We found a match
Your institution may have rights to this item. Sign in to continue.
- Title
β-Globin <i>Sleeping Beauty</i> Transposon Reduces Red Blood Cell Sickling in a Patient-Derived CD34<sup>+</sup>-Based In Vitro Model.
- Authors
Sjeklocha, Lucas M.; Wong, Phillip Y.-P.; Belcher, John D.; Vercellotti, Gregory M.; Steer, Clifford J.
- Abstract
The ultimate goal of gene therapy for sickle cell anemia (SCA) is an improved phenotype for the patient. In this study, we utilized bone marrow from a sickle cell patient as a model of disease in an in vitro setting for the hyperactive Sleeping Beauty transposon gene therapy system. We demonstrated that mature sickle red blood cells containing hemoglobin-S and sickling in response to metabisulfite can be generated in vitro from SCA bone marrow. These cells showed the characteristic morphology and kinetics of hemoglobin-S polymerization, which we quantified using video microscopy and imaging cytometry. Using video assessment, we showed that delivery of an IHK-βT87Q antisickling globin gene by Sleeping Beauty via nucleofection improves metrics of sickling, decreasing percent sickled from 53.2 ± 2.2% to 43.9 ± 2.0%, increasing the median time to sickling from 8.5 to 9.6 min and decreasing the maximum rate of sickling from 2.3 x 10-3 sickling cells/total cells/sec in controls to 1.26 x 10-3 sickling cells/total cells/sec in the IHK-βT87Q-globin group (p < 0.001). Using imaging cytometry, the percentage of elongated sickled cells decreased from 34.8 ± 4.5% to 29.5 ± 3.0% in control versus treated (p < 0.05). These results support the potential use of Sleeping Beauty as a clinical gene therapy vector and provide a useful tool for studying sickle red blood cells in vitro.
- Subjects
SICKLE cell anemia treatment; GLOBIN; TRANSPOSONS; ERYTHROCYTES; MATHEMATICAL models; GENE therapy; HEMOGLOBINS
- Publication
PLoS ONE, 2013, Vol 8, Issue 11, p1
- ISSN
1932-6203
- Publication type
Article
- DOI
10.1371/journal.pone.0080403