4-hydroxy-2-nonenal and Malaria

Oxidative stress plays an important role in the pathogenesis of falciparum malaria, a disease still claiming close to 1 million deaths and 200 million new cases per year. Most frequent complications are severe anemia, cerebral malaria, and immunodepression, the latter being constantly present in all forms of malaria. Complications are associated with oxidative stress and lipoperoxidation. Its final product 4-hydroxynonenal (4-HNE), a stable yet very reactive and diffusible molecule, forms covalent conjugates with proteins, DNA, and phospholipids and modulates important cell functions at very low concentrations. Since oxidative stress plays important roles in the pathogenesis of severe malaria, it appears important to explore the role of 4-HNE in two important malaria complications such as malaria anemia and malaria immunodepression where oxidative stress is considered to be involved. In this review we will summarize data about 4-HNE chemistry, its biologically relevant chemical properties, and its role as regulator of physiologic processes and as pathogenic factor. We will review studies documenting the role of 4-HNE in severe malaria with emphasis on malaria anemia and immunodepression. Data from other diseases qualify 4-HNE both as oxidative stress marker and as pathomechanistically important molecule. Further studies are needed to establish 4-HNE as accepted pathogenic factor in severe malaria.

Topics: Aldehydes; Anemia; Dendritic Cells; Erythrocytes; Humans; Lipid Peroxidation; Malaria

Malaria is a frequent parasitic infection becomes life threatening due to the disequilibrated immune responses of the host. Avid phagocytosis of malarial pigment hemozoin (HZ) and HZ-containing Plasmodium parasites incapacitates monocyte functions by bioactive lipoperoxidation products 4-hydroxynonenal (4-HNE) and hydroxyeicosatetraenoic acids (HETEs). CYP4F conjugation with 4-HNE is hypothesised to inhibit ω-hydroxylation of 15-HETE, leading to sustained monocyte dysfunction caused by 15-HETE accumulation. A combined immunochemical and mass-spectrometric approach identified 4-HNE-conjugated CYP4F11 in primary human HZ-laden and 4-HNE-treated monocytes. Six distinct 4-HNE-modified amino acid residues were revealed, of which C260 and H261 are localized in the substrate recognition site of CYP4F11. Functional consequences of enzyme modification were investigated on purified human CYP4F11. Palmitic acid, arachidonic acid, 12-HETE, and 15-HETE bound to unconjugated CYP4F11 with apparent dissociation constants of 52, 98, 38, and 73 µM, respectively, while in vitro conjugation with 4-HNE completely blocked substrate binding and enzymatic activity of CYP4F11. Gas chromatographic product profiles confirmed that unmodified CYP4F11 catalysed the ω-hydroxylation while 4-HNE-conjugated CYP4F11 did not. The 15-HETE dose dependently recapitulated the inhibition of the oxidative burst and dendritic cell differentiation by HZ. The inhibition of CYP4F11 by 4-HNE with consequent accumulation of 15-HETE is supposed to be a crucial step in immune suppression in monocytes and immune imbalance in malaria.

Topics: Cytochrome P450 Family 4; Gas Chromatography-Mass Spectrometry; Humans; Hydroxylation; Immunosuppression Therapy; Malaria; Monocytes; Protein Processing, Post-Translational

Severe malaria anemia is characterized by inhibited/altered erythropoiesis and presence of hemozoin-(HZ)-laden bone-marrow macrophages. HZ mediates peroxidation of unsaturated fatty acids and production of bioactive aldehydes such as 4-hydroxynonenal (HNE). HZ-laden human monocytes inhibited growth of cocultivated human erythroid cells and produced HNE that diffused to adjacent cells generating HNE-protein adducts. Cocultivation with HZ or treatment with low micromolar HNE inhibited growth of erythroid cells interfering with cell cycle without apoptosis. After HZ/HNE treatment, 2 critical proteins in cell-cycle regulation, p53 and p21, were increased and the retinoblastoma protein, central regulator of G₁-to-S-phase transition, was consequently hypophosphorylated, while GATA-1, master transcription factor in erythropoiesis was reduced. The resultant decreased expression of cyclin A and D2 retarded cell-cycle progression in erythroid cells and the K562 cell line. As a second major effect, HZ and HNE inhibited protein expression of crucial receptors (R): transferrinR1, stem cell factorR, interleukin-3R, and erythropoietinR. The reduced receptor expression and the impaired cell-cycle activity decreased the production of cells expressing glycophorin-A and hemoglobin. Present data confirm the inhibitory role of HZ, identify HNE as one HZ-generated inhibitory molecule and describe molecular targets of HNE in erythroid progenitors possibly involved in erythropoiesis inhibition in malaria anemia.

