Erythroid stem cell kinetics: experimental and clinical aspects.
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Biomedical subjects
Publications and source records attributed to M C Magli.
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The number of erythroid burst-(BFU-E) and colony-forming units (CFU-E), as well as of myeloid-macrophage colony-forming units (CFU-C), has been evaluated in tibial marrow and spleen of ex-hypoxic polycythaemic mice, at sequential time intervals after the end of hypoxia. In both marrow and spleen, the kinetics of the CFU-E pool is characterized by a sharp fall from above normal to lower than normal values. BFU-E and CFU-C however rise from below normal to higher than normal levels. These results have been correlated with both the erythropoietin (Ep) and the erythropoietic activity curves. It is apparent that Ep levels largely control both the differentiation and the amplification of the CFU-E pool and it is suggested that Ep may act as a 'survival factor' at the CFU-E level and/or increase the flow of cells from BFU-E to CFU-E. The difference in response between CFU-E and BFU-E favours a clearcut distinction between these populations, whereas the similarity between the BFU-E and CFU-C response suggest a close relationship between these two cell populations. It is also of interest that the murine spleen functions as a large reservoir of erythroid microenvironment for hypoxia-induced stress erythropoiesis.
Adenosine deaminase (ADA) deficiency, an autosomal recessive inborn error of metabolism, leads to severe combined immune deficiency in man. This enzyme, although constitutively expressed in most tissues, is expressed at high level in immature T cells, and study of the pathophysiology of the disorder indicates that increased deoxyadenosine or altered methylation capacity have toxic effects on T-cell maturation. Although bone marrow transplantation can correct the immune deficiency, this therapy is associated with graft-versus-host disease and incomplete immune restoration, and so our laboratory and others have sought to develop a method of gene replacement as a possible treatment for the disease. Moreover, characterization of the complementary DNA of the human ADA gene and some of its mutants makes it possible to design gene transfer strategies. We have now subcloned a human adenosine deaminase cDNA into the retrovirus shuttle vector pZIP-SV(B), and in this way have isolated a cell line, 4.2T, which produces high titres of replication-defective retrovirus which have been used to transfer the gene for human ADA to mouse bone marrow cells. Transfer and expression of the neomycin-resistance gene (neo) and the ADA gene in murine bone marrow colony-forming units (CFU) was demonstrated by in vitro colony formation in the presence of the antibiotic G418 or 9-xylofuranosyladenine plus deoxycoformycin, respectively. Isoenzyme analysis also showed human ADA expression in the cultured mouse bone marrow.
In a recent CD-ROM, we portrayed the microstructure of the pre-implantation human embryo (Sathananthan et al., 1999), which was a multimedia production with computer colour-enhanced electron micrographs of mainly monospermic embryos. This disk portrays light and electron micrographs of over 250 tripronuclear (3PN), dispermic, human embryos during pre-implantation development, viewed in thick and thin Araldite sections, as well as appearances of whole embryos flat embedded in Araldite blocks visualized with the light microscope. The 100 figures were computerized (IBM TIFF format), edited and labelled using Adobe Photoshop 5. Some of the figures were coloured on computer. The early development of 3PN embryos overtly resembles that of normal embryos but there are important differences in their microstructure which are portrayed in this presentation. This is a multicentric study involving researchers from four IVF centres.
OBJECTIVE: To evaluate in vitro culture of human embryos in the presence or absence of a Vero cell monolayer. DESIGN: Randomized prospective analysis. PATIENTS: Seventy-five couples were randomly allocated to either an experimental study group (embryos in co-culture) or a control group. RESULTS: A significantly higher cleavage rate (92%) was obtained in the co-culture group compared with the control group (83%). In addition, a higher percentage of morphologically superior embryos was observed in the co-culture group than in the control group, although no significant difference was found between the two groups in blastocyst development or clinical outcome. CONCLUSIONS: When taking final clinical results into consideration in a routine IVF program, the conventional method provides more practicality and efficiency if transfers are performed on either day 2 or 3. However, if embryos remain in culture, co-culture systems allow for replacements or cryopreservation at the blastocyst stage.