[Increased in vitro and ex vivo proliferation of erythroid precursors induced by calcitriol in chronic renal failure. Synergistic effect with rHuEPO]. / Aumentata proliferazione dei precursori eritroidi indotta in vitro ed ex vivo dal calcitriolo in corso di uremia cronica.Effetto sinergico con rHuEPO.

BACKGROUND: Calcitriol (C) improves anemia in chronic renal failure. This improvement may be related to the suppression of iPTH release, but also to a direct effect on erythropoiesis. MATERIALS AND METHODS: In order to verify this hypothesis, 33 patients with chronic renal failure were enrolled: 24 were undergoing hemodialysis, 9 managed conservatively. All patients were free from other chronic or hematological disease, had a negative DFO test and aluminum levels below 20 mcg/l. iPTH range was 250-480 pg/l. None had yet been treated with C. In vitro study- Samples were drawn for a basal erythroid precursors (Burst Forming Unit-Erythroid BFU-E) study. After mononuclear cells were isolated by centrifugation with Ficoll-Hypaque, they were incubated for 15 days with rHuEPO 3U/ml (A), rHuEPO 3U/l + C 30 pg (B), rHuEPO 3U/ml + C 300 pg (C), rHuEPO 30 U/ml + C 300 pg (D) was performed. Ex vivo study- After the basal evaluation, 10 pts on dialysis were treated with C (Calcijex-Abbott) 1 g three times a week. BFU-E studies were performed after 1,2 and 4 months. RESULTS: In vitro, culture B showed an increased BFU-E proliferation vs A (41+/- 23 vs 27+/-15, p less than 0.02); in C and D cultures proliferation was 61+/-31 and 78+/- 42 respectively, p less than 0.01 vs A. There was no difference among pts with renal failure and pts treated conservatively. During the in vivo study all cultures showed a progressive proliferation increase, without a plateau level (basal, after 1, 2, 4 months respectively): in A: 17+/-8, 22+/-13, 30.9+/-14.9, 41.4+/-20; in B: 27.3+/-15, 35.6+/-20, 45.5+/-21, 57+/-26; in C: 48.2+/-20.6, 63.7+/-32, 75.7+/-37, 83+/-40; in D: 72+/-24, 91+/-42, 106+/-42, 110+/-42.3 (always p less than 0.001). The hematocrit and hemoglobin increase was constant but not significant. The iPTH decrease was not related to BFU-E proliferation. CONCLUSIONS: In chronic uremia C has a direct effect on erythroid precursor proliferation, both in vitro and ex vivo, with a sinergystic effect with rHuEPO. This effect is not related to iPTH suppression. C may be a useful adjuvant therapy for rHuEPO treatment.

[Calcitriol increases burst forming unit-erythroid (BFU-E) in vitro proliferation in chronic uremia. Synergic effect with DNA recombinant erythropoietin (rHu-Epo)]. / Aumentata proliferazione dei precursori eritroidi burst forming unit-erythroid (BFU-E) indotta in vitro dal calcitriolo nell'uremia cronica. Effetto sinergico con l'eritropoietina da DNA ricombinante (rHu-Epo).

