Erythroid progenitor cell kinetics in chronic haemodialysis patients responding to treatment with recombinant human erythropoietin

The response of bone marrow and peripheral blood erythroid progenitors to human recombinant erythropoietin (rHuEPO) was studied in nine haemodialysed renal failure patients receiving this hormone for the correction of their anaemia. The haematocrit rose in all patients in response to thrice weekly injections of escalating rHuEPO doses (12-192 IU/kg). Both the numbers of CUF-e and BFU-e and their proliferative state in the bone marrow as well as BFU-e numbers in the peripheral blood were estimated before treatment and again after correction of the anaemia, at 16 h following an intravenous dose of rHuEPO. Following treatment bone marrow BFU-e numbers fell to a mean of 24.5% (P less than 0.01) of the pre-treatment values although there was no significant change in CFU-e or circulating BFU-e numbers. The mitotic rate (percentage S-phase cells) estimated by tritiated thymidine suicide rose from 45.2% to 68.4% (P less than 0.05) in the case of CFU-e and from 16.4% to 45.1% (P less than 0.05) for BFU-e following treatment with rHuEPO thus indicating in-vivo sensitivity of both the primitive as well as the mature erythroid progenitors to the hormone. The fall in BFU-e numbers in the bone marrow after several months of treatment may be due to a loss of cells from this progenitor pool by maturation that is uncompensated by replacement from the pluripotential stem cell compartment.     

Changes of the phenotype of erythroid progenitor cells (BFU-E, CFU-E) and their progenies during early steps of differentiation

Early differentiation processes of human erythroid progenitor cells (BFU-e, CFU-e) have been studied during in vitro proliferation using a panel of monoclonal antibodies with known reactivity on different levels of the erythroid cell line. Two antibodies recognizing structures on BFU-e (VIP-2b, BMA 021), two antibodies reactive with CFU-e and nucleated red cells (5F1, CLB-Ery-3) and one antibody directed against glycophorin A (VIE-G4) were used for this study. Normal human bone marrow cells were induced to proliferation in an erythroid progenitor cell assay and, after different periods of incubation, agar cultures were treated with these antibodies and complement. Thereafter, the remaining erythroid cells were incubated again to continue their proliferation with the same stimulators as before. The changes of the phenotype of BFU-e and CFU-e progenies during in vitro proliferation were determined by the reduction of colony formation in comparison with untreated control cultures. Our results indicate that the loss of HLA-DR antigens and the p45 structure is accompanied by the acquisition of structures recognized by the antibodies 5F1 and CLB-Ery-3. After 5-7 d of incubation BFU-e derived progenies exhibit the same antigenic structure as has been found for CFU-e. Glycophorin A expression could only be demonstrated at a late differentiation stage of the erythroid cell lineage.     

In vitro erythropoiesis. II. Cytochemical enumeration of erythroid stem cells (CFU-e and BFU-e) from normal mouse and human hematopoietic tissues.

A procedure which provides more accuracy in the identification and scoring of in vitro derived erythroid colonies (i.e. BFU-e and CFU-e) from normal mouse and human marrow and spleen cells has been developed. Up to now identification of erythroid colonies has been based on staining with acidified benzidine which consistently displays "background staining" of granulocyte/macrophage colonies. The staining reaction with these reagents is such that identification of erythroid colonies is less than precise. The modified benzidine procedure described herein eliminates the staining reactivity of granulocyte/macrophage colonies and specifically reacts with only those cells that contain hemoglobin. Furthermore, individual colonies and/or cells may readily be counterstained and examined by high resolution light microscopy.     

