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Objectives: The aim of this study is to analyse functional changes in murine erythropoietic tissues over 20 days post cyclophosphamide (CY) treatment. The project was focused on the capability of femoral and splenic erythropoietic responsive cells (ERC) to proliferate with human recombinant erythropoietin (rh Epo) stimulation after cytotoxic treatment.

Materials and methods: CF-1 mice were injected i.p. with CY (200 mg/Kg). Individual lots were studied at 0, 2, 5, 7, 10, 14, 17 and 20 days post cytotoxic treatment. Haematologic parameters [packed red cells (PRC) reticulocytes, white blood cells] were scored. Erythropoietic differentiation was assessed by the 59Fe uptake assay and the proliferative profiles of erythropoietic progenitors were determined by 3H-thymidine incorporation assay with several doses of rh Epo (0-250 mU/mL). Total and differential cellularities were scored in bone marrow (BM) and spleen.

Results: A drastical decrease of total nucleated BM cells was noticed at 2 days post CY, although cellularity was restored by the 7th day. Spleen, however, failed in showing significant decrease. The maintenance of PRC was achieved through a deep erythropoietic reorganization. 59Fe uptake revealed changes in erythroid differentiation in both tissues. Spleen maturative contribution to whole erythropoiesis was always less than medullary supply, except on day 10 post CY when a transient compensatory red cell contribution was noticed. Proliferative assays revealed that erythropoietic recovery in BM post CY was delayed in comparison with myelopoietic restoration. Splenic erythroid proliferative pattern correlated with differentiation data.

Conclusions: Myelopoietic and erythropoietic progenitors showed different recovery patterns post CY administration in BM and spleen. Medullary hemopoietic lineages restoration described a particular sequence: myelopoiesis restitution was previous to the erythroid one. Medullary erythropoiesis occurred without drastic changes in erythropoietin sensitivity while the spleen showed a transient dramatic increment on 10 days post CY red proliferation. Experimental data strongly suggest that erythropoietic recovery after CY insult mainly depends on microenvironmental regulations rather than on hormone titers.

Purpose: To evaluate the catalytic activity of the red cell enzyme, glutathione reductase (GR) in pregnant women with Hb AS and with Hb AA, and in a group of non-pregnant women with Hb AA, as well as the relationship of GR deficiency with Hb S.

Patients and methods: The catalytic activity was determined in presence and absence of FAD by means of a modified Long and Carson technique. 59 pregnant women with AS and 33 with AA phenotypes were studied.

Results: Differences were found in the enzyme's catalytic activity with and without FAD, both in pregnant women with Hb AS (mean values 37.17 nka/g y Hb in whites and 42.84 nkat/g HS in afro people) and in those with Hb AA, and also in non-pregnant women with Hb AA. A high frequency of GR deficiency was found in all groups due to an insufficient riboflavin supply in diet.

Conclusion: A correlation between GR deficiency and Hb S could not be demonstrated. The coefficient of activity of red cell GR shows a tendency to increase in pregnancy due to certain riboflavin deficit of diet.

Purpose: Was to establish the molecular genetics of thalassemias in Argentina.

Patients and methods: Genomic DNA was amplified by PCR and six point mutations in the beta-globin gene were investigated by Dot Blot hybridization using oligonucleotide probes. The most frequent alpha-thalassemia deletions were studied by Southern Blotting. Patients were distributed in 4 groups: a) 109 beta-thalassemic carriers; b) 15 thalassemia major patients; c) 2 thalassemia intermedia patients and d) 14 probable alpha-thalassemic carriers.

Results: The distribution of mutated alleles in the group a) was: IVS-1 nt 1: 13.76%, IVS-1 nt 6: 7.34%, IVS-1 nt 110: 23.85%, codon 39: 39.45%, IVS-2 nt 1: 3.68% e IVS-2 nt 745: 1.83%, 10.01% could not be determined with the probes used; in the group b) the allelic distribution was similar and the compound genetic genotype were predominant related to homocygous ones; in the group c): we confirmed the presence of one beta-thalassemia mutation and a alpha gene triplication (alpha alpha alpha) in the 2 patients studied. The alpha-thalassemia character was confirmed in 8 patients of the group d) (6 had -alpha 3,7/alpha alpha genotype and 2,-alpha 3,7/-alpha 3,7 genotype).

Conclusions: This study indicates that the analysis of 6 mutations in the beta-globin gene and the alpha-globin gene deletions are an effective strategy to identify thalassemias in Argentina.