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.     

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.     

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.     

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.     

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)     

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.     

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.     

Soluble Fas: a novel marker of inflammation in uremia

BACKGROUND: Inflammation has been associated with atherosclerotic cardiovascular disease (CVD) and anemia in patients with end-stage renal disease (ESRD). Recent studies have shown that serum levels of soluble Fas (sFas), an antiapoptotic and proinflammatory molecule, are elevated in patients with cardiac disease and patients with ESRD. We therefore sought to investigate serum levels of sFas in uremic patients and its correlation with known markers of inflammation, anemia and CVD. METHODS: The study included 25 ESRD patients (14 on hemodialysis, 11 on CAPD), 27 patients with chronic kidney disease (CKD; creatinine clearance <50 ml/min/1.73 m2), and 14 normal control subjects. We measured serum levels of sFas, C-reactive protein (CRP), and albumin. We also investigated the association of serum sFas levels with the presence of CVD and with erythropoietin (EPO) dosage. RESULTS: Levels of sFas were elevated in CKD and ESRD patients compared to controls. sFas levels correlated negatively with creatinine clearance. In the dialysis patients, we observed that sFas levels were higher among those with CVD. Serum levels of sFas correlated with serum levels of CRP (r=0.31; P=0.03), serum levels of albumin (r=-0.35, P=0.02), and EPO dosage (r=0.51; P=0.009). CONCLUSION: These results suggest that sFas may be a marker of inflammation in CKD and ESRD patients.     

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.     

Prostaglandin-mediated suppression of in vitro growth of erythroid progenitor cells

In vitro hematopoiesis was evaluated in 37 patients with chronic renal failure (CRF) who developed moderate to severe anemia in order to clarify the relationship between the growth of erythroid progenitor cells and CRF-associated anemia. Bone marrow cells from these patients were cultured in the presence of recombinant erythropoietin. Both early and late erythroid progenitor cells (BFU-E and CFU-E) were significantly suppressed in patients with CRF compared to those in normal controls, while myeloid progenitor cells (GM-CFC) remained normal. Suppression of CFU-E was shown to be mediated by prostaglandin(s) secreted from bone marrow adherent cells. Furthermore, the suppression of CFU-E was inversely correlated with concentrations of uremic serum or parathyroid hormone added to the assay system. These observations suggest a possibility that late erythroid progenitor cells may be preferentially suppressed by the network consisting of parathyroid hormone, bone marrow adherent cells and prostaglandin(s).     

Spermine and spermidine are non-specific inhibitors of in vitro hematopoiesis

The polyamine spermine has been reported to be the inhibitor of in vitro erythropoiesis present in uremic serum. We have employed a panel of hematopoietic colony-forming assays to evaluate the specificity of the inhibitory activity. Spermine and its precursor spermidine when added to culture inhibited mouse and human erythroid (CFU-E and BFU-E), granulocyte-macrophage, and megakaryocyte colony growth in a non-specific, dose-dependent fashion. Erythroid and non-erythroid colony growth were equally sensitive to spermine- and spermidine-induced inhibition. Increasing concentrations in culture of erythropoietin and mitogen-stimulated leukocyte-conditioned medium, a source of colony-stimulating activity, failed to overcome the in vitro inhibition. Although anemia is characteristic of chronic renal failure (CRF), leukopenia and thrombocytopenia are not. Therefore, we conclude that the non-specific inhibitory activity of spermine and spermidine, as defined by in vitro colony assays, is either of no pathophysiologic significance in the anemia of CRF, or else there are unrecognized repair mechanisms in vivo which maintain granulopoiesis and thrombopoiesis at normal levels.     