Temporal response of murine pluripotent stem cells and myeloid and erythroid progenitor cells to low-dose glucan treatment

B6D2F1 female mice were intravenously administered 0.4 mg of glucan. 1, 5, 11, and 17 days later, the total nucleated cellularity (TNC) and the numbers of pluripotent hemopoietic stem cells (CFU-s), granulocyte-macrophage progenitor cells (GM-CFC), and erythroid colony-forming (CFU-e) and burst-forming (BFU-e) cells were assayed in the bone marrow and spleen. Bone marrow TNC was not altered, but splenic TNC increased approximately twofold on day 5 and remained increased on days 11 and 17 after glucan treatment. The concentrations of bone marrow and splenic CFU-s and GM-CFC both significantly increased (p less than 0.01) by 5 days after glucan administration; however, they returned to control levels by day 17. Splenic CFU-e concentration increased on days 5, 11, and 17, whereas splenic BFU-e concentration increased only on day 11 after treatment. By contrast, bone marrow CFU-e and BFU-e concentrations were either unaffected or slightly decreased by glucan treatment. When peripheral blood was assayed for CFU-s and GM-CFC, no detectable increase in the concentrations of these progenitors was noted at any time after glucan treatment. The relevance of these effects of low-dose (0.4 mg) glucan treatment is discussed with respect to previously reported effects of higher-dose (e.g., 4.0 mg) glucan treatment.     

Erythropoiesis in vitro. III. The role of potassium ions in erythroid colony formation.

The role of potassium as an essential promotor of erythroid progenitor growth (BFU-e & CFU-e) from normal murine hematopoietic tissues was studied. Dialyzed fetal calf serum, over a wide range of concentrations, was shown to reduce the numbers of BFU-e and CFU-e that could be cultured from normal murine bone marrow. A dose-dependent addition of 1 M KC1 restored erythroid progenitor colony growth to the levels generally seen when normal, non-dialyzed fetal calf serum was used. Furthermore, when [K] was increased in some human urinary and sheep plasma erythropoietin preparations, the number of erythroid progenitor cells cultured also increased. This influence is crucial to the differentiation of committed stem cells into the erythroid pathway and must therefore be considered in the development of serum-free growth media.     

Factor-independent erythropoietic progenitor cells in leukemia induced by the myeloproliferative leukemia virus

The myeloproliferative leukemia virus (MPLV), a novel murine retroviral complex that does not transform fibroblasts, has been shown to cause an acute leukemia in adult mice accompanied by a progressive polycythemia. The present study demonstrates that, on in vivo inoculation, MPLV induces a rapid suppression of growth factor requirement for in vitro colony formation by both the late and the primitive erythroid progenitor cells. CFU-e-derived erythrocytic colonies developed and differentiated in semi-solid medium without the addition of erythropoietin (Epo). In addition, the formation of CFU-e colonies was not altered by the presence of specific neutralizing Epo antibodies. In the spleen, the CFU-e pool size increased rapidly up to 30-fold. By day 6 postinfection, 100% of these progenitor cells were Epo-independent. The in vivo effects of MPLV-infection on early erythroid progenitor cell compartments were examined in cultures grown for seven days. The concentration of erythroid progenitor cells was twofold elevated in spleen from MPLV-infected mice. As early as day 4 postinfection, 50% of these progenitors produced fully hemoglobinized colonies in serum-free cultures without the addition of interleukin-3 (IL-3) and Epo. Most spontaneous colonies were large and contained up to 10(5) cells per colony. They were composed of either erythroblasts only (16%) or erythroblasts and megakaryocytes (70%); few of them were multipotential (14%). In the marrow, the total number of BFU-e was reduced and only few factor-independent bursts were observed, suggesting a rapid migration of infected progenitors from marrow to spleen. Furthermore, the data show that abnormal erythropoiesis was due to the replication defective MPLV information and was not influenced by the Fv-2 locus.     

A case of acquired pure red cell anemia studied by cloning of erythroid progenitor cells in vitro

