The anemia of end-stage renal disease: hematopoietic progenitor cell response

Patients with the anemia of end-stage renal disease (ESRD) fail to display an appropriate compensatory increase in red cell production. In order to investigate the extent to which the impaired erythropoietic response is determined at the progenitor cell level, we determined the frequencies of marrow colony-forming cells in 11 anemic and 3 non-anemic, dialysis-dependent ESRD patients and 10 healthy individuals. In addition, we measured serum levels of erythropoietin (Epo) by radioimmunoassay. There were no significant differences (P greater than 0.1) between normal and ESRD groups in the frequencies of primitive or late erythroid (BFU-E and CFU-E, respectively), granulocyte-macrophage, and megakaryocyte progenitors, CFU-E/BFU-E ratios, or serum Epo levels. In contrast, 5 non-uremic patients with chronic anemia comparable in severity to the anemic ESRD patients had serum Epo levels and CFU-E/BFU-E ratios that were significantly increased (P less than 0.05 and P less than 0.001, respectively) in comparison to the normal controls and ESRD patients. Pre-dialysis serum and plasma from both ESRD groups were as supportive of autologous erythroid and non-erythroid colony growth in vitro as normal serum and plasma; inhibition was not observed. We conclude that the relative numbers of erythroid and non-erythroid progenitors and the majority of serum Epo levels are unchanged from normal in patients with the anemia of ESRD. However, their normal CFU-E/BFU-E ratio reflects an inadequate compensatory erythropoietic response due to their inability to appropriately increase Epo production in response to anemia. Inhibitors of autologous erythroid colony formation were not detected in ESRD serum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Erythropoietin deficiency and inhibition of erythropoiesis in renal insufficiency

The relative importance of erythropoietin (Ep) and inhibition of erythropoiesis in the anemia of chronic renal insufficiency has been investigated. Sixty patients with varying degrees of renal insufficiency, 40 normal subjects and 40 patients with anemia and normal renal function, were studied. Erythroid (CFU-E) and granulocytic (CFU-GM) progenitor cell colony formation were assayed in fetal mouse liver and human bone marrow cultures, respectively. Erythropoietin was measured by radioimmunoassay. Hematocrit and plasma creatinine concentration correlated with the degree of serum inhibition of CFU-E formation (r = 0.69, P less than 0.001, and r = 0.62, P less than 0.001, respectively). Serum erythropoietin levels in patients with renal insufficiency (34.4 +/- 6.7 mU/ml) were slightly higher than normal values (23.1 +/- 0.98 mU/ml), but showed no relationship to plasma creatinine, hematocrit, or inhibition of CFU-E formation. In contrast, serum erythropoietin concentrations increased exponentially as the hematocrit decreased below 32% (r = 0.61, P less than 0.001), and CFU-E formation was stimulated by serum in anemia patients with normal renal function. Studies of granulopoiesis showed uremic sera supported in vitro CFU-GM growth more efficiently than sera from normal subjects. These results suggest that inhibition of erythroid, but not granulocytic, progenitor cell formation, in addition to a relative erythropoietin deficiency, are the primary factors responsible for the anemia of chronic renal failure.     

