Biomedical subjects

S B Krantz

Publications and source records attributed to S B Krantz.

At least 37 records · Page 2Linked to original sources

PubMed1995

Inhibition of heme synthesis induces apoptosis in human erythroid progenitor cells.

Heme synthesis by erythroid progenitor cells is maintained by erythropoietin (EP), insulin-like growth factor-I (IGF-I), and stem cell factor (SCF), and without these growth factors apoptosis (programmed cell death) occurs. To clarify the possible interaction between heme synthesis and programmed cell death of human erythroid progenitor cells, the effect of specific inhibition of heme synthesis on apoptosis of highly purified human erythroid colony forming cells (ECFC) was studied. When the amount of uncleaved DNA was determined as a measure of apoptosis, the heme synthesis inhibitors, succinylacetone (SA) (0.1 mmol/L) or isonicotinic acid hydrazide (INH) (10 mmol/L), significantly decreased the amount of uncleaved DNA (P < 0.01) in the presence of erythropoietin (EP). Addition of recombinant heavy-chain ferritin (rHF) (10 nmol/L), or deprivation of transferrin from the culture medium, which decreased heme synthesis, also reduced the amount of uncleaved DNA (P < 0.01). The production of apoptosis by diverse inhibitors of heme synthesis was in each case reversed by the addition of hemin (0.1 mmol/L) and did not occur with HL-60 cells. When the colony-forming capacity of ECFC was determined by plasma clot assay, SA, INH, or rHF reduced the number of CFU-E (P < 0.01), and the effect of SA was reversed by hemin. The addition of SA did not alter the c-myc response of ECFC to EP. These data indicate that inhibition of heme synthesis induces apoptosis of human erythroid progenitor cells, in a manner independent of an early c-myc response, and suggest that the presence of apoptosis in ineffective erythropoiesis may be secondary to impaired heme synthesis.

Apoptosis↗

PubMed1995

Stem cell factor can overcome inhibition of highly purified human burst-forming units-erythroid by interferon gamma.

Highly purified human blood burst-forming units-erythroid (BFU-E) were used to study the effects of interferon gamma (IFN gamma). IFN gamma inhibited erythroid colony formation, cell proliferation, and differentiation of day 3 to day 6 mature BFU-E in a dose-dependent manner. The primitive BFU-E (day 1 and day 2 cells) and later day 7 cells were less affected. IFN gamma dose-response experiments demonstrated that the number and size of erythroid colonies were reduced at a concentration of 500 U/ml with more complete inhibition at 1,000 U/ml. Inhibition of day 4 to day 6 erythroid progenitors was first noted by 72 h of incubation with IFN gamma, and target cell growth and differentiation continued to decrease with further incubation. IFN gamma also induced erythroblast apoptosis which was demonstrated by both nuclear condensation and fragmentation plus flow cytometry with in situ end-labelling. Because day 3 to day 6 cells need stem cell factor (SCF) for development in serum-free culture, the relationship of IFN gamma inhibition to this growth factor was investigated. The reduction in the number of erythroid colonies by IFN gamma was reversed by SCF although the colony size was not completely re-established. In contrast, interleukin-3 did not have the capacity to overcome the inhibitory effects of IFN gamma. Since IFN gamma blood levels are elevated in some anemias of chronic disease, IFN gamma may have a role in promoting this anemia and its inhibitory effect might be better overcome by SCF plus EP. However, the mechanism by which these growth factors overcome the inhibition of IFN gamma, or vice versa, is unknown at the present time.

Apoptosis↗

PubMed1995

Erythropoietin and the anaemia of chronic disease.

