TRANSGENIC MICE CONTAINING HUMAN HEMOGLOBIN S

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While the molecular and cellular properties of hemoglobin S and sickle cells have been studied in detail, the processes leading to vaso- occlusion, painful crisis and other pathophysiological aspects of sickle cell disease remain poorly understood. The ultimate aim of this project is to produce a mouse model of sickle cell disease, which would be of great value for in vivo studies of acute and chronic vaso-occlusive processes, and for developing therapies. The transgenic models that we an others have produced to date generally resemble sickle cell trait, but do not display many of the severe symptoms of homozygous sickle cell disease. One immediate aim is to further characterize the transgenic mouse models we have produced so far, and to breed together different transgenic lines to increase the expression of Hb S (or the variant Hb SAD) and potentially increase the phenotypic effects. However, the success of transgenic approach appears to be limited by the endogenous mouse globins, which combine with human globin chains and dilute the human Hb S. The technology now exists to produce new mouse models in which endogenous hemoglobin is entirely replaced by human Hb S. For this purpose, we will produce targeted mutations in the murine alpha- and beta-globin loci through homologous recombination in embryonic stem (ES) cells, deleting the mouse adult alpha- and beta-globin genes. these mutant globin loci will be introduced into the mouse germ line, where they will be combined (by breeding) with human alpha and human betaS globin transgenes. In an alternative strategy, we will directly replace the mouse adult beta-globin genes with the human betaS gene, and the mouse adult alpha globin genes with the two human adult alpha globin genes, through gene targeting in ES cells. The resulting mice will be characterized to determine the extent to which replacement of mouse Hb with human Hb S results in a model of sickle cell disease.