Between 28 and 34 weeks gestation, a switch from prodution of Hemoglobin F (alpha 2 gamma 2) to Hemoglobin A (alpha 2 beta 2) occurs. As Hgb F functions well, this normal developmental phenomenon causes great morbidity in the beta chain hemoglobinopathies and beta thalassemias. Understanding the mechanisms that produce this fetal globin switch may be of potential benefit for patients with these prevalent diseases and is considered to have general applicability to understanding of developmental gene control. This proposal seeks to elucidate the biologic processes that produce this globin switch during its natural evolution in normal premature to full-term infants and in infants of diabetic mothers (IDM), a newly identified population of infants in whom the switch does not occur. Two mechanisms associated with renewed production of Hb F in adults will be studied. Changes in the erythroid progenitor cell populations that produce globin will be ascertained in the different infants by culture of mononuclear cells in methycellulose medium with erythropoietin. DNA methylation patterns surrounding the beta globin genes will be elucidated in cord blood nucleated erythroblasts from the different infants as the switch occurs. This will be performed by double DNA digestion with methylation-sensitive and insensitive restriction enzymes and Southern blot analysis. Causal changes should precede the increase in beta globin synthesis that occurs by 36-40 weeks gestation. The role of insulin on DNA methylation may be further elucidated, as IDM have marked hyperinsulinemia. The effects of specific metabolic and hormonal abnormalities of the diabetic intrauterine environment on beta globin production will be studied in vitro in erythroid progenitor cultures under normal and hypoxic conditions, as occurs in vivo. Finally, an animal model for preventing the fetal globin switch will be established to attempt to 1) more specifically identify the physiologic factors that may modulate this process, and 2) hopefully, to prevent the globin switch from occurring after birth. This will be attempted by continuous insulin infusion into chronically catheterized fetal sheep.