Anemia in new congenital adult type polycystic kidney mice

Mechanisms for the development of anemia and the effects of recombinant human erythropoietin (r-HuEPO) on hematological parameters were studied in new congenital adult type polycystic kidney (DBA/2FG-pcy) mice. The majority of DBA/2FG-pcy mice showed progressive anemia and an elevation of blood urea nitrogen, while a minority showed progressive anemia following polycythemia. Kidneys with numerous cysts in the cortex and medulla occupied virtually the entire abdominal cavity, and the combined kidney weight taken as a percentage of body weight reached 13.5% in the DBA/2FG-pcy mouse. The osmotic fragility of DBA/2FG-pcy mice erythrocytes was significantly increased compared with that of normal control mice. In addition, two-fold increases in serum EPO levels, determined by radioimmunoassay, and a decreased number of colony forming unit-erythroid (CFU-E) were observed in the DBA/2FG-pcy mice. The administration of r-HuEPO during anemia significantly increased the red blood cell count, hemoglobin concentration, hematocrit and reticulocyte percentage in a dose-dependent manner. These findings indicate that anemia in the DBA/2FG-pcy mouse is due to increased fragility of erythrocytes, a deficiency in EPO for the degree of anemia and a decreased number or a decreased response of erythroid progenitor cells. We suggest that the DBA/2FG-pcy mouse is a useful spontaneous model of chronic progressive renal failure.     

The effect of i.v. iron alone or in combination with low-dose erythropoietin in the rapid correction of anemia of chronic renal failure in the predialysis period

BACKGROUND: It is now more and more evident that anemia of predialysis chronic renal failure (CRF) should be actively treated, since long-standing anemia may cause irremediable damage to the heart. The most common form of treatment of this anemia is subcutaneous erythropoietin (EPO). iron (Fe) deficiency can also contribute to anemia in predialysis CRF, and intravenous iron (i.v. Fe) can frequently improve it. It is possible, therefore, that the combination of EPO and i.v. Fe may have an additive effect, and cause a rapid improvement in anemia with relatively small doses of EPO. PURPOSE: The purpose of this study was an initial study: to assess the ability of a combination of low-dose EPO and i.v. Fe, given weekly for 5 doses, to correct the anemia of predialysis CRF patients compared to the use of i.v. Fe alone in a randomized study. In the follow-up study: to assess the ability of the maintenance of adequate iron stores for one year to achieve and maintain the target Hct of 35% with the minimum dose of EPO. Initial study: METHOD: Ninety predialysis CRF patients (creatinine clearance 10-40 ml/min/1.73 m2 received either: Group A (45 patients): 200 mg i.v. Fe as Fe sucrose (Venofer, Vifor Int.) once per week for 5 doses in combination with 2,000 international units (IU) EPO (Eprex, Cilag-Janssen), subcutaneously given simultaneously also for 5 doses. Group B (45 patients): the same dose of i.v. Fe as in Group A but without EPO. RESULTS: The mean increase in hematocrit (Hct) and hemoglobin (Hb) by one week after the last dose was greater in group A, 4.54 +/- 2.64% (p < 0.01) and 1.37 +/- 0.84 g% (p < 0.01), respectively, than in Group B, 2.74 +/- 2.72% (p < 0.05) and 0.91 +/- 0.78 g% (p < 0.05), respectively. 80% of those in Group A had an increase in Hct of 3 vol% or more compared to 48.9% in Group B (p < 0.01). 40% of those in Group A reached the target Hct of 35% compared to 28.9% in Group B (p > 0.05). Follow-up study: During a 12-month follow-up period, enough i.v. iron was given to maintain the Hct at 35%, while keeping the serum ferritin at < 400 ug/l and % Fe Sat at < 40%. If the i.v. Fe alone was not capable of maintaining the target Hct, EPO was given in increasing doses. Eighteen patients required dialysis. Of the 72 patients who did not require dialysis, 24 (33.3%) maintained the target Hct with i.v. Fe alone, without EPO. All the remaining 48 patients (66.7%) continued to receive EPO in addition to the i.v. Fe, and 47 achieved and maintained the target Hct with a mean EPO dose of 2,979 +/- 1,326 IU/week. CONCLUSION: The combination of low-dose EPO and i.v. Fe had a rapid and additive effect on the correction of anemia in CRF predialysis patients. Maintaining adequate iron stores with i.v. Fe during a subsequent maintenance phase allowed the target Hct of 35% to be reached and maintained with low-dose EPO in two-thirds of the predialysis patients and with no EPO at all in one-third.     