BACKGROUND: It has been suggested that calcitriol (C) could improve anemia in chronic renal failure. However it remains debatable whether vitamin D has a specific effect on erythropoiesis, or it acts via suppression of hyperparathyroidism. METHODS: We enrolled 29 patients with chronic renal failure, free from malignancies, iron deficiency or other chronic or hematological diseases. Aluminium accumulation was also excluded by DFO test. 22 were on hemodialysis and 7 on conservative management, creatinine clearance ranging 22-48 ml/min. Their mean age was 62+/-28 years and duration of renal disease was 98+/-51 months. No patient under-went rHu-Epo or Vitamin D treatment. 4 subjects were enrolled as controls. Samples of peripheral blood were drawn for the Burst Forming Unit-Erythroid (BFU-E) assay. After isolation of mononuclear cells by density gradient centrifugation with Fycoll-Hypaque, a 15-day incubation was set up with four different conditions: a) adding standard dose, 3 U/ml, of r-HuEpo (Dompè Biotec), standard colture; b) combined doses of r-HuEpo, 3 U/ml, and C (Abbott), 30 pg; c) standard dose, 3 U/ml, of r-HuEpo and high dose, 300 pg, of C; and lastly d) combined high doses of r-HuEpo, 30 U/ml, and C, 300 pg. RESULTS: In the b colture (combined low doses) a higher BFU-E proliferation was found vs standard (a) colture (33.2+/-15.5 vs 17.1+/-9.2, p<0.02); interestingly, either in the c and d studies BFU-E showed an even higher proliferation (52.3+/-24 and 86.3+/-37.8 respectively, p<0.01 vs a). No difference was found when evaluating separately preterminal and hemodialysis patients. In control subjects only colture d showed an increased BFU-E proliferation. CONCLUSIONS: C has a direct effect on erythroid precursors proliferation in vitro, acting in a sinergystic manner with rHuEpo. C may be useful as adjuvant therapy for renal anemia.

Desferrioxamine improves burst-forming unit-erythroid (BFU-E) proliferation in haemodialysis patients.

BACKGROUND: In chronic renal failure, desferrioxamine (DFO) may improve erythropoiesis independent from its aluminium (Al) chelating effect. The mechanism of this action is still unknown. METHODS: To verify whether DFO influences proliferation of erythropoietic precursors, we studied 10 patients on chronic haemodialysis, free from malignancies or other haematological diseases, iron deficiency, bone marrow fibrosis, and Al toxicity. Al accumulation was excluded by the DFO test. Peripheral blood samples were drawn for basal burst-forming unit erythroid (BFU E) assay. Mononuclear cells were isolated by density gradient centrifugation with Ficoll Hypaque, and incubated for 15 days with three different experimental conditions: (a) low-dose recombinant human erythropoietin (rHuEpo) (3 U/ml); (b) high dose rHuEpo, (30 U/ml); (c) both DFO (167 microg/ml) and rHuEpo (3 U/ml). We determined TIBC, transferrin, ferritin, reticulocytes, hypochromic erythrocytes, soluble transferrin receptor (sTR), haemoglobin (Hb), and haematocrit (Hct) at baseline and then every 14 days. Patients received 5 mg/kg DFO infused during the last hour of each dialysis session for 6 weeks; six patients remained in the study for an additional 6 more weeks. BFU E assays were set up after 6 and 12 weeks of DFO therapy. RESULTS: At baseline DFO had small effect on BFU E proliferation (33.9+/-25 vs 30.4+/-25.9) and high-dose rHuEpo had a significant effect (45.15+/-27 vs 30.4+/-25.9, P<0.01). After 6 weeks of DFO therapy a significant increase in BFU E proliferation was observed in all culture conditions (78.25+/-32 vs 30.45+/-25.9 standard culture, P<0.01; 110.9+/-30 vs 45.15+/-27 high dose rHuEpo, P<0.01; 98.75+/-32 vs 45.15+/-27 DFO culture, P<0.01). Moreover, the increase in BFU E proliferation was significant greater with DFO culture than standard culture (P<0.01). The same trend was found at the third BFU E assay, performed in only six patients, when all culture conditions showed a further increase of erythroid precursor proliferation. However, the DFO culture was not significantly greater than the standard culture, while the high-dose rHuEpo was significantly greater than the DFO culture. Patients in group I (n=10), had a significant increase in reticulocytes (1.5+/-0.6 vs 1.72+/-0.3, P<0.01) and of hypochromic erythrocytes (HE) (5.6+/-5.1 vs 14.4+/-12.7, P<0.01), while sTR, Epo, Hb, and Hct were only minimally increased. Ferritin decreased significantly (448+/-224 vs 196+/-215, P<0.01) and TIBC and transferrin were unchanged. CONCLUSIONS: Thus DFO increases erythroid activity by BFU E proliferation and increases reticulocytes in haemodialysis patients. Such an effect may be related to increased iron utilization. DFO may be a useful tool for anaemic patients with good iron stores and without Al overload.