A 70-year-old woman developed typical clinical symptoms of pure red cell anemia (PRCA) following a history of rheumatoid arthritis (RA). The patient's bone marrow erythropoietic progenitors cells were cloned in a micro agar culture system several times over a period of 11 months, revealing a diminished frequency of bone marrow erythroblasts paralleled by a markedly reduced number of CFU-e and BFU-e in vitro. No inhibitory activity in the patient's IgG fraction could be detected either by preincubation with IgG and/or rabbit complement, or in the continuous presence of IgG. Depletion of T lymphocytes from the patient's bone marrow cells led to an improved in vitro erythroid proliferation. Cytostatic therapy with cyclophosphamide clinically induced a marked increase in the bone marrow erythroblast and reticulocyte number, correlated in vitro by normalization of CFU-e levels and increase in the number of BFU-e. Nevertheless, BFU-e values never attained normal levels, which could be attributed to a reduced stem cell pool resulting from previous therapy with cyclophosphamide and/or antirheumatic drugs. Two independent factors, a reduced pool of committed stem cells as well as an autoimmune cell-mediated suppression, may both contribute to the pathomechanism of the disease in this patient.     

Circulating Granulocytic and Erythroid Progenitor Cells in Chronic Granulocytic Leukaemia

S ummary . We used a standard methyl cellulose method to assay erythroid progenitor cells in the blood of 35 patients with untreated CGL and of 18 normal controls. In 28 patients we simultaneously assayed granulocyte/nionocyte committed progenitor cells (CFU-c) by an agar method. Circulating erythroid burst-forming units (HFU-e) in CGL wcre increased above normal by a factor of about 180; CFU-c were increascd by a factor of about 9000. Both BFU-e and CFU-c numbers were linearly related to the total leucocyte count in individual patients but not to numbers of circulating blast cells. There was a positive correlation in individual patients between CFU-c and BFU-e numbers. Circulating BFU-e and erythroid colony-forming cells (CFU-e) were unable to proliferate in vitro in the absence of erythropoietin. We conclude that erythroid progenitor cells are involved in the'clonal expansion'that characterizes CGL, but apparently to a lesser extent than are granulocyte/monocyte progenitor cells.     

Comparison of hemin enhancement of burst-forming units-erythroid clonal efficiency by progenitor cells from normal and HIV-infected patients.

The ability of peripheral-blood hematopoietic progenitor cells from AIDS patients and normal controls to respond to erythropoietin (Epo) was assessed for burst-forming units-erythroid (BFU-E). BFU-E colony formation from AIDS patients' peripheral blood responded to a wide range of Epo concentrations (0.5-4 U) in a similar manner as erythroid progenitors obtained from normal peripheral blood. The optimum dose response of BFU-E to Epo was 2 U which resulted in generation of 71 +/- 4 BFU-E in AIDS patients (n = 10), as compared to 77 +/- 5 BFU-E in normal donors (n = 3). The optimum concentration range of hemin enhancement of erythroid progenitor BFU-E was 10-50 microM. In all instances, Epo was essential for BFU-E growth. Inclusion of hemin at a concentration of 10 microM in AIDS patients' peripheral-blood erythroid progenitor cells resulted in enhancement of BFU-E by 136-215%. Similarly, inclusion of hemin (10-100 microM) in normal bone marrow erythroid progenitor cell cultures resulted in enhancement of BFU-E. Inclusion of an equivalent amount of iron or tin protoporphyrin to progenitors cells from AIDS patients' peripheral blood had no effect on the number of colonies observed. On the other hand, inclusion of another heme analogue, zinc protoporphyrin, in AIDS or normal cultures resulted in a 50% suppression of BFU-E colony formation. These results demonstrate that peripheral-blood mononuclear cells from AIDS patients retain the capacity to generate erythroid precursors such as BFU-E in the presence of Epo, and that hemin has a specific enhancement effect on growth of BFU-E colony formation obtained from peripheral blood or bone marrow cells.     

Abnormalities of T lymphocyte subsets in Beh?et's disease demonstrated with anti-CD45RA and anti-CD29 monoclonal antibodies.