Bone marrow failure following severe injury in humans

BACKGROUND: Hematopoietic failure has been observed in experimental animals following shock and injury. In humans, bone marrow dysfunction has been observed in the red cell component and characterized by a persistent anemia, low reticulocyte counts, and the need for repeated transfusions despite adequate iron stores. While a quantitative defect in white blood cell count has not been noted, an alteration in white blood cell function manifesting as an increased susceptibility to infection is well established. Since the etiology of this anemia remains unknown and the bone marrow has been rarely studied following injury, we measured various parameters of hematopoiesis directly using bone marrow from trauma patients and tested the hypothesis that trauma results in profound bone marrow dysfunction, which could explain both the persistent anemia and the alteration in white blood cell function. METHODS: Bone marrow aspirates and peripheral blood were obtained between day 1 and 7 following injury from 45 multiple trauma patients. Normal volunteers served as controls. Peripheral blood was assayed for hemoglobin concentration, reticulocyte count, erythropoietin levels, white blood cell count, and differential. Peripheral blood and bone marrow were cultured for hematopoietic progenitors (CFU-GM, BFU-E, and CFU-E colonies). RESULTS: Bone marrow CFU-GM, BFU-E, and CFU-E colony formation was significantly reduced while peripheral blood CFU-GM, BFU-E, and CFU-E was increased in the trauma patients compared with normal volunteers. Bone marrow stroma failed to grow to confluence by day 14 in >90% of trauma patients. In contrast, bone marrow stroma from volunteers always reached confluence between days 10 and 14 in culture. The mean hemoglobin concentration and reticulocyte counts of the trauma patients were 8.6 +/- 1.0 g/dL and 2.75 +/- 0.7% respectively, while their plasma erythropoietin levels were 2 to 10 times greater than control values. CONCLUSIONS: Release of immature white blood cells into the circulation may also contribute to a failure to clear infection and an increased propensity to organ failure. Concomitantly, profound changes occur within the bone marrow, which include the increased release of erythroid and myeloid progenitors into the circulation, a decrease in progenitor cell growth within the bone marrow, and an impaired growth of the bone marrow stroma. Erythropoietin levels are preserved following trauma, implying that the persistent anemia of injury is related to the failure of the bone marrow to respond to erythropoietin.     

Anemia of chronic renal failure: inhibition of erythropoiesis by uremic serum

The pathogenesis of anemia in patients with end-stage renal disease was studied by assessing the effect of uremic serum on the proliferation and maturation of erythroid progenitor cells, BFU-E and CFU-E, into colonies in vitro. Nucleated peripheral blood cells from 10 anemic patients produced normal or increased numbers of BFU-E colonies in response to added erythropoietin when cultured in control serum, but declined a mean of 63% when autologous uremic serum was substituted. Uremic sera from 90 patients cultured with normal human marrow produced a mean decrease in BFU-E colony growth of 72%, and of CFU-E colony growth of 82%, compared to control serum. Neither hemodialysis nor peritoneal dialysis was effective in removing the inhibitor. We conclude that patients with uremia have adequate circulating erythroid progenitors that respond to erythropoietin normally when removed from the uremic environment, and that uremic serum is toxic and inhibitory to erythropoiesis. This may be an important mechanism in the anemia of chronic renal failure.     

Insulin-like growth factor I plays a role in regulating erythropoiesis in patients with end-stage renal disease and erythrocytosis

Erythroid progenitor growth, the serum hormones that regulate erythropoiesis, and the effect of patient's serum on the growth of normal erythroid progenitors were assessed in eight patients with end-stage renal disease (ESRD) and erythrocytosis. All patients were male and had been on maintenance dialysis, they had a hematocrit >50% and/or a red blood cell count >6 x 10(12)/L and an arterial oxygen saturation >95%. Four had acquired cystic disease of the kidney (ACDK), and four other non-ACDK patients did not have known causes of secondary erythrocytosis after appropriate investigations and long-term follow-up. The methylcellulose culture technique was used to assay the erythroid progenitor (BFU-E/CFU-E) growth. Serum erythropoietin (EPO) and insulin-like growth factor I (IGF-I) levels were measured by RIA. Paired experiments were performed to determine the effects of 10% sera from ESRD patients and control subjects on normal marrow CFU-E growth. The numbers of EPO-dependent BFU-E in marrow and/or blood of patients with ESRD and erythrocytosis were higher than those of normal controls. No EPO-independent erythroid colonies were found. Serum EPO levels were constantly normal in one patient and elevated in three patients with ACDK; for non-ACDK patients, EPO levels were normal or low in two patients and persistently increased in one, but fluctuated in the remaining one on serial assays. There was no correlation between serum EPO levels and hematocrit values. The serum IGF-I levels in patients with ESRD and erythrocytosis were significantly increased compared with normal subjects or ESRD patients with anemia. We found an inverse correlation between serum EPO and IGF-I levels. Sera from patients with ESRD and erythrocytosis exhibited a stimulating effect on normal marrow CFU-E growth. The stimulating effect of sera from patients who had a normal serum EPO level and an elevated IGF-I level could be partially blocked by anti-IGF-I. The present study suggests that IGF-I plays an important role in the regulation of erythropoiesis in patients with ESRD and erythrocytosis who did not have an increased EPO production.     

