Hypoxia-Inducible Factor (HIF-1alpha) Has a Crucial Role in Microvascular Regulation in Transgenic/Knockout Sickle Mouse Models.

In sickle cell anemia (SCA), abnormal red cell rheology, hemolytic anemia and chronic hypoxia will impact endothelial function and result in compromised microvascular regulation. In SCA, chronic hypoxia and hypoxic events are likely to activate hypoxia-inducible factor-1alpha (HIF-1alpha), a transcriptional factor that is involved in regulation of several genes including genes for vascular endothelial growth factor (VEGF) and vasoactive molecules (e.g., inducible NOS, neuronal NOS, heme oxygenase [HO-1], endothelin). However, the status and role of this critical factor remains unexplored in SCD. We hypothesize that in SCA, chronic hypoxia caused by intravascular sickling (hemoglobin S polymerization), low hematocrit and vaso-occlusive events will induce HIF-1alpha, which will lead to activation of vasoregulatory genes and affect vascular tone mechanisms. To this end, we have explored the expression of HIF-1alpha: under normoxic conditions in transgenic-knockout sickle BERK mice (express exclusively human alpha- and beta S-globins; severe pathology) and in transgenic sickle (NY1DD) mice (mild pathology) subjected to prolonged hypoxia. In BERK mice, low hematocrit coupled with intravascular sickling will contribute to chronic hypoxia. We selected the cremaster muscle preparation used for our microcirculatory studies in BERK mice that have shown in vivo sickling, red cell adhesion and frequent episodes of transient ischemic events (Kaul et al. JCI, 2004). Western blotting of cytoplasmic and nuclear extracts of rapidly excised cremaster muscle from normoxic C57BL and BERK mice was carried out using mouse-reactive anti-HIF-1alpha antibodies. HIF-1alpha positive bands were detected at 90 kDa. Densitometric analysis confirmed average increases of 1.6-and 2.3-fold, respectively, in cytoplasmic and nuclear extracts of BERK mice ( P P P P