We assessed T cell subpopulations using 2-color flow cytometry with phycoerythrin conjugated anti-CD45RA and anti-CD29 and fluorescein conjugated anti-CD4 and anti-CD8 monoclonal antibodies, on peripheral blood lymphocytes from 19 patients with Beh?et's disease (BD) and 18 healthy control subjects. The percentage of CD4+ cells was significantly lower in patients with BD (34 +/- 2%) than in control subjects (46 +/- 3%) (p less than 0.001). Among CD4+ cells, the percentage of suppressor-inducer (CD4+CD45RA+) cells was significantly lower in patients with BD (31 +/- 4%) than in control subjects (45 +/- 2%) (p less than 0.01), while the percentages of helper-inducer (CD4+CD29+) cells were similar in patients and controls. The percentage of CD8+ cells was significantly higher in patients with BD (36 +/- 2%) than in control subjects (26 +/- 2%) (p less than 0.001) involving both CD45RA+ and CD29+ subpopulations. Within CD4+ cells, the percentage of suppressor-inducer (CD4+CD45RA+) cells was significantly decreased in patients with active BD (26 +/- 3%) compared with control subjects (45 +/- 2%) (p less than 0.001), whereas in patients with inactive BD the difference was statistically insignificant. Our results suggest that the defective suppressive function in patients with active BD may be related to the decreased suppressor-inducer subpopulation (CD4+CD45RA+).     

Charcoal-dextran treatment of fetal bovine serum removes an inhibitor of human CFU-megakaryocytes

Serum has been shown to have an inhibitory effect on the growth of human megakaryocyte colony-forming units (CFU-Meg), both in the methylcellulose and plasma clot systems. Charcoal-dextran (CD) treatment of fetal bovine serum (FBS), a method that is known to adsorb free hormones and various other substances from their protein-bound counterparts, has been used for the improvement of human erythroid colony growth. We now report a CD treatment of FBS to improve human CFU-Meg growth, using a modified plasma clot colony assay. We found 22.9 +/- 1.9 (SEM) colonies in cultures containing CD-treated FBS, as compared to 7.1 +/- 0.9 colonies with control FBS (different at p less than or equal to 0.0001). Serum treated with dextran alone produced 5.0 +/- 1.6 colonies, which did not differ from control FBS. Cultures containing CD-treated human AB serum, human AB plasma, and citrated bovine plasma yielded significantly (p less than 0.05) fewer colonies than those with CD-treated FBS. Endotoxin, cortisol, T3, T4, insulin, estradiol, testosterone, and erythropoietin levels were not changed in a consistent direction by the CD treatment. Our treatment of FBS with CD allows improved growth of human CFU-Meg colonies and likely represents the removal of an as-yet-undefined inhibitor(s) of CFU-Meg. This technique provides the ability to assay for stimulators or potentiators less hampered by the influence of an undefined inhibitor(s).     

Separation of erythroid progenitor cells in mouse bone marrow by isokinetic-gradient sedimentation.

Isokinetic-gradient sedimentation employing a shallow linear gradient of Ficoll in tissue culture medium was used to isolate erythroid progenitor cells (CFU-e) from mouse bone marrow. Following gradient sedimentation, 34% of the total nucleated cells and 48% of the CFU-e applied to the gradient were recovered, and three distinct modal populations of CFU-e could be distinguished. The slowest-migrating population did not require exposure to exogenous erythropoietin in order to form erythroid colonies in vitro. The other two modal populations of CFU-e required exposure to exogenous erythropoietin for differentiation. One of these, constituting 64% of the hormone-dependent CFU-e recovered, migrated with the bulk of the marrow cells, whereas the other migrated ahead of the bulk of the marrow cells. This latter population, which contained 34% of the CFU-e, was recovered with 11% of the marrow cells, representing a twofold to threefold enrichment. BFU-e migrated more slowly than the erythropoietin-dependent CFU-e. Resedimentation studies suggested that the two erythropoietin-dependent CFU-e populations were distinct modal populations. When cells from the fastest-migrating population of erythropoietin-dependent CFU-e were cocultured with unseparated marrow cells, a further twofold to threefold enhancement of erythroid colony formation was obtained. Comparison of isokinetic-gradient sedimentation with discontinuous and continuous albumin density-gradient sedimentation revealed that isokinetic-gradient sedimentation was a more efficient method than the former and a more rapid method than the latter for isolating CFU-e from mouse bone marrow.     