Vitamin E-bonded hemodialyzer improves atherosclerosis associated with a rheological improvement of circulating red blood cells

BACKGROUND: Vitamin E-bonded hemodialyzer is known to improve oxidative stress in patients with hemodialysis. However, there is little information available as to whether or not this membrane clinically improves atherosclerosis. Furthermore, it remains unknown whether there is any effect of the membrane on rheology of circulating red blood cells. METHOD: We conducted a randomized, open-labeled, prospective control study (N = 34) for 1 year to investigate the effect of vitamin E-bonded cellulose membrane dialyzer (EE) (N = 17) on carotid atherosclerotic changes [intima-media thickness (IMT) of carotid arteries] and the viscosity, percentage of dysmorphism (%DMR) of red blood cells (RBCs) and their distribution width-standard deviation (RDW-SD), in comparison with cellulose membrane (SU) (N = 17) identical to EE without vitamin E-bonded membrane. Erythropoietin (EPO) dose used for the treatment of uremic anemia was also calculated. RESULTS: The IMT significantly decreased in the EE group, while in the SU group the IMT significantly increased. The viscosity of RBCs in hemodialysis patients (4.70 +/- 0.45 cP) was greater than that in healthy individuals (3.73 +/- 0.15 cP). EE significantly improved the viscosity (from 4.84 +/- 0.41 cP to 4.51 +/- 0.54 cP, P < 0.01), %DMR (from 2.29 +/- 2.17% to 1.90 +/- 1.49%, P < 0.01), and RDW-SD (from 54.4 +/- 7.6 fL to 49.3 +/- 5.9 fL, P < 0.01). On the contrary, these parameters all worsened in the SU group. EPO dose needed for the treatment of anemia was significantly (P < 0.05) reduced from 5383 +/- 2655 U/week to 4235 +/- 3103 U/week in the EE group. During these period, mean blood pressure, Kt/V urea, and serum beta2-microglobulin were not changed between the two groups. CONCLUSION: These findings suggest that vitamin E-bonded hemodialyzer is very useful for improving atherosclerosis from a clinical point of view. As one of the underlying mechanisms, as well as antioxidant effects, we want to address an important role of the improvement of rheology of circulating RBCs, which may also help to reduce the requirement of EPO dose in the treatment of anemia of ESRD patients.     

Integrated therapies including erythropoietin decrease the incidence of dialysis: lessons from mapping the incidence of end-stage renal disease in Japan.

BACKGROUND: Erythropoietin (EPO) has been reported to slow the decline of renal function in predialysis chronic kidney disease (CKD) patients. On the contrary, in the recent large-scale randomized controlled trial (RCT), CREATE and CHOIR, which aimed to keep a higher haemoglobin (Hb) level than former trials, the renoprotective effect of EPO was not observed. Today, the renoprotective effect of EPO has become controversial. In order to test the hypothesis that the usage of EPO in predialysis CKD patients may ameliorate the progression of renal disease, we conducted a macro-level observational study dealing with all Japanese predialysis CKD patients. METHODS: Annually since 1982, the Japanese Society for Dialysis Therapy reports the number of patients that have entered maintenance dialysis in each prefecture of Japan. Based on the 2002-2004 data, we calculated the annual incidence of end-stage renal disease (ESRD) in each of the 47 prefectures. The annual amounts paid for EPO by each prefecture, presumably corresponding to the amounts used, corrected for the estimated predialysis CKD patients, were calculated. We examined the relationship between the incidence of new dialysis and the usage of EPO in each prefecture. Furthermore, the usage of EPO was compared with that of antihypertensive agents including angiotensin converting enzyme inhibitor (ACE-I), and that of statin. RESULTS: There were prefectural differences in the annual incidence of ESRD from 2002 to 2004. We also found prefectural differences in the usage of EPO for the three consecutive years. The usage of EPO in predialysis patients was negatively correlated with the incidence of ESRD on linear and multiple regression analyses. At the same time, the usage of EPO had strong positive correlations with the usage of antihypertensive agents including ACE-I and with that of statin. CONCLUSION: Our nationwide epidemiologic study revealed that a higher use of EPO was associated with a decreased incidence of new dialysis in daily clinical practice. In addition, there were strong correlations among the usage of EPO, antihypertensive agents and statin. These data are supportive of, but do not prove, the hypothesis that EPO may be renoprotective, when used in combination with other strategies.     