The anaemia of chronic disease is the second most common anaemia in the world and is an underproduction anaemia with relatively low erythropoietin (EPO) values for the degree of the anaemia. This anaemia occurs with inflammation, infection, or malignancy and a principle question has been whether it would respond to recombinant human EPO (r-HuEPO). Several studies are now available to answer this question. In one study 12 of 16 patients with rheumatoid arthritis receiving r-HuEPO increased their haematocrits 6 percentage points or more and 11 of 12 reached normal haematocrits. Investigations of the effect of r-HuEPO on the anaemia of AIDS showed that patients with EPO levels of 500 U/L or less had an increase in the mean haematocrit of 4.6 percentage points with a decrease in red cell transfusions from 5.3 to 3.2 units per patient. Quality of life indices significantly improved in responders. When 413 patients with anaemia due to a wide variety of malignancies were randomized to r-HuEPO treatment, 58% of those receiving chemotherapy increased their haematocrits by at least 6 points over 12 weeks. Quality of life parameters of responders also significantly improved. Anaemia in three patients with inflammatory bowel disease also responded in 8-14 weeks to r-HuEPO and two of the three reached normal haemoglobin levels. It is clear that r-HuEPO can correct the anaemia of chronic disease and can improve the quality of life of responders.

Acquired Immunodeficiency Syndrome↗

PubMed1994

Erythroid-specific processing of human beta spectrin I pre-mRNA.

Erythroid cells express a unique form of beta spectrin I as a result of tissue-specific alternative pre-mRNA processing. Nonerythroid cells that express the beta spectrin I gene include four additional exons at the 3' end of the mature transcript, leading to elongation of the carboxyl terminus of the protein. The nonerythroid beta spectrin I isoform is not present in the red blood cell membrane skeleton; the erythroid isoform is not detected in other cell types. Therefore, developing erythroid cells acquire this tissue-specific pre-mRNA processing activity during differentiation. In the present study, we investigated the developmental timing of erythroid-specific pre-mRNA processing in human erythroid precursors. Partially purified human peripheral blood burst forming uniterythroid (BFU-E) cells were grown in culture for 5 to 12 days. beta Spectrin I mRNA transcripts were analyzed at different time points by S1 nuclease mapping. The processing of beta spectrin I transcripts was found to be exclusively erythroid from day 5 onward, indicating that erythroid-specific processing is not linked temporally to assembly of the mature erythroid membrane skeleton. Human erythroleukemia (HEL) cells had both erythroid and nonerythroid transcripts, indicating that both processing patterns can coexist. Induction of erythroid differentiation in HEL cells using hemin resulted in a partial switch toward the erythroid processing pattern of beta spectrin I transcripts. Using a genomic S1 probe that spans the erythroid polyadenylation signal, we found that a substantial portion of the transcripts detected by the erythroid cDNA S1 probe (in both cultured BFU-E and HEL cells) is incompletely processed pre-mRNA precursors. Poly(A) RNA selection before S1 analysis showed that the unprocessed transcripts are not polyadenylated. We conclude that (1) erythroid-specific pre-mRNA processing activity is present early in erythroid differentiation; (2) beta spectrin I transcripts that are unprocessed at the 3' end accumulate, awaiting either erythroid or nonerythroid processing pathways, from which observation we infer that the regulated alternative pathways are both inefficient; and (3) HEL cells offer a human cell culture model in which to study the balance between the two pre-mRNA processing pathways. We speculate that erythroid cells evolved this tissue-specific pre-mRNA processing machinery for other erythroid genes in addition to beta spectrin I.

Base Sequence↗

PubMed1994

Inhibition of murine erythroid colony formation in vitro by interferon gamma and correction by interferon receptor immunoadhesin.

It has been previously reported that inhibition of human erythroid colony-forming units (CFU-E) in vitro by interleukin-1 (IL-1) is an indirect effect, occurring through the production of interferon gamma (IFN gamma). IFN gamma, in turn, inhibits CFU-E colony formation directly, and its inhibitory effect can be overcome by exposure to high concentrations of erythropoietin (EPO). To develop an in vitro animal model for investigating inhibition of erythropoiesis by IFN gamma, the effects of recombinant murine (rm) IFN gamma on highly purified CFU-E from the spleens of mice infected with the anemia strain of the Friend virus (FVA) were studied. rmIFN gamma inhibited CFU-E colony formation in a dose-dependent manner. This inhibition occurred with large (> or = 8 cell) colonies only; smaller colonies were not affected. The inhibitory effect was corrected to 72% of control by high EPO concentrations of 64 U/mL. Murine CFU-E were then cultured with rmIFN gamma in the presence of a soluble murine IFN gamma receptor fused to the hinge and Fc domains of the human IgG1 heavy chain (mIFN gamma R-IgG). Inhibition of CFU-E colony formation by rmIFN gamma (100 U/mL) was corrected by mIFN gamma R-IgG in a dose-dependent manner, with an approximate IC50 of 0.05 nmol/L, and complete or near complete correction at 0.5 nmol/L. Similarly, a human IFN gamma R-IgG greatly reduced the inhibitory effect of recombinant human IFN gamma on human CFU-E. These experiments provide an in vitro animal model for studying the inhibitory effects of IFN gamma on erythropoiesis and indicate that IFN gamma R-IgG may be a useful agent for reducing the toxicity of IFN gamma in vivo.