Effects of an aplastic anemia urinary extract on mouse erythroid progenitor cells in vivo

We investigated the in vivo effects of a crude extract from the urine of aplastic anemia patients (AA urinary extract) on erythroid precursor cells in the femoral bone marrow and spleens of normal adult mice. A single intraperitoneal injection of AA urinary extract induced a significant increase in the number of splenic erythroid burst-forming units (BFU-e) and erythroid colony-forming units (CFU-e) within 24 h after injection. We then injected pure recombinant erythropoietin (Epo) equivalent to the amount present in the urinary extract. This addition increased the number of splenic CFU-e by almost the same degree as the amount induced by the AA urinary extract 24 h after injection, but failed to elicit any change in the number of splenic BFU-e. In other studies, mice were injected with the same amount of lipopolysaccharide (LPS) and/or pure Epo as that present in the AA urinary extract. Experiments with Limulus amebocyte lysate-adsorbed (endotoxin-depleted) or nonadsorbed (endotoxin-containing) AA urinary extracts showed that endotoxin contamination interfered with the increase in numbers of marrow CFU-e and enhanced the increase in splenic CFU-e numbers induced by pure Epo or Epo activity in the AA urinary extract. The number of splenic BFU-e, however, was not affected by administration of LPS and/or Epo or by adsorbed endotoxin. These data suggest that AA urinary extract contains a stimulating activity for mouse splenic BFU-e, and that this activity is not attributable to the Epo activity or endotoxin contamination within the urinary extract.     

In vitro myelotoxicity of 2',3'-dideoxynucleosides on human hematopoietic progenitor cells

Three nucleoside analogues, 2',3'-dideoxyadenosine (ddA), 2',3'-dideoxyinosine (ddI), and 2',3'-dideoxycytosine (ddC), were evaluated for their potential myelotoxic effects to normal human hematopoietic progenitor cells. The myeloid (granulocyte-monocyte colony-forming units, CFU-gm) and erythroid (erythroid burst-forming units, BFU-e: and erythroid colony-forming units, CFU-e) committed progenitor cells were exposed to the agents for a 1-h period prior to culture in a microcapillary assay or continuously exposed during the entire culture period. Both ddA and ddI (100 microM) were mildly toxic (less than 50% colony inhibition) to human CFU-gm, BFU-e, and CFU-e following either 1-h or continuous exposures. Marrow progenitor sensitivities to ddA and ddI were indistinguishable. Colony inhibition ranged from 47% to 67% for 1-h ddC exposure (100 microM), values that were comparable to ddA and ddI. Continuous exposure to ddC was highly myelotoxic to human hematopoietic progenitors, with concentrations of 10 and 100 microM suppressing colony formation by 79%-92% and 93%-97%, respectively. These results demonstrate that 1-h and continuous exposures to ddA and ddI were similarly myelotoxic to human hematopoietic cells, whereas a 1-h exposure to ddC was equivalent to ddA and ddI, yet continuous ddC exposure was extremely toxic to marrow cell progenitors.     

Association between urinary zinc excretion and lymphocyte dysfunction in patients with lung cancer

