Molecular, biochemical and biophysical aspects of sickle cell disease (SCD) have been studied extensively by researchers from various fields in the past 20 years. However, factors that initiate sickling in vivo are not well understood, and the relationship between sickling and the pathophysiology of SCD is unclear. In addition, no safe, effective means of preventing sickling or reversing already sickled cells in vivo, has been found, and the development and testing of antisickling agents is complicated by difficulties involved with clinical trials. One of the main constraints in studying SCD has been the lack of animal models. Animal models for SCD would be extremely helpful to study the initiation of vasoocclusive crises, pathological consequences of deformed cells, and the effects of antisickling agents in vivo. Rapid advances in molecular biology and transgenic technology have made possible the long-pursued goal of an animal model for SCD. Although numerous applications can be considered for experimental use of transgenic mice that produce Hb S (Tg-Hb S) mice, we will focus on four specific aims. We will first perform studies to examine basic biochemical and biophysical properties of various mixtures of mouse and human hemoglobins including measurements of solubility and gelation kinetics of Hb S in the presence of various mouse hemoglobins. These studies will provide us with important information regarding the influence of endogenous mouse hemoglobins on the polymerization of Hb S and the sickling properties of mouse RBC. Sickling properties of RBC containing various levels of human beta-s-globin and different mouse alpha- and beta-globin will be investigated under various hypoxic conditions. The size, shape and percentage of sickled cells will be analyzed quantitatively using a computer-assisted image analysis system. We will study various hematological parameters and pathological changes after exposure of Tg-Hb S mice to of hypoxic, hyperthermic and dehydration stress. Another important application of the Tg-Hb S mice is for screening and evaluation of antisickling agents, at both the cellular level and the whole body level. Since RBC from Tg-Hb S mice, with more than 50% human beta-s sickle almost completely upon deoxygenation, such animals could provide a readily available laboratory source for screening of new antisickling agents. Preliminary studies with several antisickling agents showed that the compounds inhibit sickling of Tg-Hb S mouse RBC as well human SS cells.
Showing the most recent 10 out of 140 publications