Animals↗

PubMed1994

Changes in cytoskeletal proteins and their mRNAs during maturation of human erythroid progenitor cells.

We have used highly purified human early erythroid progenitors to study changes in cytoskeletal proteins during their maturation and terminal differentiation. When erythroid progenitors at the burst-forming unit-erythroid (BFU-E) stage of development are grown in the presence of erythropoietin, the cells mature and terminally differentiate into reticulocytes during a 14-15-day culture period. We have shown by immunofluorescence that spectrin is present in day 3 BFU-E, at which time proteins band 3, ankyrin, and band 4.1 cannot be detected. Ankyrin and band 4.1 were detected in the majority of the cells by day 7 of culture, at the colony-forming unit (CFU)-E stage, whereas only 15% of the cells were positive for band 3 protein on day 7 of culture. The mRNA level for spectrin was already at its maximum on day 8 whereas the mRNAs for band 3, ankyrin, and band 4.1 were just beginning to accumulate. After enucleation, spectrin, band 3, ankyrin, and band 4.1 fluorescence were all associated with the reticulocytes. Actin was localized at the constriction between the extruding nucleus and the incipient reticulocyte in enucleating erythroblasts suggesting a key role for actin in the enucleation of human erythroblasts. Our investigations have also shown that purified human erythroid progenitors cultured in serum-free suspension media are capable of enucleating without the requirement of an extracellular matrix. These results demonstrate that the synthesis and expression of major cytoskeletal proteins in the human erythrocyte membrane occur in an asynchronous manner and that the remodeling of the membrane skeleton begins at a very early stage during erythrocyte development.

Actins↗

PubMed1994

The anemia of microgravity and recumbency: role of sympathetic neural control of erythropoietin production.

We hypothesize that reduced sympathetic stimulation of erythropoietin production may maintain the anemia which develops in virtually all space travellers. We tested this hypothesis in a human model of reduced sympathetic activity. Thirty-three patients with the Bradbury-Eggleston syndrome were divided into three groups according to their hemoglobin (Hgb) level. Patients with low Hgb had lower upright norepinephrine and lower upright renin. Patients with anemia also had inappropriately low plasma erythropoietin levels. We administered recombinant erythropoietin (Epogen) 25-50 units/kg s.c. 3 times per week and found that the anemia seen in autonomic failure could be reversed by this treatment. These results support the hypothesis that erythropoiesis is modulated by the sympathetic nervous system and that at such mechanisms may also operate in the microgravity environment where sympathetic activity is reduced.

Anemia↗

PubMed1994

Pathogenesis and treatment of the anemia of chronic disease.

The anemia of chronic disease may be viewed simply as the anemia that accompanies chronic inflammatory, infectious, or neoplastic disorders. Because these conditions are very common, the anemia of chronic disease is one of the most frequent anemias encountered, and is only second in incidence to iron-deficiency anemia. The anemia of chronic disease is primarily an anemia due to underproduction of red cells, with low reticulocyte production, and is most often a normochromic, normocytic anemia. However, in 30% to 50% of patients, the red cells are hypochromic and microcytic and, most often, the serum iron, total iron-binding capacity, and transferrin saturation are reduced in the presence of adequate iron stores. Although the differential diagnosis includes other underproduction anemias, such as those caused by vitamin and mineral deficiencies, renal failure, endocrinopathies, and myelodysplasia, it generally is easily distinguished from these conditions. Nevertheless, an understanding of the pathogenesis of this condition, as well as a means of alleviating the anemia when the chronic disorder persists, has remained elusive. Recently, major advances have occurred toward understanding the pathogenesis of the anemia of chronic disease and its treatment, and these advances are reviewed.