Trauma inhibits erythroid burst-forming unit and granulocyte-monocyte colony-forming unit growth through the production of TGF-beta1 by bone marrow stroma

OBJECTIVE: To examine the effect of trauma plasma on clonogenic progenitor cultures. SUMMARY BACKGROUND DATA: Severely injured trauma patients often experience altered hematopoietic functions, manifested by an increased susceptibility to infection and the development of a persistent anemia. Experimental and clinical data suggest that trauma results in the release of cytokines into the plasma that have hematopoietic regulatory function, but few studies have examined human bone marrow. METHODS: Plasma was obtained from 42 severely injured patients admitted to the surgical intensive care unit from days 1 to 15 after injury. Bone marrow and normal plasma were obtained from volunteers. Bone marrow mononuclear cells were isolated and plated for granulocyte-monocyte colony-forming unit (CFU-GM) and erythroid burst-forming unit (BFU-E) growth. Parallel cultures were incubated with 2% (v/v) trauma or normal plasma. Additional cultures were plated with neutralizing concentrations of antibodies to transforming growth factor (TGF)-beta1 and MIP-1alpha. Circulating plasma TGF-beta1 was determined by bioassay. mRNA from bone marrow stromal cultures was extracted and probed for TGF-beta1 and macrophage inflammatory protein (MIP)-1alpha. RESULTS: Trauma plasma suppressed CFU-GM and BFU-E colony growth by 40% to 60% at all time periods after injury compared with cultures incubated with normal plasma. Using a noncontact culture system, the authors showed that this inhibition of BFU-E and CFU-GM colony growth was mediated by bone marrow stroma. The inhibition appeared to be due to soluble plasma-induced bone marrow stromal products that did not require direct cell-cell contact. The addition of anti-TGF-beta1 antibodies reversed the suppressive effect of trauma plasma on CFU-GM and BFU-E colony growth during the early but not late time points after injury. Trauma but not normal plasma induced TGF-beta1 mRNA in bone marrow stroma. CONCLUSIONS: Trauma plasma inhibits bone marrow BFU-E and CFU-GM colony growth for up to 2 weeks after injury. This inhibition is mediated through the interaction of trauma plasma with bone marrow stroma. TGF-beta1 production by bone marrow stroma appears to plays an important role in the early but not late bone marrow suppression after injury.     

The impact of a hypercatecholamine state on erythropoiesis following severe injury and the role of IL-6

BACKGROUND: Severe traumatic injury can lead to hemorrhagic shock-induced bone marrow (BM) dysfunction resulting in persistent anemia. The hypercatacholamine state that accompanies severe injury has been shown to impact the growth of erythroid progenitors. IL-6 has a role both in the acute phase response of trauma and has been implicated in the development of anemia. The aim of this study was to investigate the severity of a hyper-adrenergic stimulus on pluripotent progenitors (GEMM-CFU) as well as erythroid progenitors (BFU-E and CFU-E) and the potential regulatory role of IL-6. METHODS: Normal human BM mononuclear cells were isolated and erythropoiesis was assessed by the growth of GEMM-CFU, BFU-E and CFU-E in the presence of adrenergic agonists, norepinephrine (NE) and epinephrine (EPI), at increasing concentrations. Similarly, normal BM stroma cells were grown to confluence then incubated with NE and EPI. Supernatant was harvested and IL-6 levels were determined using ELISA. RESULTS: Under physiologic conditions (10(-7) M), NE and EPI increase BFU-E and CFU-E growth (374% and 177% versus 100% control). At severe stress levels (10(-3) M), NE and EPI completely inhibited BFU-E and CFU-E growth (5% and 4% versus 100% control). GEMM-CFU growth was increased by NE and not EPI at 10(-7) M. The presence of NE and EPI increased IL-6 levels in a dose-dependent fashion. CONCLUSIONS: The proliferative effect of adrenergic agonists at physiologic levels on normal erythropoiesis begins early during erythroid differentiation. At severe stress levels, BFU-E and CFU-E growth is inhibited. The erythropoietic dysfunction and resultant anemia seen following severe injury may be due to the presence of a severe hypercatecholamine state and may be mediated by IL-6.     

