 5AGo). These results are consistent with those of others who used in vivo systems (11, 14, 28) and extend this concept to bone marrow tissue of aged rats. Although myeloid progenitor cells are much less plentiful in the spleen, GH increased the number of myeloid CFUs in this tissue. These results are consistent with the potent increase in splenic erythroid and granulocyte-macrophage CFUs that has been reported in transgenic mice that overexpress bovine GH (42) and in GH-deficient adult humans who display an increase in erythropoiesis following long-term replacement therapy with GH (43).

There are many possibilities that could explain the mechanism by which GH stimulates formation of erythroid and granulocytic cells in the bone marrow (44). For example, GH is synthesized by a variety of lymphoid organs (45, 46), and this locally produced GH may act directly on these tissues (47). Endocrine and paracrine GH may also increase IGF-I that is produced by cells in both the thymus (48) and bone marrow ( 34). Alternatively, GH may act indirectly, such as by reducing lipogenesis throughout all animal tissues (49), including the bone marrow. However, this effect is likely to be complicated because adipocytes produce proteins that both enhance (e.g. leptin [50 ]) and suppress (e.g. TNF-{alpha} [51 ]) hematopoiesis. We favor the view that IGF-I is the critical molecule that acts directly on progenitor cells to promote hematopoiesis. For example, the enhancing effects of GH on both human erythropoiesis and granulopoiesis in the bone marrow are blocked in vitro by an antibody to the IGF-I receptor (52). Addition of antibodies to the IGF-I receptor block differentiation of double-positive thymocytes from their double-negative precursors (48). Progenitor cells of the B, T, and myeloid lineages express receptors for IGF-I (31). We have not detected GH receptors or biological responses of promyeloid cells to GH in vitro. However, IGF-I receptors are abundant on promyeloid cells (53), and, once activated in vitro, IGF-I promotes their replication (53, 54), inhibits their death ( 55, 56), and promotes their differentiation into both granulocytes (55) and macrophages (57).

Although the thymus gland involutes substantially during the aging process, the total number of peripheral T cells does not decline (58). Instead, there is a change in the subpopulations of peripheral T cells, which is characterized by a loss of naïve cells and an increase in memory cells. It is now recognized that the production of naïve T cells from the human thymus continues even in very old age, albeit at a reduced output (59, 60). IL-7 has recently been shown to reverse the reduction in thymopoiesis that occurs in aged mice (61), so GH/IGF-I may act by increasing the synthesis of this cytokine. Although involution of the thymus has been hypothesized to simply represent good housekeeping (62), the major objective of many past and current experiments is to increase functional capabilities of the thymus during old age. For example, a novel GH secretagogue has recently been shown to increase the generation of cytotoxic T lymphocytes, augment resistance to the establishment of tumors, and inhibit subsequent metastases in aged mice (63). Similarly, we have shown that GH/IGF-I primes human, porcine, and bovine neutrophils (64, 65) and porcine and rat macrophages ( 66, 67) for enhanced secretion of free oxygen radicals. This leads to enhanced bacterial killing by neutrophils from aged rats treated in vivo with GH (68). Similarly, treatment of GH-deficient adult humans with GH increases blood hemoglobin concentrations (43), consistent with findings of enhanced erythropoiesis in the present study. Although GH treatment for the critically ill has been shown to increase mortality (69), it continues to be recommended for patients with GH deficiency. Indeed, long-term, low-dose GH treatment has been shown to significantly increase life expectancy of aged mice (70).

The thymotropic properties of GH are well documented (71), even in aged animals. The ability of GH treatment in vivo to stimulate proliferation of hematopoietic cells in the bone marrow compartment of young animals was shown to occur in the original studies of Nagy and Berczi (27). Here we extend these and other reports to hematopoietic tissue of aged animals by showing that in vivo treatment with GH significantly reverses bone marrow hypocellularity. These data establish that the loss of hematopoietic cells and the accumulation of adipocytes in the bone marrow during aging are not irreversible events and that manipulation of the endocrine system is one way to retard these age-associated changes.


   Acknowledgments
 
The authors gratefully acknowledge the technical expertise of Wenhong Shen in the statistical evaluation of these data.


   Footnotes
 
This work was supported by grants (to K.W.K.) from the NIH (AG-06246, MH-51569) and the Pioneering Research Project in Biotechnology financed by the Japanese Ministry of Agriculture, Forestry, and Fisheries.