Acquired Immunodeficiency Syndrome↗

PubMed1994

Treatment of aplastic anemia with an investigational antilymphocyte serum prepared in rabbits.

The authors evaluated antilymphocyte serum prepared in rabbits (ALS-R) as an alternative to antilymphocyte serum prepared in horses (ALG-H) in the therapy of aplastic anemia. Between 1980 and 1993, 57 evaluable patients received ALS-R and prednisone +/- cyclosporine +/- androgens. Standard response criteria were used and patients were evaluated at 3 months from the start of therapy. Median age was 43 years. Disease was present for up to 2 months in 24 patients, 2-5 months in 14 patients, and 6 months or more in 19 patients. Disease was severe in 30 patients and moderate in 27. Responses occurred in 16 (28%) of 57 patients. Responses were more frequent in females, in patients treated within 6 months of diagnosis, and in patients with severe disease. Among patients receiving ALS-R and cyclosporine within 2 months of diagnosis, 46% responded. After ALS-R therapy, 20 patients received ALG-H; 8 (40%) of 20 responded. Eight patients receiving ALS-R previously had received ALG-H; 2 (25%) of these 8 patients responded. Toxicity of ALS-R was minimal. Antilymphocyte serum prepared in rabbits, in conjunction with other immunosuppressive agents, represents an effective alternative to ALG-H in aplastic anemia, especially in patients previously treated with ALG-H.

Adolescent↗

PubMed1994

Polycythaemia vera. III. Burst-forming units-erythroid (BFU-E) response to stem cell factor and c-kit receptor expression.

We previously demonstrated that highly purified normal human blood burst-forming units-erythroid (BFU-E) need the direct action of recombinant human stem cell factor (rSCF) in the presence of recombinant human erythropoietin (rEP) and recombinant human interleukin-3 (rIL-3) for further development in a serum-free medium. To study the response of polycythaemia vera (PV) BFU-E to rSCF, we performed dose-response experiments in a serum-free medium using highly purified BFU-E from PV patients. A marked increase in the number of PV bursts occurred with increasing concentrations of rSCF, compared to normal burst formation, when the cells were cultured in the presence of rIL-3 at 1 U/ml. The percentage of maximum growth for normal BFU-E was 31 +/- 11% while for PV it was 64 +/- 9% at the highest concentration of rSCF (P < 0.01). Without rIL-3, only 11% of maximum normal BFU-E growth occurred as the rSCF concentration was increased and the size of the colonies was very small, but PV BFU-E still expressed 48% of the maximum number of large erythroid bursts (P < 0.001). This demonstrated an enhanced sensitivity of PV BFU-E to rSCF, compared to normal BFU-E. The pattern of 59Fe incorporation into haem after 8 d of cell culture indicated that PV BFU-E had a time course of maturation and a degree of cellular maturity similar to normal BFU-E. The percentage positivity and intensity of c-kit receptors on PV erythroid cells were examined using immunofluorescence flow cytometry. When BFU-E, CFU-E, or erythroblasts were incubated with phycoerythrin-conjugated SR-1 anti-c-kit receptor monoclonal antibody, 90% of the PV and normal BFU-E displayed c-kit receptor at comparable intensities, as well as 80% of the PV and normal CFU-E. A distinct loss of c-kit expression occurred with erythroid differentiation beyond the CFU-E stage, but at all stages no difference of c-kit receptor expression was evident for PV erythroid precursors compared to normal precursors. These results indicate that the hypersensitivity to rSCF did not appear to be related to the number of c-kit receptors. Since we have previously shown that highly purified PV BFU-E are hypersensitive to rIL-3 and rGM-CSF, as well as rEP, it is now evident that PV BFU-E are hypersensitive to each of the cytokines that have a prominent role in guiding their normal proliferation and differentiation.(ABSTRACT TRUNCATED AT 400 WORDS)

Cells, Cultured↗

PubMed1994

Polycythaemia vera. IV. Specific binding of stem cell factor to normal and polycythaemia vera highly purified erythroid progenitor cells.