Patients with bronchogenic carcinoma often have low serum zinc concentrations and sometimes have markedly elevated renal zinc losses. Since normal zinc metabolism is critical for the proper function of T lymphocytes and natural killer cells, the effect of zinc status on T cell phytohemagglutinin response and peripheral blood lymphocyte natural killer cell activity was studied in patients with lung cancer. Mean (+/- SEM) serum zinc concentration in 75 patients with cancer was 67.4 +/- 2.2 micrograms/dl versus 96.0 +/- 8.0 micrograms/dl for normal subjects. Patients with low serum zinc levels (less than 70 micrograms/dl) had significantly higher urine zinc excretion than patients with normal serum zinc levels (1,385 +/- 240 micrograms per 24 hours versus 392 +/- 107 micrograms per 24 hours) (p less than 0.001). This pattern of zinc concentrations (i.e., low serum zinc in combination with high urine zinc) is typical of patients with mild zinc deficiency, and suggests that a mild chronic zinc deficiency state was present in some of these patients. When lymphocyte data were analyzed according to serum zinc concentrations and urinary zinc excretion, low serum zinc concentration and high urine zinc excretion both correlated with depressed T cell phytohemagglutinin response (p less than 0.005 and p less than 0.001, respectively). For instance, mean maximal phytohemagglutinin response in patients with urinary zinc excretion of more than 700 micrograms per 24 hours was 22,132 +/- 3,201 cpm (n = 14) compared with 68,130 +/- 6,850 cpm for patients with normal zinc excretion (n = 7). Peripheral blood lymphocyte natural killer cell activity did not correlate with either serum or urine zinc values. Oral zinc sulfate (220 mg, three times daily for six weeks) was then administered to patients with hyperzincuria (mean = 992 micrograms per 24 hours). Zinc-supplemented patients had normalization of T cell phytohemagglutinin response after zinc therapy, whereas control patients demonstrated continued T cell dysfunction. Natural killer cell activity did not change in either group during the study period. These data suggest that a mild subclinical zinc deficiency state may exist in some patients with lung cancer and may be an important cause of abnormal T cell function. Furthermore, zinc supplementation may be useful to improve lymphocyte function in selected patients. Whether zinc supplementation would alter the course of the disease or the patient's prognosis is presently unknown.     

Effects of T cells and monocytes on globin chain synthesis in erythroid bursts cultured from human peripheral blood burst-forming units (BFU-e)

Globin chain synthesis was studied in mature erythroid bursts cultured from the peripheral blood null cells of 21 normal individuals. Although coculture of autologous T-lymphocytes with null cells resulted in a fivefold increase in the yield of erythroid bursts, the proportion of gamma chains synthesized (gamma/gamma + beta) was not significantly different from that seen without T cells. Coculture with autologous monocytes also resulted in increased burst-forming unit (BFU-e) proliferation but the ratio gamma/gamma + beta (0.25 +/- 0.03) was significantly higher than that seen with null cells alone (0.08 +/- 0.006) or with T cells (0.10 +/- 0.008). The relative increase in gamma-chain synthesis correlated with the severity of megaloblastic changes in erythroid progeny of BFU-e (P less than 0.01) but showed no significant relationship to the extent of colony maturation assessed by erythroblast maturity.     

Improved diabetic control enhances erythroid stem cell proliferation in vitro

Patients with type I diabetes mellitus treated with continuous sc insulin infusion (CSII) have improved glucose homeostasis, metabolic control, and linear growth. To determine the influence of CSII on cellular growth in vitro, we used a clonal stem cell assay for proliferation of erythroid progenitors, [burstforming units-erythroid (BFU-E)] in peripheral blood. Eight patients were studied before and after 1 week of CSII. Improvement in metabolic control was demonstrated by a decrease in mean 24-h plasma glucose from 232 +/- 29 (+/- SEM) mg/dl before treatment to 112 +/- 3 mg/dl after treatment (P = 0.01). Somatomedin-C levels increased from 1.1 +/- 0.4 to 1.4 +/- 0.4 U/ml (P less than 0.001). Numbers of BFU-E-derived colonies were not different from normal during conventional treatment, but increased 300% after 1 week of CSII. Our findings indicate that the acute metabolic and hormonal improvements that accompany short term CSII therapy in vivo are associated with a striking increase in the proliferation of erythroid committed stem cells in vitro.     

HLA-DR positive T lymphocytes in blood and synovial fluid in rheumatoid arthritis

HLA-DR expression was found on 6.7 +/- 0.7% of blood T lymphocytes from 34 patients with rheumatoid arthritis (RA) and 2.6 +/- 0.3% of T cells from normals. In synovial fluid (SF) of RA patients high percentages were found (56 +/- 3%, n = 18); the SF T lymphocytes showed an acid alpha-naphthyl acetate esterase pattern and cell morphology compatible with activated T lymphocytes. The helper function for pokeweed mitogen induced B cell differentiation was higher in blood T lymphocytes from RA patients than from normal controls; in SF T lymphocytes helper function could not be established. These differences between blood and SF T lymphocytes may represent differences in recirculating and homing properties of T cells in these compartments.     

