Comparative Analysis of Gene Expression Changes Mediated by Individual Constituents of Hemozoin

Abstract

Plasmodium protozoa, the source of malarial infections, catabolize large quantities of hemoglobin during an intra-erythrocytic phase. During this process, free heme is detoxified through biomineralization into an insoluble heme aggregate, hemozoin (Hz). In its native state, Hz is associated with a variety of lipid peroxidation products including 4-hydroxy-2-nonenal (HNE). In the present study, gene expression profiles were used to compare responses to two of the individual components of Hz in a model macrophage cell line. LPS-stimulated RAW 264.7 cells were exposed to HNE and the synthetic form of Hz, β-hematin (BH), for 6 or 24 h. Microarray analysis identified alterations in gene expression induced by exposure to HNE and opsonized BH (fold change ≥1.8, p-value ≤0.01). Patterns of gene expression were compared to changes induced by an opsonized control latex bead challenge in LPS-stimulated cells and revealed that the BH response was predominantly phagocytic. Ingenuity Pathway Analysis demonstrated that HNE mediated a short term oxidative stress response and had a prolonged effect on the expression of genes associated with categories of ‘Cell Cycle’, ‘Cellular Assembly and Organization’, ‘DNA Replication, Recombination, and Repair’, and ‘Cellular Development’. Comparisons of expression changes caused by BH and HNE with those observed during malarial infection suggest that BH and HNE are involved in inflammatory response modulation, altered NF-κB signal transduction, extracellular matrix (ECM) degradation, and dyserythropoiesis. HNE exposure led to several significant steady-state expression changes including repressed chemokine (C-C motif) ligand 5 (Ccl5) indicative of dyserythropoiesis, and a severe matrix metalloproteinase 9 (Mmp9)/tissue inhibitor of metalloproteinase 1 (Timp1) imbalance in favor of ECM proteolysis.

Metabolic Impact of 4-Hydroxynonenal on Macrophage-Like RAW 264.7 Function and Activation

“…39 The complex biochemical interactions that occur between cellular macromolecules, such as these, and HNE result in a variety of cellular perturbations, including disruption of cytoskeletal integrity, impairment of mitochondrial respiration, inhibition of DNA, RNA and protein synthesis, as well as impairment of specialized cellular functions such as oxidative burst in professional phagocytes. 12, 18, 41 It is also thought that HNE is linked to foam cell formation and the development of atherosclerosis. 42 Inhibiting PKC would disrupt the formation of NADPH oxidase and slow down formation of ROS.…”

Matrix Metalloproteinase-9 and Haemozoin: Wedding Rings for Human Host andPlasmodium falciparumParasite in Complicated Malaria

“…Experiments by Real Time RT-PCR have shown that HZ and HZ-containing trophozoites enhanced the mRNA expression of MMP-9 [23]. HZ-dependent MMP-9 gene induction was also confirmed by additional microarray [92] and macroarray [38] studies. As a consequence, pro-MMP-9 protein expression was found to be increased [23].…”

Severity of anaemia is associated with bone marrow haemozoin in children exposed to Plasmodium falciparum

“…Recent in vitro studies have also reported that Hz stimulates the release of cytokines, chemokines and lipo‐peroxides by macrophages, and this has been suggested to contribute indirectly to anaemia and dyserythropoiesis by negatively affecting the development of erythroid cells (Awandare et al , ; Skorokhod et al , ). Moreover, in vitro studies have also shown that Hz may have a direct harmful effect on erythroid precursors by delaying the expression of erythroid markers and increasing the apoptosis of progenitor cells (Lamikanra et al , ; Schrimpe & Wright, ). In addition, peripheral blood Hz may also have a deleterious effect on circulating erythrocytes by decreasing their deformability and thus increasing their phagocytic elimination (Nuchsongsin et al , ; Skorokhod et al , ).…”

“…However, the presence and clinical impact of Hz in the bone marrow of children exposed to malaria (not necessarily currently infected with the parasite) has not yet been studied ex vivo. In spite of the evidence suggesting the role of bone marrow dysfunction in the pathogenesis of malarial anaemia (Casals-Pascual et al, 2006;Keller et al, 2009;Were et al, 2006), there is little detailed ex vivo information on the presence of malaria parasites and/or Hz in this tissue, and there is only in vitro data on the negative effects of Hz on the development of erythroid cells (Awandare et al, 2011;Lamikanra et al, 2009;Schrimpe & Wright, 2009;Skorokhod et al, 2010).…”