Polycythaemia vera (PV) patients' blood burst-forming units-erythroid (BFU-E) have an enhanced sensitivity to stem cell factor (SCF) compared to normal BFU-E. To characterize SCF receptors on erythroid progenitors from normal individuals and PV patients, we performed binding experiments using radioiodinated recombinant SCF (rSCF), day 1 BFU-E and day 8 erythroid colony-forming cells (ECFC), which are mostly colony-forming units-erythroid (CFU-E). 125I-rSCF binds to a single class of cell surface receptors (23,000/ECFC) at 0 degrees C with a high-binding affinity (Kd = 17 pM). Saturation occurred at 0.5 nM (10 ng/ml) which produces a nearly maximum biological effect. One half of the radiolabelled rSCF was internalized by the cells after 30 min at 37 degrees C. No significant differences in the receptor number, dissociation constant, or internalization rate were found between normal and PV ECFC. Autoradiographic analysis of 125I-rSCF binding to normal BFU-E and ECFC showed that no differences were present in either the percentage of positive cells or the number of radioactive grains/cell between the normal and PV erythroid progenitors. The enhanced sensitivity of PV BFU-E and CFU-E to SCF does not appear to be related to changes in SCF receptor number, binding affinity or internalization and the hypersensitivity of PV erythroid progenitors to SCF must reside in a further internal cellular abnormality.

Autoradiography↗

PubMed1994

Distinct roles of erythropoietin, insulin-like growth factor I, and stem cell factor in the development of erythroid progenitor cells.

Erythropoietin (EP), insulin-like growth factor I (IGF-I) and stem cell factor (SCF) each reduce apoptosis of human erythroid progenitor cells. To determine if these growth factors have additional roles in stimulating erythropoiesis, the proliferation, maturation, and survival of highly purified human erythroid colony-forming cells (ECFCs) were studied during the application of different combinations of these growth factors in a serum-free liquid culture. EP maintained cell viability and supported heme synthesis during erythroid maturation, with little increase in viable cell number or stimulation of DNA synthesis. The addition of SCF with EP resulted in a substantial increase in DNA synthesis, which was greater than that seen with the addition of EP and was associated with a large expansion in the number of ECFCs. Thus EP, by itself, produces little increase in cell proliferation, and expansion of the number of erythroid cells depends upon the presence of SCF with EP. The addition of IGF-I with EP led to enhanced heme synthesis and moderate cellular proliferation, but also greatly enhanced nuclear condensation and enucleation in the late erythroblasts. Thus EP, by itself, is not sufficient for complete end-terminal nuclear condensation/enucleation and the presence of IGF-I is necessary for this complete process. While EP greatly reduced apoptosis during 16 h of incubation at 37 degrees C, the addition of SCF and IGF-I with EP had little additional effect, but these additions enhanced DNA synthesis > 3.4-fold. Thus SCF may have an additional role in directly stimulating proliferation through a process that is distinct from apoptosis. Our observations indicate that EP prevents apoptosis and maintains erythroid cell viability and development. IGF-I enhances erythroid maturation and proliferation, but the proliferation of erythroid progenitors is mainly controlled by the addition of SCF with EP, independent of an effect on apoptosis.

Apoptosis↗

PubMed1993

Apoptosis of human erythroid colony-forming cells is decreased by stem cell factor and insulin-like growth factor I as well as erythropoietin.