慢性肾功能衰竭对大鼠红细胞生成素受体基因表达的影响

目的研究５／６肾切除引起大鼠慢性肾功能衰竭并出现贫血时骨髓细胞红细胞生成素（ＥＰＯ）受体基因表达的变化。方法用二步法切除５／６肾组织诱发大鼠肾功能衰竭并出现贫血，另以盐酸苯肼导致的急性贫血大鼠和正常大鼠为对照。用ＲＴＰＣＲ检测３组大鼠骨髓有核细胞ＥＰＯ受体ｍＲＮＡ表达，并经灰度扫描，与共扩增的β肌动蛋白基因相比，再用ＳｏｕｔｈｅｒｎＢｌｏｔｉｎｇ证实特异性。结果５／６肾切除后５周大鼠出现明显的肾功能衰竭和贫血。正常大鼠、急性贫血大鼠和肾性贫血大鼠骨髓有核细胞ＥＰＯ受体ｍＲＮＡ表达量分别为０２６±００６、０５１±０２４、０１５±００６，急性贫血大鼠ＥＰＯ受体ｍＲＮＡ表达量高于正常大鼠（Ｐ＜００５），而肾性贫血大鼠明显低于正常大鼠（Ｐ＜００１）。结论大鼠在出现慢性肾功能衰竭并出现贫血时，骨髓有核细胞ＥＰＯ受体ｍＲＮＡ表达明显减少，这可能在慢性肾功能衰竭并发贫血过程中起着一定作用。     

Aluminium accumulation in chronic renal failure affects erythropoiesis

Summary: Aluminium (Al) toxicity has been associated with anaemia in exposed patients with chronic renal failure (CRF). the present study was undertaken to determine whether the ingestion of Al citrate was able to affect erythropoiesis in rats with normal or impaired renal function. the renal insufficiency was induced by surgical procedures and control rats were sham operated. Twenty-four rats were allocated to four groups of six rats each: (A) Sham; (B) Sham+Al; (C) CRF; and (D) CRF+Al. the groups B and D received daily doses of Al citrate (0.5 μmol/g bodyweight) and the groups A and C, deionized water, via the intragastric route. At the end of the experimental period (15 weeks) cultures of late erythroid progenitor cells (CFU-E) stimulated with erythropoietin were performed and haematological parameters determined. the liver, kidney, brain, bone and serum Al amounts were quantified. the results are expressed as median and interquartile range. the CFU-E growth was found inhibited in B and D groups (A: 100; B: 74/54-83; C: 86/54-98; D: 46/39-53 %). the haematocrit values were significantly diminished in rats with renal insufficiency when compared to controls (A: 42/40-43; B: 45/42-46; C: 37/32-40 and D: 37/24-39 %). Serum Al accumulation was observed in B and D groups receiving Al (A: 8/5-12; B: 36/36-44; C: 5/5-6; D: 45/26-132 μg Al/l). No differences among groups were found in the liver and kidney Al contents, but uraemic state favoured Al accumulation in brain (A: 6/5.0-9.0; B: 4/3.8-4.3; C: 2/1.0-3.0; D: 15/12.0-21.0 μg Al/g tissue) and bone (A: 29/27-31; B: 30/29-39; C: 42/33-48; D: 68/56-79 μg Al/g tissue). We suggest that the heavy accumulation of Al in the bone compartment may result in a protracted endogenous exposure of bone marrow cells, affecting the erythropoiesis in vivo.     