1 Present address: Department of Pathobiology and Veterinary Science, University of Connecticut, Storrs, Connecticut 06269-3089. Back

2 Present address: Department of Life Sciences, University of Limerick, Limerick, Ireland. Back

Abbreviations: CFU, Colony-forming unit; EMH, extramedullary hematopoiesis; GM-CSF, granulocyte-macrophage colony stimulating factor; H&E, hematoxylin and eosin.

Received July 6, 2001.

Accepted for publication October 5, 2001.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Lamberts SW, van den Beld AW, van der Lely AJ 1997 The endocrinology of aging. Science 278:419–424[Abstract/Free Full Text]
  2. Miller RA 1996 The aging immune system: primer and prospectus. Science 273:70–74[Abstract]
  3. Mackall CL, Fleisher TA, Brown MR, Andrich MP, Chen CC, Feuerstein IM, Horowitz ME, Magrath IT, Shad AT, Steinberg SM, et al. 1995 Age, thymopoiesis, and CD4+ T-lymphocyte regeneration after intensive chemotherapy. N Engl J Med 332:143–149[Abstract/Free Full Text]
  4. Goya RG, Gagnerault MC, De Moraes MC, Savino W, Dardenne M 1992 In vivo effects of growth hormone on thymus function in aging mice. Brain Behav Immun 6:341–354[CrossRef][Medline]
  5. Kelley KW, Brief S, Westly HJ, Novakofski J, Bechtel PJ, Simon J, Walker EB 1986 GH3 pituitary adenoma cells can reverse thymic aging in rats. Proc Natl Acad Sci USA 83:5663–5667[Abstract/Free Full Text]
  6. Knyszynski A, Adler-Kunin S, Globerson A 1992 Effects of growth hormone on thymocyte development from progenitor cells in the bone marrow. Brain Behav Immun 6:327–340[CrossRef][Medline]
  7. Goff BL, Roth JA, Arp LH, Incefy GS 1987 Growth hormone treatment stimulates thymulin production in aged dogs. Clin Exp Immunol 68:580–587[Medline]
  8. Murphy WJ, Durum SK, Longo DL 1992 Human growth hormone promotes engraftment of murine or human T cells in severe combined immunodeficient mice. Proc Natl Acad Sci USA 89:4481–4485[Abstract/Free Full Text]
  9. Murphy WJ, Durum SK, Anver M, Frazier M, Longo DL 1992 Recombinant human growth hormone promotes human lymphocyte engraftment in immunodeficient mice and results in an increased incidence of human Epstein Barr virus-induced B-cell lymphoma. Brain Behav Immun 6:355–364[CrossRef][Medline]
  10. Murphy WJ, Durum SK, Longo DL 1992 Role of neuroendocrine hormones in murine T cell development. Growth hormone exerts thymopoietic effects in vivo. J Immunol 149:3851–3857[Abstract]
  11. Murphy WJ, Durum SK, Anver MR, Longo DL 1992 Immunologic and hematologic effects of neuroendocrine hormones. Studies on DW/J dwarf mice. J Immunol 148:3799–3805[Abstract]
  12. Murphy WJ, Durum SK, Longo DL 1993 Differential effects of growth hormone and prolactin on murine T cell development and function. J Exp Med 178:231–236[Abstract/Free Full Text]
  13. Berczi I, Nagy E, de Toledo SM, Matusik RJ, Friesen HG 1991 Pituitary hormones regulate c-myc and DNA synthesis in lymphoid tissue. J Immunol 146:2201–2206[Abstract]
  14. Murphy WJ, Tsarfaty G, Longo DL 1992 Growth hormone exerts hematopoietic growth-promoting effects in vivo and partially counteracts the myelosuppressive effects of azidothymidine. Blood 80:1443–1447[Abstract/Free Full Text]
  15. Beschorner WE, Divic J, Pulido H, Yao X, Kenworthy P, Bruce G 1991 Enhancement of thymic recovery after cyclosporine by recombinant human growth hormone and insulin-like growth factor I. Transplantation 52:879–884[Medline]
  16. Davila DR, Brief S, Simon J, Hammer RE, Brinster RL, Kelley KW 1987 Role of growth hormone in regulating T-dependent immune events in aged, nude, and transgenic rodents. J Neurosci Res 18:108–116[CrossRef][Medline]