Isolation and purification of erythropoiesis inhibitory activity from uremic sera

In vitro erythropoiesis inhibitory activity (EIA) was isolated and partially purified from sera obtained from a patient with chronic renal failure (CRF). EIA was purified from human sera by ammonium sulfate precipitation, column chromatography and PAGE electrophoresis, and the corresponding biological activity was assayed in erythroid colony (CFU-E) cultures. Ammonium sulfate precipitation revealed that the EIA resided in two high molecular weight fractions which yielded a single peak on G-200 Sephadex. PAGE electrophoresis demonstrated that the EIA fraction separated into 3 bands with a molecular weight range of 12,000-50,000 daltons. Antibody to EIA was prepared in rabbits, and Ouchterlony immunodiffusion assays employing the antibody material revealed that several CRF sera generated intense double precipitin bands when tested in the assay. In contrast, normal sera consistently yielded a weak precipitin band. These results suggest the presence of reduced amounts of EIA-like material in normal humans and elevated levels in CRF sera. The significance of EIA in the anemia of CRF is discussed.     

Effect of different modes of dialysis on serum erythropoietin levels in pediatric patients

The relative importance of erythropoietin (Ep) and inhibitors of erythropoiesis in the development of anemia in pediatric patients with end-stage renal disease (ESRD) was assessed in 82 patients: 41 treated with peritoneal dialysis (PD) and 41 with hemodialysis (HD). Serum Ep was determined with a sensitive radioimmunoassay. Potential serum inhibition of erythroid (CFU-E) and granulocytic (CFU-GM) progenitor cell growth was assessed using human bone marrow cell cultures. The mean Ep level for all 82 patients was 33.1±3.1 mU/ml, which was significantly higher (P<0.05) than the values obtained in 29 normal children (26.2±2.4 mU/ml). Serum Ep in the PD group (41.6±5.6 mU/ml) was significantly higher (P=0.007) than that of the HD group (24.6±2.1 mU/ml). The mean hematocrit in the PD group (25.2±0.8%) was also significantly higher (P<0.002) than that of the HD group (22.2±0.5%). The mean serum parathyroid hormone (PTH) level as measured by a mid-terminal radioimmunoassay was not significantly different (P=0.79) in the HD group (17,298±3,998 pg/ml) from that of the PD group (15,747±4,227 pg/ml). Neither serum Ep nor PTH concentration correlated with hematocrit or degree of inhibition of erythroid progenitor cell colony (CFU-E) formation in either group of dialysis patients, nor did the hematocrit correlate, with the degree of serum inhibition of CFU-E formation. The higher level of Ep in the PD group may indicate more effective removal by PD of some enzymatic substance which reduces the immunologic and biologic activities of Ep. The higher hematocrit in the PD group may be due to the higher serum level of biologically active Ep in the PD patients. The lack of any difference in PTH level in the HD and PD groups suggests that differences in PTH activity are not responsible for the higher hematocrit in PD patients. Southwest Pediatric Nephrology Study Group (SPNSG Central Office, Baylor University Medical Center at Dallas, Tex. USA): Director, Ronald J. Hogg; Statistician, Joan S. Reisch; Administrative Assistant, Kaye Green. SPNSG Centers and Clinicians participating in this study: Baylor College of Medicine, Houston, Tex, Phillip L. Berry, L. Leighton Hill, Sami A. Sanjad: Tulane University Medical Center, New Orleans, La., Frank Boineau, John E. Lewy, Radhakrishna Baliga; University of Arkansas, Little Rock, Watson Arnold, Eileen Ellis; University of Colorado Health Science Center, Denver, Gary M. Lum; University of Oklahoma Medical Center, Oklahoma City, James Wenzl, James Matson; University of Tennessee Center for the Health Sciences, Memphis, F. Bruder Stapleton, Shane Roy, III, Robert J. Wyatt, Charles McKay; University of Texas Health Science Center at Dallas, Billy S. Arant, Jr.; University of Texas Medical School, Houston, Susan B. Conley, Ann Ince; University of Texas Health Science Center at San Antonio, Michael Foulds, Fred A. McCurdy; University of Texas Medical Branch, Galveston, Ben H. Brouhard, Alok Kalia, Luther B. Travis, Lisa Hollander; University of Utah Medical Center, Salt Lake City, Eileen Brewer, Richard Siegler. Offprint requests to: R. J. Hogg, Department of Pediatrics, Baylor University Medical Center, 3500 Gaston Avenue, Dallas TX, 75246, USA     

