A comprehensive study of immune function and immunophenotyping of white blood cells from ß-thalassaemia/HbE patients on hydroxyurea supports the safety of the drug.

Hydroxyurea (HU) (hydroxycarbamide) is used as a therapeutic option in ß-thalassaemia to increase fetal haemoglobin, which results in a reduced requirement for blood transfusion. However, a potential serious adverse effect of HU is neutropenia. Abnormal neutrophil maturation and function in ß-thalassaemia/HbE patients are well documented. This raises questions about the effect of the drug with regards to the immune response these patients. This study investigated the effects of HU treatment on both innate and adaptive immunity in a cross-sectional study of 28 ß-thalassaemia/HbE patients who had received HU treatment (BE+HU) as compared with 22 ß-thalassaemia/HbE patients who had not received HU (BE-HU) and 26 normal subjects. The expression of PU.1 and C/EBPß, transcription factors, which are associated with neutrophil maturation, was significantly reduced in BE+HU patients as compared with BE-HU patients and normal subjects. Interestingly, C3bR expression on neutrophils and their oxidative burst activity in BE+HU were restored to close to normal levels when compared with BE-HU. There was no observed effect of HU on monocytes, myeloid derived suppressor cells (both granulocytic and monocytic subsets), CD4+ T cells, CD8+ T cells, complement levels and serum immunoglobulin levels in this study. The full immunophenotyping analysis in this study indicates that HU therapy in ß-thalassaemia/HbE patients does not significantly compromise the immune response.

Impaired neutrophil extracellular trap formation in ß-thalassaemia/HbE.

Neutrophil dysfunction contributes to a high susceptibility to severe bacterial infection which is a leading cause of morbidity and mortality in ß-thalassaemia/HbE, especially in splenectomised patients. This study demonstrated another abnormality of neutrophil function, namely neutrophil extracellular trap (NET) formation in splenectomised and non-splenectomised ß-thalassaemia/HbE patients who had iron overload. A classification system of morphological NET formation using confocal microscopy was developed, and samples were categorized into early and late phases which were subdivided into web-like and non-web structures. At baseline, neutrophils from non-splenectomised patients (58 ± 4%) and splenectomised patients (65 ± 3%) had higher early phase NETs than those from normal subjects (33 ± 1%). As a mimic of iron overload and infection, haemin/PMA/LPS treatment led to a significant reduction of early NETs and an increase of late NETs in neutrophils from normal and non-splenectomised patients. Interestingly, neutrophils from splenectomised patients had impaired development of late NETs. This suggests that during infection bacteria might not be trapped and may spread from the site of infection resulting in higher susceptibility to severe bacterial infection in splenectomised patients.

Increased ferritin levels in non-transfusion-dependent ß°-thalassaemia/HbE are associated with reduced CXCR2 expression and neutrophil migration.

Severe bacterial infection is a major complication causing morbidity and mortality in ß-thalassaemia/HbE patients. Innate immunity constitutes the first line of defence against bacterial infection. This study aimed to comprehensively investigate the innate immune phenotype and function related to factors predisposing to infection in non-transfusion-dependent (NTD) ß°-thalassaemia/HbE patients. Twenty-six patients and 17 healthy subjects were recruited to determine complement activity (C3, C4, mannose-binding lectin and CH50) and surface receptor expression including markers of phagocytosis (CD11b, CD16 and C3bR), inflammation (C5aR) and migration (CD11b, CXCR1 and CXCR2) on neutrophils and monocytes. In addition, phagocytosis and oxidative burst activity of neutrophils and monocytes against Escherichia coli and neutrophil migration were examined. Decreased C3 and surface expression of CD11b and C3bR on neutrophils were found in patients. However, phagocytosis of neutrophils in patients was still in the normal range. Interestingly, patients displayed a significant reduction of surface expression of CXCR2 [1705 ± 217 mean fluorescent intensity (MFI)] on neutrophils, leading to impaired neutrophil migration (9·2 ± 7·7%) when compared to neutrophils from healthy subjects (2261 ± 627 MFI and 27·8 ± 9% respectively). Moreover, surface expression of CXCR2 on neutrophils was associated with splenectomy status, serum ferritin and haemoglobin levels. Therefore, impaired neutrophil migration could contribute to the increased susceptibility to infection seen in NTD ß°-thalassaemia/HbE patients.