Haematopoietic progenitor cells in an infant who developed pancytopenia following an extensive burn

We observed a 24-month-old infant who developed anaemia, thrombocytopenia and neutropenia while recuperating from an extensive burn. In order to determine the mechanism(s) responsible for the pancytopenia, we quantified marrow-derived haematopoietic progenitor cells, assessed the relative proliferative rate of haematopoietic progenitor cells, and sought the presence of progenitor cell inhibitors. The concentration and relative proliferative rate of pluripotent progenitors (CFU-GEMM) were elevated. No inhibitors of progenitor cells were observed; in fact, the patient's serum contained very high levels of stimulatory activity for CFU-GEMM as well as for granulocyte-macrophage progenitors (CFU-GM). However, the marrow concentration of erythroid progenitors (BFU-E and CFU-E) was diminished. We conclude that the anaemia in this patient was the result of either hypoproduction of differentiated erythroid progenitors or intramyeloid destruction of early erythroid cells. In contrast, the neutropenia was likely to be due to accelerated neutrophil consumption at a rate that exceeded the capacity for increasing neutrophil production.     

Ribonuclease inhibition of erythropoiesis in anemia of uremia

The anemia of chronic renal failure was studied by assessing the effect of uremic serum on proliferation of human marrow erythroid stem cells into colonies in vitro. Of 50 sera tested, 46 inhibited "CFU-E" colony formation by a mean of 72%, and 42 inhibited "BFU-E" colonies by a mean of 53.5%, compared to normal sera. Analysis of the uremic sera revealed a striking increase of ribonuclease activity in every patient. Mean activity in the study group was 17,346 U/ml serum (range 6,700-36,250) compared to control mean of 1,047 +/- 247 U/ml. Purified ribonuclease added to marrow cultures in concentrations simulating uremic serum produced a dose-dependent decrease in CFU-E colonies suggesting that the substance has a role in the production of anemia of renal failure.     

Hematopoietic inhibitors in chronic renal failure: lack of in vitro specificity

To assess the potential role of retained inhibitors in the pathogenesis of the anemia of chronic renal failure, we have studied simultaneously the effects of increasing concentrations of normal or uremic sera on the growth of erythroid colonies (from CFU-E), granulocyte-macrophage colonies (from CFU-GM), and megakaryocytic colonies (from CFU-Meg) in mouse marrow cell cultures. As compared to normal human serum, increasing concentrations of uremic sera induced a dose-dependent inhibition in the growth of all colony types. Significant correlations (P less than 0.001) were found between the ability of any individual uremic serum to support CFU-E, CFU-GM, and CFU-Meg growth, and, whenever significant inhibition was seen, all three progenitor types were affected. The inhibitory effect on CFU-E growth was significantly greater (P less than 0.01) in patients with serum creatinine concentrations greater than or equal to 7 mg/dl, but no correlation was found between CFU-E inhibition and hematocrit. Likewise, inhibition of CFU-GM and CFU-Meg growth was not associated with leukopenia or thrombocytopenia, respectively. Sera from patients undergoing chronic intermittent hemodialysis were assayed before and after one hemodialysis session. In each case, the degree of inhibition of CFU-E and CFU-GM growth decreased after hemodialysis, but improvement in CFU-Meg growth was more variable. These data indicate that uremic sera contain dialyzable inhibitors of in vitro hematopoiesis which increase with the severity of renal dysfunction, but these inhibitors lack specificity. If uremic inhibitors of erythropoiesis are of pathophysiologic significance in vivo, there must be unrecognized repair mechanisms for granulopoiesis and thrombopoiesis.     