To clarify the manner by which erythropoietin (EP), stem cell factor (SCF), or insulin-like growth factor I (IGF-I) regulate erythropoiesis, apoptosis of human erythroid progenitor cells was investigated. Human burst-forming units-erythroid (BFU-E) partially purified from peripheral blood were cultured for 6 days to generate erythroid colony-forming cells (ECFC), which consist mainly of colony-forming units-erythroid (CFU-E). The cells were labeled with [3H]thymidine, incubated in serum-free liquid media, at 37 degrees C, for 16 h, and the pattern of DNA breakdown was analyzed by agarose gel electrophoresis. When these cells were incubated without EP, 70% of the total cellular DNA was broken down into DNA fragments of less than 5 kilobases and nuclear condensation and fragmentation, characteristic of apoptosis, were evident. While EP greatly reduced the amount of DNA breakdown to 23%, SCF and IGF-I each reduced the amount of DNA breakdown to 38-46% and, when added together, to 24%. Dose-response experiments with SCF and IGF-I showed a dose-dependent reduction in DNA fragmentation at concentrations that stimulate colony formation in serum-free semisolid cultures. Finally, assays of ECFC performed by the plasma clot method, after serum-free liquid culture, at 37 degrees C, for 16 h, demonstrated marked protection of erythroid colony-forming capacity by SCF or IGF-I in the absence of EP, as well as by EP itself. These data indicate that human erythroid progenitor cells undergo apoptosis which is reduced by SCF and IGF-I as well as EP and suggest that the control of apoptosis by each of these factors has a prominent role in the regulation of erythropoiesis.

Apoptosis↗

PubMed1993

Inhibition of human erythroid colony-forming units by tumor necrosis factor requires beta interferon.

We have previously reported that inhibition of human CFU-erythroid (E) colony formation by tumor necrosis factor (TNF) is an indirect effect mediated by a soluble factor released from a fraction of marrow accessory cells which are predominantly stromal elements (Means, R. T., Jr., E. N. Dessypris, and S. B. Krantz. 1990. J. Clin. Invest. 86:538-541). Further studies reported here identify a mediator of this effect. The inhibitory effect of recombinant TNF on marrow CFU-E is ablated by neutralizing antibodies to human beta IFN, but not by antibodies to gamma IFN or IL-1. Anti-beta IFN also neutralizes the inhibitory effect of conditioned medium prepared from marrow cells exposed to TNF. Human beta IFN inhibits colony formation by unpurified marrow CFU-E as well as highly purified CFU-E generated from peripheral blood progenitors, and limiting dilution analysis shows that this is a direct inhibitory effect. TNF has been implicated in the pathogenesis of the anemia of chronic diseases since blood TNF levels are elevated in many patients with this syndrome, and since exposure to TNF produces a similar anemia in either humans or mice. The present study demonstrates that beta IFN is a required mediator of this inhibitory effect on erythropoiesis.

Erythroid Precursor Cells↗

PubMed1992

Differentiation and erythropoietin receptor gene expression in human erythroid progenitor cells.

Partially purified human burst-forming unit-erythroid (BFU-E) cells from peripheral blood were cultured for 6 to 8 days to obtain colony-forming unit-erythroid (CFU-E) cells. When these BFU-E-derived CFU-E were further purified and recultured in liquid suspension cultures with erythropoietin (EPO), they matured and differentiated into reticulocytes in vitro. A maximum rate of hemoglobin synthesis was observed at day 10 of cumulative culture time by measuring 59Fe incorporation into heme. Withdrawal of EPO from erythroblast cultures at various times during development showed that between day 10 and day 11 (when the majority of the cells are in the polychromatic erythroblast stage), these cells became independent of EPO. The timing of the disappearance of the EPO requirement in these cells coincided with the marked decline in proliferation. Measurement of EPO receptor messenger RNA (mRNA) levels by Northern analysis showed that there is a slight decline during the day 8 to day 10 time period, followed by a rapid decline between days 10 and 14. Binding of 125I-EPO to erythroblasts also showed a steady decline of the cell surface binding during maturation and terminal differentiation. The half-life of the human EPO receptor was 90 minutes in the presence of the transcriptional inhibitor actinomycin D and the half-life measured at two different times during the 8- to 14-day culture period remained constant. These results indicate that human EPO receptor mRNA must be transcribed continuously to maintain the levels seen by Northern analysis. The human cell system described here is well suited for the study of a wide variety of biochemical events during late erythroid differentiation.

Blotting, Northern↗

PubMed1992

Progress in understanding the pathogenesis of the anemia of chronic disease.