Increased response of erythroid progenitors to interleukin-3 in sickle cell anemia: CFU-E-like behavior of circulating erythroid progenitors in liquid culture

Erythroid progenitors circulating in peripheral blood and their response to erythropoietin (EPO), interleukin-3 (IL3), and phytohemagglutinin-stimulated, lymphocyte-conditioned medium (PHALCM) were assessed in sickle cell anemia (SCA) patients and controls. SCA patients have significantly higher numbers of circulating burst-forming unit-erythroid (BFU-E) compared with controls (mean +/- SEM, 940.27 +/- 129.11 per ml and 86.56 +/- 19.74 per ml, respectively; P less than 0.0001). At low doses of EPO, BFU-E-derived colonies were significantly increased in SCA patients compared with controls (each P less than 0.05). The EPO dose required to produce 50% of maximum colony numbers was 47 times greater in control subjects than in SCA patients. Moreover, in 11 of 17 patients with SCA, spontaneous BFU-E-derived colonies were formed without added erythropoietin. This phenomenon was not observed in control subjects (P = 0.035). PHALCM developed from mononuclear cells of SCA patients had significantly greater stimulatory effect than did that derived from controls regardless of the source of target cells (each P less than 0.05). A two-step study of IL3 sensitivity of erythroid progenitors was conducted. First, in a liquid culture system, circulating erythroid progenitors of SCA patients and controls were incubated in the presence of varying doses of IL3. During a second step, CFU-E-like colonies were observed in methylcellulose cultures of these cells. The mean numbers of colony-forming unit-erythroid (CFU-E)-like colonies was significantly higher in SCA patients compared with control subjects at low doses of IL3 (each P less than 0.02). The increased response of erythroid progenitors to IL3 and the increased production of hemopoietic growth factors (IL3 or non-IL3) contribute to the hemopoietic response in SCA patients. These mechanisms and increased sensitivity of the BFU-E to EPO may explain lower than expected EPO levels in SCA patients.     

T-lymphocyte subsets as noninvasive markers of cardiomyopathy

Fifty-eight patients with symptomatic congestive heart failure were examined for T-lymphocyte subsets in the peripheral blood using two-color laser flow cytometry as a noninvasive diagnostic procedure. The final diagnosis established by catheterization and endomyocardial biopsy were dilated cardiomyopathy (DCM, n = 24), myocarditis (MC) by the Dallas criteria (n = 12), and coronary heart disease (CHD, n = 16). The CD8+CD11- (cytotoxic T) subset was significantly low in patients with DCM (13.9 +/- 4.4 vs. controls, p less than 0.05) in comparison with MC (20.7 +/- 10.9) and CHD (22.3 +/- 5.9). Moreover, the CD4+2H4+ (suppressor/inducer T) subsets were higher in patients with DCM (27.3 +/- 6.9 vs. controls, p less than 0.01) than in those with MC (17.3 +/- 7.8) and CHD (15.6 +/- 7.9). The CD4/CD8 and CD4+2H4+/CD8+CD11- ratio were examined and compared with those of normal controls (NC n = 16). The CD4+2H4+/CD8+CD11- ratio was clearly higher in patients with DCM (2.2 +/- 0.9 vs. controls, p less than 0001) than in those with MC (1.1 +/- 0.6) CHD (0.9 +/- 0.7). A CD4+2H4+/CD8+CD11- ratio of greater than 1.6 was considered to facilitate diagnosis of dilated cardiomyopathy with 79% sensitivity and 70% specificity. There was no significant increase in the ratios between MC and CHD. However, the proportion of the CD8+Leu7+ (natural suppressor) subset of circulating T lymphocytes in patients with MC was statistically higher (19.1 +/- 6.3% vs. controls, p less than 0.05) than in DCM or CHD. An elevated ratio of CD4+2H4+/CD8+CD11- among peripheral blood lymphocytes may thus be a useful marker for differential diagnosis of dilated chronic cardiomyopathy from myocarditis and coronary heart disease.