Polyamines in the anemia of end-stage renal disease

The improvement in the anemia in patients with end-stage renal disease (ESRD) on continuous ambulatory peritoneal dialysis (CAPD) suggests that dialyzable substances present in the sera of uremic patients either inhibit erythropoiesis directly or inactivate erythropoietin (EPO). In the present study predialysis sera from patients with ESRD inhibited erythroid colony (CFU-E) (N = 10) formation to a significantly (P less than 0.01) greater degree than granulocyte-macrophage (CFU-GM) (N = 7) colony formation in mouse bone marrow (MBM) cultures. The polyamines spermine (SP) (18 to 560 nm/ml) and spermidine (SD) (4 to 648 nm/ml) exerted a more significant (P less than 0.05) inhibition of CFU-E (N greater than or equal to 5) than that of CFU-GM (N greater than or equal to 5) growth. Concentrations of 0.80, 1.0, and 1.5 nm/ml of putrescine (PU) were 92%, 85%, and 77% of erythroid colony (CFU-E) controls (N = 4) and 104%, 130%, and 127% of CFU-GM controls (N = 4). Putrescine (PU) at 1.5 nm/ml also produced a significant (P less than 0.05) inhibition of CFU-E, whereas CFU-GM were stimulated by PU. These data suggest that predialysis sera from uremic patients, as well as SP, SD, and PU, are selectively more inhibitory to CFU-E than CFU-GM growth. The immunoreactivity of EPO was not significantly changed when it was coincubated with SP, SD and PU and measured by radioimmunoassay. PU was found to inhibit noncompetitively the bioactivity of EPO in a CFU-E assay. These data support the hypothesis that polyamines may be important uremic toxins in the anemia of ESRD.     

Bone Sialoprotein (BSP) is a Crucial Factor for the Expression of Osteoblastic Phenotypes of Bone Marrow Cells Cultured on Type I Collagen Matrix

In this study, we demonstrated that type I collagen matrix induced the expression of osteoblastic phenotypes of bone marrow cells, and that antibone sialoprotein (BSP) monoclonal antibody suppressed the expression of these phenotypes. On the other hand, BSP accelerated the expression of osteoblastic phenotypes of bone marrow cells. The adherent bone marrow cells were harvested from rat femur and cultured on type I collagen matrix gels in medium containing 15% fetal calf serum, neither β-glycerophosphate nor glucocorticoid. Cells showed osteoblastic phenotypes (high alkaline phosphatase activity, osteocalcin synthesis, and responsiveness against parathyroid hormone) on collagen matrix gels at week 3 after the inoculation, and simultaneously, BSP was detected in the conditioned medium by Western blotting using an anti-BSP monoclonal antibody. However, cells in the conventional culture dishes did not show osteoblastic phenotypes during the experimental period. To investigate the physiological function of BSP in osteoblastic differentiation, bone marrow cells were cultured on collagen matrix with an anti-BSP monoclonal antibody for 3 weeks. This treatment suppressed the expression of the osteoblastic phenotypes, and the effect of the antibody was abolished by the addition of bovine bone BSP. Furthermore, bovine bone BSP stimulated the expression of osteoblastic phenotypes of bone marrow cells. Our results indicate that BSP plays a crucial role in the expression of osteoblastic phenotypes of bone marrow cells. Received: 17 February 1999 / Accepted: 14 December 1999     

Inhibition of erythropoiesis in chronic renal failure: the role of parathyroid hormone

Sera from 20 anemic patients with chronic renal failure (CFR) were studied for their effect on bone marrow in vitro erythroid colony formation (CFUE) and the observations correlated with parathyroid hormone (PTH) and ionized calcium levels in the patients' sera. Results demonstrated that 17 out of 20 patients' sera significantly inhibited in vitro erythropoiesis by 47% to 97%. No significant elevation in ionized calcium was found in 16 of the patients tested. Furthermore, assay of PTH levels in these patients revealed that 9 out of 20 had elevated levels of PTH. No correlation was found between PTH serum levels and the degree of in vitro inhibition of erythropoiesis (CFUE) by the patients' sera. Addition of up to 2,000 pg/mL (far above the patients' levels) of exogenous N-terminal or C-terminal PTH with in vitro bone marrow cultures resulted in no inhibitory effect on CFUE. It is concluded that the circulating inhibitor of erythropoiesis which has been shown to exist in the sera of this particular group of patients with CRF, is not PTH.     