  17. Li YM, Brunke DL, Dantzer R, Kelley KW 1992 Pituitary epithelial cell implants reverse the accumulation of CD4-CD8- lymphocytes in thymus glands of aged rats. Endocrinology 130:2703–2709[Abstract/Free Full Text]
  18. Dubiski S, Ponnappan U, Cinader B 1989 Strain polymorphism in progression of aging: changes in CD4, CD8 bearing subpopulations. Immunol Lett 23:1–7[CrossRef][Medline]
  19. Gagnerault MC, Postel-Vinay MC, Dardenne M 1996 Expression of growth hormone receptors in murine lymphoid cells analyzed by flow cytofluorometry. Endocrinology 137:1719–1726[Abstract]
  20. Dardenne M, Mello-Coelho V, Gagnerault MC, Postel-Vinay MC 1998 Growth hormone receptors and immunocompetent cells. Ann NY Acad Sci 840:510–517[CrossRef][Medline]
  21. Kooijman R, Scholtens LE, Rijkers GT, Zegers BJ 1995 Type I insulin-like growth factor receptor expression in different developmental stages of human thymocytes. J Endocrinol 147:203–209[Abstract/Free Full Text]
  22. Ban E, Gagnerault MC, Jammes H, Postel-Vinay MC, Haour F, Dardenne M 1991 Specific binding sites for growth hormone in cultured mouse thymic epithelial cells. Life Sci 48:2141–2148[CrossRef][Medline]
  23. Timsit J, Savino W, Safieh B, Chanson P, Gagnerault MC, Bach JF, Dardenne M 1992 Growth hormone and insulin-like growth factor-I stimulate hormonal function and proliferation of thymic epithelial cells. J Clin Endocrinol Metab 75:183–188[Abstract]
  24. Eren R, Globerson A, Abel L, Zharhary D 1990 Quantitative analysis of bone marrow thymic progenitors in young and aged mice. Cell Immunol 127:238–246[CrossRef][Medline]
  25. McCormick KR, Haar JL 1991 Bone marrow-thymus axis in senescence. Am J Anat 191:321–324[CrossRef][Medline]
  26. Montecino-Rodriguez E, Clark R, Dorshkind K 1998 Effects of insulin-like growth factor administration and bone marrow transplantation on thymopoiesis in aged mice. Endocrinology 139:4120–4126[Abstract/Free Full Text]
  27. Nagy E, Berczi I 1989 Pituitary dependence of bone marrow function. Br J Haematol 71:457–462[Medline]
  28. Tian ZG, Woody MW, Sun R, Welniak LA, Raziuddin A, Funakoshi S, Tsarfaty G, Longe DL, Murphy WJ 1998 Recombinant human growth hormone promotes hematopoietic reconstitution after syngeneic bone marrow transplantation in mice. Stem Cells 16:193–199[Abstract/Free Full Text]
  29. Welniak LA, Tian ZG, Sun R, Keller JR, Richards S, Ruscetti FW, Murphy WJ 2000 Effects of growth hormone and prolactin on hematopoiesis. Leuk Lymphoma 38:435–445[Medline]
  30. Woody MA, Welniak LA, Richards S, Taub DD, Tian Z, Sun R, Longo DL, Murphy WJ 1999 Use of neuroendocrine hormones to promote reconstitution after bone marrow transplantation. Neuroimmunomodulation 6:69–80[CrossRef][Medline]
  31. Burgess W, Liu Q, Zhou J, Tang Q, Ozawa A, VanHoy R, Arkins S, Dantzer R, Kelley KW 1999 The immune-endocrine loop during aging: role of growth hormone and insulin-like growth factor-I. Neuroimmunomodulation 6:56–68[CrossRef][Medline]
  32. Davila DR, Edwards CKI, Arkins S, Simon J, Kelley KW 1990 Interferon-gamma-induced priming for secretion of superoxide anion and tumor necrosis factor-{alpha} declines in macrophages from aged rats. FASEB J 4:2906–2911[Abstract]
  33. Moloney WC, Boschetti AE, King V 1969 Observations on leukemia in Wistar Furth rats. Cancer Res 29:938–946[Abstract/Free Full Text]
  34. Arkins S, Rebeiz N, Brunke-Reese DL, Minshall C, Kelley KW 1995 The colony-stimulating factors induce expression of insulin-like growth factor-I messenger ribonucleic acid during hematopoiesis. Endocrinology 136:1153–1160[Abstract]
  35. French RA, Zachary JF, Dantzer R, Frawley LS, Chizzonite R, Parnet P, Kelley KW 1996 Dual expression of p80 type I and p68 type II interleukin-I receptors on anterior pituitary cells synthesizing growth hormone. Endocrinology 137:4027–4036[Abstract]
