AbstractThalassaemia is one of the most prevalent inherited haemoglobin disorders with a broad clinical spectrum. The diversity in symptoms cannot be explained purely by patients’ genetic background. Several factors have been recognised to contribute to disease severity, and this work has set out to undertake a comprehensive study of some of these contributors in HbE/β-thalassaemia patients from Thailand.
Extracellular vesicles (EVs) are one factor that may indicate and/or contribute to disease severity. The levels of EVs observed in the plasma of thalassaemic patients are reported to be, on average, four times higher than in healthy controls. Moreover, it is well established that these EVs are associated with an increase in clinically significant procoagulant activity. The work presented in this thesis has investigated the EVs produced from both in vitro and in vivo origins derived from thalassaemic patients and the quantification and characterisation of their protein constituents.
An erythroid culture system was developed for growing thalassaemic patient progenitor cells into reticulocytes. Using this culture system, thalassaemic and age-matched control reticulocytes were produced, EVs subsequently isolated, and their proteomic profiles assessed using quantitative mass spectrometry. This thesis reports the first in vitro reticulocyte EV proteome derived from adults with β-thalassaemia.
Furthermore, in vivo EVs isolated from plasma of thalassaemic patients were investigated using quantitative proteomic analysis. Amongst 21 proteins identified with significantly altered abundances in HbE/β-thalassaemia EVs, haptoglobin, hemopexin, and cathepsin S had the potential to be used as clinical biomarkers. A pilot clinical follow-up trial was designed to examine their application, with promising results. All three protein biomarkers had significantly altered levels in groups of patients with different severity of symptoms, i.e., patients with transfusion-dependent thalassaemia, non-transfusion dependent thalassaemia, thalassaemic carriers, and healthy controls. Additionally, both haptoglobin and hemopexin showed a significant correlation to other haemolytic blood parameters. These findings suggested haptoglobin and hemopexin have utility in thalassaemic patients as a tool for clinical monitoring and as indicators of blood transfusion requirements.
Therefore, in summary, the investigation and characterisation of the extracellular vesicles generated by HbE/β-thalassaemic patients were successfully completed, protein biomarkers were identified, and their potential clinical application explored.
|Date of Award||28 Nov 2019|
|Supervisor||Ash M Toye (Supervisor) & Vanja Karamatic Crew (Supervisor)|