Hemodialysis and continuous ambulatory peritoneal dialysis effects on erythropoiesis in renal failure

Parameters of erythropoiesis were studied in patients with endstage renal disease established on continuous ambulatory peritoneal dialysis (CAPD) and regular hemodialysis treatment (RDT). Serum erythropoietin was measured by radioimmunoassay, and erythroid progenitor cell (CFU-E) formation was assayed in fetal mouse liver cultures. Serum erythropoietin concentrations in both CAPD (35.3 +/- 4.0 mU/ml) and RDT (31.9 +/- 1.9 mU/ml) patients were significantly higher (P less than 0.01) than normal values (23.1 +/- 1.0 mU/ml). The serum erythropoietin concentration did not correlate with either hematocrit or inhibition of CFU-E formation in either group of dialysis patients. In both CAPD and RDT patients the hematocrit correlated significantly (P less than 0.001) with the degree of serum inhibition of CFU-E formation. CFU-E formation decreased from 74.5 +/- 2.5 to 62.5 +/- 3.5% of control with increasing concentrations of uremic serum in cell cultures from 5 to 20%. In RDT patients a single hemodialysis produced a decrease in the mean serum erythropoietin concentration from 31.8 +/- 2.1 to 27.4 +/- 1.8 mU/ml (P less than 0.01) but no significant change in CFU-E formation. In conclusion, although serum immunoreactive erythropoietin levels are elevated above the normal range in dialysis patients, the response remains inadequate for the severity of the anemia, and it is the degree of serum inhibition of erythropoiesis in both CAPD and RDT patients which correlates with and possibly determines the degree of anemia.     

In-vitro growth patterns of bone marrow erythroid progenitors from patients with post-renal transplant erythrocytosis

Background: Erythrocytosis is relatively common after renal transplantation and is associated with a higher risk of thromboembolism. Its aetiology is unclear and there is still debate about the most frequently suggested causes. The culture in vitro of erythroid progenitors is regarded as a useful tool for the differential diagnosis of patients with unclear erythrocytosis. We studied the growth in vitro of bone marrow erythroid progenitor from renal transplant patients with erythrocytosis and controls without erythrocytosis. Subjects and methods: Thirteen renal transplant patients with erythrocytosis and 12 normocythaemic renal transplant controls were studied. The clinical characteristics of these patients were evaluated and serum erythropoietin (Epo) and ferritin levels were determined. Bone marrow erythroid progenitors were cultured both with and without the addition of Epo to the medium. Results: Samples from six polycythaemic patients and seven controls did not grow spontaneously in the absence of exogenous Epo. Three cases of post-transplant erythrocytosis and five controls produced CFU-E, but not BFU-3. A few CFU-E and BFU-E grew spontaneously in samples from four polycythaemic patients but not in samples from the controls. Addition of 1 unit per millilitre Epo caused similar increases in the number of colonies in both polycythaemic patients and controls. Of the nine patients eligible for follow-up, all four with spontaneous growth of BFU-E had transient erythrocytosis and four of the five patients with no spontaneous growth or spontaneous growth of CFU-E only had persistent erythrocytosis requiring treatment with ACE inhibitors. Conclusions: Pathophysiology of post-transplant erythrocytes is heterogenous. In one-third of the patients, there was unexpected, spontaneous and transient growth of BFU-E which was not predictive of permanent erythrocytosis. The results of stem-cell studies suggest that in these cases erythrocytosis may be caused by defective regulation of erythroid progenitor proliferation, possibly due to particular cellular interactions or the effect of cyclosporin on erythropoiesis.     