  36. French RA, VanHoy RW, Chizzonite R, Zachary JF, Dantzer R, Parnet P, Bluthé RM, Kelley KW 1999 Expression and localization of p80 and p68 interleukin-1 receptor proteins in the brain of adult mice. J Neuroimmunol 93:194–202[CrossRef][Medline]
  37. Heyworth C, Spooncer E 1993 In vitro clonal assays for murine multi-potential and lineage restricted myeloid progenitor cells. In: Testa NG, Molineux G, eds. Hematopoiesis: a practical approach, Oxford: Oxford University Press; 37–53
  38. SAS 2000 SAS/STAT Release 8.1. Cary, NC: SAS Institute
  39. Fernholm R, Bramnert M, Hagg E, Hilding A, Baylink DJ, Mohan S, Thoren M 2000 Growth hormone replacement therapy improves body composition and increases bone metabolism in elderly patients with pituitary disease. J Clin Endocrinol Metab 85:4104–4112[Abstract/Free Full Text]
  40. Lamberts SW 2000 The somatopause: to treat or not to treat? Horm Res 53:42–43
  41. Dorshkind K, Horseman ND 2000 The roles of prolactin, growth hormone, insulin-like growth factor-I, and thyroid hormones in lymphocyte development and function: insights from genetic models of hormone and hormone receptor deficiency. Endocr Rev 21:292–312[Abstract/Free Full Text]
  42. Blazar BR, Brennan CA, Broxmeyer HE, Shultz LD, Vallera DA 1995 Transgenic mice expressing either bovine growth hormone (bGH) or human GH releasing hormone (hGRH) have increased splenic progenitor cell colony formation and DNA synthesis in vitro and in vivo. Exp Hematol 23:1397–1406[Medline]
  43. Ten Have SM, van der Lely AJ, Lamberts SW 1997 Increase in haemoglobin concentrations in growth hormone deficient adults during human recombinant growth hormone replacement therapy. Clin Endocrinol (Oxf) 47:565–570[CrossRef][Medline]
  44. Kooijman R, Hooghe-Peters EL, Hooghe R 1996 Prolactin, growth hormone, and insulin-like growth factor-I in the immune system. Adv Immunol 63:377–454[Medline]
  45. Vines CR, Weigent DA 2000 Identification of SP3 as a negative regulatory transcription factor in the monocyte expression of growth hormone. Endocrinology 141:938–946[Abstract/Free Full Text]
  46. Recher S, Raccurt M, Lambert A, Lobie PE, Mertani HC, Morel G 2001 Prenatal and adult growth hormone gene expression in rat lymphoid organs. J Histochem Cytochem 49:347–354[Abstract/Free Full Text]
  47. Weigent DA, Blalock JE 1997 Production of peptide hormones and neurotransmitters by the immune system. Chem Immunol 69:1–30[Medline]
  48. Kecha O, Brilot F, Martens H, Franchimont N, Renard C, Greimers R, Defresne MP, Winkler R, Geenen V 2000 Involvement of insulin-like growth factors in early T cell development: a study using fetal thymic organ cultures. Endocrinology 141:1209–1217[Abstract/Free Full Text]
  49. Etherton TD 2000 The biology of somatotropin in adipose tissue growth and nutrient partitioning. J Nutr 130:2623–2625[Abstract/Free Full Text]
  50. Fantuzzi G, Faggioni R 2000 Leptin in the regulation of immunity, inflammation, and hematopoiesis. J Leukoc Biol 68:437–446[Abstract/Free Full Text]
  51. Sachs L 1996 The control of hematopoiesis and leukemia: from basic biology to the clinic. Proc Natl Acad Sci USA 93:4742–4749[Abstract/Free Full Text]
  52. Merchav S 1998 The haematopoietic effects of growth hormone and insulin-like growth factor-I. J Pediatr Endocrinol Metab 11:677–685[Medline]
  53. Minshall C, Arkins S, Freund GG, Kelley KW 1996 Requirement for phosphatidylinositol 3'-kinase to protect hemopoietic progenitors against apoptosis depends upon the extracellular survival factor. J Immunol 156:939–947[Abstract]