Improved understanding of the inflammatory response and the identification and characterization of the specific cytokines involved, as well as improved understanding of erythropoiesis, and the availability of recombinant human growth factors such as EPO, have greatly enhanced our appreciation of the pathogenesis of ACD by allowing development of a number of informative models for studying this syndrome. It appears that a variety of cytokines are involved in all aspects of the pathogenesis of ACD, from the inhibition of erythroid progenitors and EPO production to impairment of iron release. A schematic of the contributions of some of these cytokines to the development of ACD is shown in Fig 6. The exact biochemical mechanisms by which these effects occur is still to be determined. The progress outlined in this report has allowed us to develop a more precise understanding of the pathogenesis of this common and important clinical syndrome. In 1983, Hansen subtitled a review of ACD "A Bag of Unsolved Questions." Although this description is still accurate, our understanding of ACD has now developed to the point where we can offer a more defined subtitle: "A Bag of Cytokines."

Anemia↗

PubMed1992

Polycythemia vera. II. Hypersensitivity of bone marrow erythroid, granulocyte-macrophage, and megakaryocyte progenitor cells to interleukin-3 and granulocyte-macrophage colony-stimulating factor.

Polycythemia vera (PV) is a clonal disease of the hematopoietic stem cell characterized by a hyperplasia of marrow erythropoiesis, granulocytopoiesis, and megakaryocytopoiesis. We previously reported that highly purified PV blood burst-forming units-erythroid (BFU-E) are hypersensitive to recombinant human interleukin-3 (rIL-3). Because these cells may be only a subset, and not representative of marrow progenitors, we have now studied partially purified marrow hematopoietic progenitor cells. Dose-response experiments with PV marrow BFU-E showed a 38-fold increase in sensitivity to rIL-3 and a 4.3-fold increase in sensitivity to recombinant human erythropoietin (rEpo) compared with normal marrow BFU-E. In addition, PV marrow colony-forming units-granulocyte-macrophage (CFU-GM) and CFU-megakaryocyte (CFU-MK) also showed a marked hypersensitivity to rIL-3 and to human recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF). Dose-response curves with rGM-CSF and blood BFU-E showed a 48-fold increase in sensitivity. No effect of rIL-4, rIL-6, human recombinant granulocyte-CSF (rG-CSF), or macrophage-CSF (rM-CSF) was evident, nor was there any effect of PV cell-conditioned medium on normal BFU-E, when compared with normal cell-conditioned medium. Autoradiography with 125I-rEpo showed an increase in Epo receptors after maturation of PV BFU-E to CFU-E similar to that shown with normal BFU-E, but no increase of specific binding of 125I-rIL-3 by PV CD34+ cells was seen compared with normal CD34+ cells. These studies show that PV marrow hematopoietic progenitor cells are hypersensitive to rIL-3 and rGM-CSF, similar to PV blood BFU-E. While the mechanism does not appear to be due to enhanced binding of rIL-3, the hypersensitivity of PV progenitor cells to IL-3 and GM-CSF may be a key factor in the pathogenesis of PV.

Bone Marrow↗

PubMed1992

Inhibition of human erythroid colony-forming units by interleukin-1 is mediated by gamma interferon.

IL-1 inhibits erythropoiesis in vivo and in vitro. This inhibition was studied by comparing the effect of recombinant human IL-1 (rhIL-1) on highly purified CFU-erythroid (E) generated from peripheral blood burst-forming units-erythroid (BFU-E) (mean purity 44.4%) with its effect on unpurified marrow CFU-E (mean purity 0.36%). Colony formation by marrow CFU-E was significantly inhibited by rhIL-1, while colony formation by highly purified CFU-E was not inhibited. However, purified CFU-E colonies were inhibited by rhIL-1 in the presence of autologous T-lymphocytes, and also by cell-free conditioned medium prepared from T-lymphocytes stimulated by rhIL-1. This inhibitory effect was ablated by neutralizing antibodies to gamma interferon (IFN), but not by antibodies to human IL-1, tumor necrosis factor, or beta IFN. Colony formation by highly purified CFU-E was also inhibited by recombinant human gamma IFN (rh gamma IFN). IL-1 and gamma IFN play significant roles in the pathogenesis of the anemia of chronic disease. These studies indicate that rhIL-1 inhibits CFU-E colony formation by an indirect mechanism involving T-lymphocytes and requiring gamma IFN and that gamma IFN itself is most probably the direct mediator of this effect.

Animals↗