Adrenergic modulation of erythropoiesis following severe injury is mediated through bone marrow stroma

BACKGROUND: Severe trauma leads to hematopoietic failure and bone marrow (BM) dysfunction that manifests clinically as a persistent anemia and leukopenia. The impact of severe trauma and its associated hyperadrenergic state on erythropoiesis has not been described. The aim of this study was to demonstrate the effects of adrenergic agonists and antagonists on erythropoiesis, both in normal bone marrow mononuclear cells (BMNC) and stroma-depleted BM. METHODS: Urine epinephrine (EPI) and norepinephrine (NE) excretion from severely injured patients was assessed via enzyme-linked immunoadsorbent assay (ELISA). Erythropoiesis was assessed by the growth of erythroid progenitors-erythroid burst forming units and colony forming units (BFU-E and CFU-E)-in normal human BM in the presence of adrenergic agonists and antagonists at varying concentrations. Parallel cultures, depleted of BM stroma by passage through nylon wool columns, were compared. RESULTS: Urine NE excretion was elevated in all samples from days 1 to 10 following injury (average 139 +/- 59 mcg/day vs. control 35 +/- 9 mcg/day). In vitro doses of NE, EPI, and isoproterenol (ISO) exerted a stimulatory effect on BFU-E colony growth in BMNCs (expressed as percentage of control: 324 +/- 30, 272 +/- 16, 212 +/- 95, vs. 100%), but had no effect on stroma-depleted BM. CONCLUSIONS: There is a substantial and persistent hyperadrenergic state seen after severe injury that may last for up to a week. Adrenergic agonists have a clear stimulatory effect on the growth of primitive erythroid precursors in normal BM. The adrenergic stimulus appears to be mediated via BM stroma.     

Erythropoiesis and hemolysis in hemodialysis patients

In order to investigate the pathogenesis of renal anemia, erythroid marrow cellularity, factors affecting erythropoiesis and hemolysis, hemolysis starting point by Parpart method and red cell life-span were studied in 21 patients undergoing hemodialysis (HD). Mean value of serum erythropoietin level (EPO) in HD patients was 28.4 mU/ml, which value was nearly equal to that in healthy subjects. Total erythroblast count was higher than normal up to 25.2% in HD patients with Ht below 25% (A group), on the other hand, in HD patients with Ht above 25% (B group) it was 21 6%, nearly equal to normal. Total erythroblast counts positively correlated to EPO level, but did not correlate to ribonuclease, aluminium and parathyroid hormone. Red cell life-span was 23.4 days in A group, and it was 19.8 days in B group Hemolysis starting point was observed at 0.61% NaCl in B group, and at 0.56% in A group. Hemolysis starting point negatively correlated to red cell life-span, but did not correlate to BUN, serum creatinine and serum guanidino compound. Hb level negatively correlated to nuclear cell counts of bone marrow in HD patients, and positively correlated to hemolysis starting point. These results suggested that erythroblast count was controlled by both erythropoietin and hemoglobin levels in HD patients. Hemoglobin level in HD patients was maintained by balance of counteracting factors between erythropoiesis and hemolysis.     

Hyperparathyroidism and anemia in uremic subjects: a combined therapeutic approach

Several factors are involved in conditioning renal anemia, and a critical role is attributed to parathyroid hormone (PTH) oversecretion, which has some direct effects on endogenous erythropoietin (EPO) synthesis, bone marrow erythroid progenitors, and red cell survival. Indirect effects are mainly based on the induction of bone marrow fibrosis. Indirect evidence of the role of PTH is based on the observation that parathyroidectomy, when performed in uremic patients, is often followed by restoration of the hematocrit. The interpretations of such positive results are based on the observation of the restored bone marrow space after operation and also in a rise of immunoreactive EPO serum concentrations observed in the first weeks after gland removal. Another field of clinical interest is the possible beneficial effects of vitamin D therapy in controlling PTH secretion, which in turn determines an improvement of anemia of uremic subjects. Several uncontrolled studies confirmed this possibility, indicating that patients who respond to calcitriol or its analogs also show an increase of their hemoglobin levels. Thus, a combined therapeutic approach to PTH oversecretion and anemia is possible by intravenous calcitriol or parathyroidectomy pointing to the possible reversibility of bone marrow fibrosis, which is a common feature of secondary hyperparathyroidism. The increased sensitivity to EPO therapy can also induce a successful reduction of its dosage, thus allowing an interesting reduction of costs.