  54. Liu Q, VanHoy RW, Zhou JH, Dantzer R, Freund GG, Kelley KW 1999 Elevated cyclin E levels, inactive retinoblastoma protein, and suppression of the p27KIP1 inhibitor characterize early development of promyeloid cells into macrophages. Mol Cell Biol 19:6229–6239[Abstract/Free Full Text]
  55. Liu Q, Schacher D, Hurth C, Freund GG, Dantzer R, Kelley KW 1997 Activation of phosphatidylinositol 3'-kinase by insulin-like growth factor-I rescues promyeloid cells from apoptosis and permits their differentiation into granulocytes. J Immunol 159:829–837[Abstract]
  56. Minshall C, Arkins S, Dantzer R, Freund GG, Kelley KW 1999 Phosphatidylinositol 3'-kinase, but not S6-kinase, is required for insulin-like growth factor-I and IL-4 to maintain expression of Bcl-2 and promote survival of myeloid progenitors. J Immunol 162:4542–4549[Abstract/Free Full Text]
  57. Liu Q, Ning W, Dantzer R, Freund GG, Kelley KW 1998 Activation of protein kinase C-{zeta} and phosphatidylinositol 3'-kinase and promotion of macrophage differentiation by insulin-like growth factor-I. J Immunol 160:1393–1401[Abstract/Free Full Text]
  58. Aspinall R, Andrew D 2000 Thymic involution in aging. J Clin Immunol 20:250–256[CrossRef][Medline]
  59. Haynes BF, Sempowski GD, Wells AF, Hale LP 2001 The human thymus during aging. Immunol Res 22:253–261
  60. Haynes BF, Markert ML, Sempowski GD, Patel DD, Hale LP 2000 The role of the thymus in immune reconstitution in aging, bone marrow transplantation, and HIV-1 infection. Annu Rev Immunol 18:529–560[CrossRef][Medline]
  61. Andrew D, Aspinall R 2001 IL-7 and not stem cell factor reverses both the increase in apoptosis and the decline in thymopoiesis seen in aged mice. J Immunol 166:1524–1530[Abstract/Free Full Text]
  62. George AJ, Ritter MA 1996 Thymic involution with ageing: obsolescence or good housekeeping? Immunol Today 17:267–272[CrossRef][Medline]
  63. Koo GC, Huang C, Camacho R, Trainor C, Blake JT, Sirotina-Meisher A, Schleim KD, Wu TJ, Cheng K, Nargund R, McKissick G 2001 Immune enhancing effect of a growth hormone secretagogue. J Immunol 166:4195–4201[Abstract/Free Full Text]
  64. Fu YK, Arkins S, Wang BS, Kelley KW 1991 A novel role of growth hormone and insulin-like growth factor-I. Priming neutrophils for superoxide anion secretion. J Immunol 146:1602–1608[Abstract]
  65. Fu YK, Arkins S, Fuh G, Cunningham BC, Wells JA, Fong S, Cronin MJ, Dantzer R, Kelley KW 1992 Growth hormone augments superoxide anion secretion of human neutrophils by binding to the prolactin receptor. J Clin Invest 89:451–457
  66. Edwards CKI, Ghiasuddin SM, Schepper JM, Yunger LM, Kelley KW 1988 A newly defined property of somatotropin: priming of macrophages for production of superoxide anion. Science 239:769–771[Abstract/Free Full Text]
  67. Edwards CKI, Yunger LM, Lorence RM, Dantzer R, Kelley KW 1991 The pituitary gland is required for protection against lethal effects of Salmonella typhimurium. Proc Natl Acad Sci USA 88:2274–2277[Abstract/Free Full Text]
  68. Fu YK, Arkins S, Li YM, Dantzer R, Kelley KW 1994 Reduction in superoxide anion secretion and bactericidal activity of neutrophils from aged rats: reversal by the combination of gamma interferon and growth hormone. Infect Immun 62:1–8[Abstract/Free Full Text]
  69. Gelato MC 2000 The growth hormone/insulin-like growth factor axis in critical illness. J Pediatr Endocrinol Metab 13(Suppl 2):1023–1029
  70. Khansari DN, Gustad T 1991 Effects of long-term, low-dose growth hormone therapy on immune function and life expectancy of mice. Mech Ageing Dev 57:87–100[CrossRef][Medline]
  71. Murphy WJ, Longo DL 2000 Growth hormone as an immunomodulating therapeutic agent. Immunol Today 21:211–213[CrossRef][Medline]



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