Transgenic Human α-Hemoglobin Stabilizing Protein Could Partially Relieve β IVS-2-654 -Thalassemia Syndrome in Model Mice

Abstract

β-Thalassemia is an anemia caused by a relative excess of α-hemoglobin (αHb) due to absent or reduced β-hemoglobin (βHb) synthesis. In this study, we explore whether the introduction of α-hemoglobin stabilizing protein (AHSP), a chaperone protein for proper folding and stabilization of free αHb in red blood cells, thus aiding hemoglobin A (HbA) assembly, could relieve the pathogenic state of red blood cells in β-thalassemia. For that, a human ahsp vector was constructed to generate transgenic human ahsp mice in a model of β^IVS-2-654-thalassemia by microinjecting the vector into fertilized eggs, resulting in the production of double heterozygous mice (h-ahsp ⁺/β^IVS-2-654+). Real-time quantitative RT-PCR and Western blot analysis confirmed AHSP expression in three h-ahsp ⁺/β^IVS-2-654+ mice. Hematologic determination showed an improvement in the red blood cell indices of these h-ahsp ⁺/β^IVS-2-654+ mice. The red blood cell count and hemoglobin level were elevated to various extents as compared with their diseased siblings. A dramatic reduction in anisocytosis in the peripheral blood of h-ahsp ⁺/β^IVS-2-654+ mice was observed (16.2 ± 4.6 vs. 30.0 ± 5.2%). Few erythroid precursors appeared in the liver sinusoids of h-ahsp ⁺/β^IVS-2-654+ mice. Splenomegaly with extramedullary hematopoiesis was also ameliorated. Significantly, serum iron concentration was remarkably reduced as compared with that of h-ahsp ⁻/β^IVS-2-654+ mice (43.2 ± 14.9 vs. 82.4 ± 12.9 μM), and iron deposition in the liver was decreased in h-ahsp ⁺/β^IVS-2-654+ mice. All these results suggested amelioration of the anemia phenotype in h-ahsp ⁺/β^IVS-2-654+ mice after introduction of the ahsp gene. We therefore propose that an ahsp transgene could provide an adjuvant method for gene therapy of β-thalassemia.