Scientists at the U.S. Food and Drug Administration (FDA) identified chemical reactions inside the red blood cells from a transgenic mouse model with sickle cell disease (SCD) that help trigger the release of microparticles (MPs), small packets of the highly reactive form of SCD hemoglobin (Hb), called HbS, that contribute to some of the symptoms of this disease.

The FDA scientists also reported for the first time a link between the abnormal chemical environment of SCD red blood cells and the destruction of HbS by a cellular structure called the proteasome.

The findings of the study suggest new strategies using hydroxyurea (an FDA approved drug), that would lessen the intensity of the symptoms of this disease by blocking specific abnormal chemical reactions that occur in the red blood cells of SCD patients.

SCD is caused by genetic mutations in Hb, the iron-containing protein inside red blood cells that bind oxygen molecules and carry them to tissues. HbS distorts the cells into sickle-shaped structures that inhibit their movement through the blood vessels. This disrupts delivery of blood and oxygen to tissues and organs, causing pain and damage to tissues and organs. Eventually the sickled cells break apart, releasing HbS into the bloodstream. This free HbS, which is highly chemically reactive, damages blood vessels, contributing to the symptoms of SCD.

The abnormal chemical environment of the cell and its effects on HbS has previously been known to be linked to production of MPs. However, until now the contribution of HbS to MP formation was poorly defined. Using mouse models, the FDA scientists showed for the first time that a chemical reaction called oxidation is a key early step in their formation. Specifically, the scientists showed that the iron atoms in HbS with two positive charges (Fe²⁺) undergo oxidation so each one gains a third charge (Fe³⁺). In the abnormal chemical environment of the HbS red cell, Fe³⁺ undergoes further oxidation to Fe⁴⁺. The Fe⁴⁺ form of HbS then interacts with the red cell membrane at a site called Band 3, causing the formation of the HbS -containing MPs, which bud off into the bloodstream. Once in the bloodstream, the MP can release its contents, which damage the blood vessels and reduce the flow of blood.

The study also showed for the first time that oxidation of the iron in HbS to Fe⁴⁺ enables several subsequent chemical reactions that lead to the destruction of this molecule. In one of these reactions, a key amino acid βCys93, undergoes irreversible oxidation. Next, a molecule called ubiquitin binds to some lysine amino acids. These changes to HbS label it for destruction by the proteasome, which acts as the cell’s protein shredder. This reduces the amount of the oxygen- carrying protein and contributes to symptoms of SCD.

The FDA scientists also demonstrated how the drug hydroxyurea (an FDA-approved drug), which is commonly used to reduce symptoms in SCD patients, blocked the oxidation of Fe³⁺ to Fe⁴⁺ and shielded the βCys93 target. This reduced both the binding of HbS to Band 3 that triggers MP production and the tagging of HbS for destruction in the proteasome.

Title

Hemoglobin oxidation-dependent reactions promote interactions with band 3 and oxidative changes in sickle cell-derived microparticles.

JCI Insight. 2018 Nov 2;3(21). doi: 10.1172/jci.insight.120451.

Authors

Jana S¹, Strader MB¹, Meng F¹, Hicks W¹, Kassa T¹, Tarandovskiy I¹, De Paoli S², Simak J², Heaven MR³, Belcher JD⁴, Vercellotti GM⁴, Alayash AI¹.

1 Laboratory of Biochemistry and Vascular Biology and.
2 Laboratory of Cellular Hematology, Division of Blood Components and Devices, Center for Biologics Evaluation and Research, FDA, Silver Spring, Maryland, USA.
3 Vulcan Analytical LLC, Birmingham, Alabama, USA.
4 Division of Hematology, Oncology and Transplantation, University of Minnesota Medical School, Minneapolis, Minnesota, USA.