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Exploring the role of low-density neutrophils and neutrophil extracellular traps in cerebral malaria

Attipa, Charalampos (2023) Exploring the role of low-density neutrophils and neutrophil extracellular traps in cerebral malaria, Thesis (Doctoral), Liverpool School of Tropical Medicine.

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Background: In 2022 Plasmodium falciparum and its most severe complication, cerebral malaria (CM) caused >500,000 deaths. The pathogenic mechanisms are poorly understood, however neutrophils have recently been implicated with neutrophil extracellular traps (NETs) detected in the cerebrovascular in fatal cases. Low-density neutrophils (LDNs) are associated with disease severity in P. vivax, as well as in cancer and sepsis in which they have increased NETotic ability, endothelial cytotoxicity or suppression of NK-cells. Still, the presence and function of LDNs and the role of NETs in CM have not been investigated.

Hypothesis: I hypothesise that the interactions of Plasmodium in the bone marrow (BM) leads to the formation and release of increased numbers of LDNs in circulation. The malaria induced LDNs are pro-inflammatory with increased NETotic capacity at the site of infected RBCs sequestration. The NETs formation at the site of sequestration acts as a key amplifier promoting further endothelial cell activation leading to BBB dysfunction and leak, as well as chemoattraction of other inflammatory cells which contributes to the development of CM and of a fatal outcome.

Methods: Paediatric CM patients and controls were recruited at a specialist unit in Malawi. Purified LDNs and normal-density neutrophils (NDNs) were isolated from venous blood and plasma was used to measure cytokines and NETs. C57BL/6 female mice infected with P. berghei ANKA were used to investigate the ontogeny and trajectory progression of LDNs. On Day 5 and Day 6 post-infection, the BM and blood were harvested and LDNs and NDNs were isolated from these two organs of mice. The isolated human and murine neutrophils sub-sets underwent cytomorphological analysis, transmission electron microscopy, flow cytometry and ex-vivo NETs formation.

Results: Increased LDNs were associated with CM compared to non-CM coma and healthy controls (HC). Furthermore, elevated LDNs were weakly correlated with delayed coma resolution in the survival CM cases. Unlike NDNs, the LDNs showed increased ex-vivo spontaneous NETs formation, immaturity, and decreased expression of maturity markers on flow cytometry. Similarly, the P. berghei caused the formation of mainly immature LDNs in the BM with increased mitochondria content, primary and secondary granules. The LDNs increased in both BM and peripheral blood as the infection was progressing.

Discussion: CM is associated with elevated NETs plasma concentrations and LDNs. The latter are activated neutrophils with increased NETotic capacity, carry an increased granular and mitochondrial content, and represent an accelerated and dysregulated maturation granulopoiesis process in the BM. NETs have been identified in the brain microvasculature of CM patients highlighting their potential role in BBB dysfunction and leak.

Item Type: Thesis (Doctoral)
Subjects: QU Biochemistry > Cells and Genetics > QU 300 General works
QU Biochemistry > Cells and Genetics > QU 375 Cell physiology
WC Communicable Diseases > Tropical and Parasitic Diseases > WC 680 Tropical diseases (General)
WC Communicable Diseases > Tropical and Parasitic Diseases > WC 750 Malaria
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Faculty: Department: Biological Sciences > Department of Tropical Disease Biology
Depositing User: Lynn Roberts-Maloney
Date Deposited: 08 Nov 2023 11:31
Last Modified: 08 Nov 2023 11:31


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