This is a continuation of the effort to produce a mouse model of sickle anemia. During the past project years, we have succeeded in constructing a sickle cell mouse in which the predominant hemoglobin produced in the adult mouse is HbS. This model is different from others that have been made in the beta/S globin transgene lies within a YAC which contains the locus control region and the whole beta-globin gene cluster. Because the control regions of the globin genes and putative enhancers and silencers sequences are in their native context, these mice may mimic more closely to humans for testing drugs that affect HbF production. One problem in the preparation of these mice is that the yield is extremely low. This may be due to the fact that these mice turn off gamma-globin gene expression early in the embryo and the high level of HbS is not tolerated in utero.
Our aim i n the coming years is to brain sickle cell mice that survive the neonatal period using several approaches. First, we will continue to breed them in order to see if we can obtain more sickle cell mice that survive. It has been the experience of others that in spite of initial differences in obtaining sickle cell mice because of perinatal mortality, upon repeatedly breeding, mice that survive could eventually be obtained probably due to the interaction of yet undefined modifying factors. Because it is difficult to predict how long it will take to obtain longer surviving mice from continued breeding, we will also use alternative strategies. We will rescue the sickle cell mouse by breeding in transgenes that express high levels of human Hb F, preferably only in fetal life. One strategy is to use an alpha- LCR (HS-40) construct to direct gamma-globin gene expression. Previously, it has been shown that when the HS4-40 was used to direct an alpha-globin expression was high in the embryo but declined soon after birth. Another alternative is to breed in a beta-chimeric transgene in which the expression of the gamma-globin gene is directed by the beta promoter. The rescuing transgenes will be marked at both the RNA and protein levels to allow their expression to be differentiated from that of the YAC. To validate the utility of this model, the sickle cell line will be given agents that have been found to affect gamma-globin gene expression in humans. The model will also be available to other investigators to test new agents that increase fetal hemoglobin or inhibit hemoglobin S polymerization.
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