Long-term effects of recombinant human erythropoietin on bone marrow progenitor cells

Eleven uraemic patients were treated with recombinant humanerythropoietin (rHuEpo). Seven haemodialysis patients and fourperitoneal dialysis patients received a starting dose of 80IU/kg i.v. and 40 IU/kg s.c. respectively, thrice weekly. Thenumber of burst-forming-unit erythroid (BFU-E), colony-forming-uniterythroid (CFU-E), granulocyte-monocyte (CFU-GM) and megakaryocyte(CFU-Mk) were assayed 2 weeks before (DO), and 1 (M1) and 6months (M6) after the initiation of rHuEpo treatment by meansof a commonly applied in-vitro clonal assay. All the patientsshowed the same haematopoietic response. A significant increaseof CFU-E and CFU-Mk could be observed within 1 month of treatment.At this time, no significant modification was observed in BFU-Eand CFU-GM number. At the 6th month the increase of CFU-E wasmaintained, whereas a significant fall of BFU-E, CFU-GM andCFU-Mk was observed. These results suggest that in-vivo effects of rHuEpo are notrestricted to the erythroid lineage but that erythropoietinmight also act as a co-factor of megakar-yopoiesis. In the longterm erythropoietin might induce erythroid differentiation inmultipotent progenitor cells at the expense of the non-erythroidprogenitors.     

Red blood cell membrane lipid peroxidation and resistance to erythropoietin therapy in hemodialysis patients.

BACKGROUND: Chronic hemolysis, inadequate production of erythropoietin (EPO) or an impaired response of erythroid stem cells to EPO are the main factors of anemia in end-stage renal disease (ESRD) patients. Oxidative damage of red blood cell (RBC) membrane is a well-established cause of chronic hemolysis in hemodialysis (HD) patients. Administration of high-dose recombinant human EPO (rHuEPO) fails to correct anemia in 5 to 10% HD patients although all established factors of resistance to rHuEPO therapy have been previously ruled out or corrected. PATIENTS AND METHODS: We investigated the degree of RBC membrane oxidative damage in 9 HD patients who failed to respond to maximal rHuEPO administration (more than 200 UI/Kg weekly for 4 months consecutively, group A), compared to 10 patients who showed a good response to standard rHuEPO therapy (group B) and to 10 patients who needed no treatment (group C). RBC malondialdehyde (MDA) was assumed as the index of oxidative stress in erythrocyte membrane. RESULTS: No significant difference in erythrocyte MCV and MCHC, iron status, parathyroid function, aluminum and dialysis-related blood loss was observed between patients of group A, B and C. RBC MDA, reticulocyte count, plasma-free hemoglobin (fhb) and serum lactate dehydrogenase (LDH) were significantly higher while plasma haptoglobin was significantly lower in patients of group A compared to patients of groups B and C. Moreover, a significant inverse relationship was observed between RBC MDA and either plasma hemoglobin, RBC count and hematocrit when all patients were evaluated together. CONCLUSION: In conclusion, increased oxidative damage of RBC membrane is often detectable in HD patients who fail to respond to rHuEPO administration even in the absence of all established factors of resistance to EPO. Peripheral response to rHuEPO may be normal in these patients and persistent anemia may be related to enhanced hemolysis due to oxidative stress. Oxidative damage itself may therefore be considered a factor of resistance to EPO.