Topics: Aldehydes; Anemia; Biomarkers; Cell Cycle; Cell Differentiation; Cell Line; Cell Proliferation; Coculture Techniques; Colony-Forming Units Assay; Cyclin A; Cyclin D2; Cyclin-Dependent Kinase Inhibitor p21; Erythroid Cells; Erythropoiesis; GATA1 Transcription Factor; Glycophorins; Hemeproteins; Hemoglobins; Humans; Malaria; Monocytes; Receptors, Immunologic; Tumor Suppressor Protein p53

Plasmodium protozoa, the source of malarial infections, catabolize large quantities of hemoglobin during an intraerythrocytic 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, beta-hematin (BH), for 6 or 24 h. Microarray analysis identified alterations in gene expression induced by exposure to HNE and opsonized BH (fold change, > or = 1.8; p value, < or = 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-kappaB 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.

Topics: Aldehydes; Animals; Cell Line; Gene Expression; Gene Expression Profiling; Hemeproteins; Hemoglobins; Humans; Lipid Peroxidation; Macrophages; Malaria; Mice; Oxidative Stress; Phagocytosis

The most common and deadly form of the malaria parasite, Plasmodium falciparum, is responsible for 1.5-2.7 million deaths and 300-500 million acute illnesses annually [Bremen in J. Trop. Med. Hyg. 64:1-11 (2001); World Health Organization (2002)]. Hemozoin, the biomineral formed to detoxify the free heme produced during parasitic hemoglobin catabolism, has long been suspected of contributing to the pathological immunodeficiencies that occur during malarial infection. While there is a growing consensus in the literature that native hemozoin maintains immunosuppressive activity, there is considerable controversy over the reactivity of the synthetic form, beta-hematin (BH). Given the emerging importance of hemozoin in modulating a host immune response to malarial infection, a careful examination of the effects of the constitutive components of the malaria pigment on macrophage response has been made in order to clarify the understanding of this process. Herein, we present evidence that BH alone is unable to inhibit stimulation of NADPH oxidase and inducible nitric oxide synthase, the key enzymes involved in oxidative burst, and is sensitive to the microbicidal agents of these enzymes both in vitro and in vivo. Further, by systematically examining each of the malaria pigment's components, we were able to dissect their impact on the immune reactivity of a macrophage model cell line. Reactions between BH and red blood cell (RBC) ghosts effectively reconstituted the observed immunomodulatory reactivity of native hemozoin. Together, these results suggest that the interaction between hemozoin and the RBC lipids results in the generation of toxic products and that these products are responsible for disrupting macrophage function in vivo.

Topics: Aldehydes; Animals; Cell Line; Cells, Cultured; Erythrocyte Membrane; Hemeproteins; Hydroxyeicosatetraenoic Acids; Immunosuppressive Agents; Macrophages; Malaria; Mice; Microscopy, Confocal; Molecular Structure; NADPH Oxidases; Nitric Oxide Synthase Type II; Oxidation-Reduction; Reactive Nitrogen Species; Reactive Oxygen Species

beta-Hematin is an important heme metabolite of malarial infection. Its role as an agent mediating the formation of the reactive electrophile 4-hydroxynonenal (HNE) from polyunsaturated fatty acids was investigated. In vitro formation of HNE was found to be facilitated by the presence of hemozoin in a concentration-dependent fashion. The reactivity of HNE derived from reaction with beta-hematin was confirmed through its ability to form protein adducts on myoglobin.

Topics: Aldehydes; Animals; Fatty Acids, Unsaturated; Heme; Hemeproteins; Malaria; Myoglobin; Plasmodium falciparum

Malarial anemia involves destruction of parasitized and non-parasitized red blood cells and dyserythropoiesis. Malarial pigment, hemozoin (HZ), is possibly implicated in dyserythropoiesis. We show that supernatants of HZ and HZ-fed-monocytes, and 4-hydroxynonenal generated by them, inhibited progenitor growth.

Topics: Aldehydes; Anemia; Animals; Cells, Cultured; Erythropoiesis; Hemeproteins; Humans; Malaria; Plasmodium