Lakshmi M. Kumaratilake

GM‐CSF‐induced priming of human neutrophils for enhanced phagocytosis and killing of asexual blood stages of Plasmodium falciparum: synergistic effects of GM‐CSF and TNF

Granulocyte macrophage‐colony stimulation factor (GM‐CSF), which is a haematopoietic cytokine generated by activated T lymphocytes and macrophages during infection, was investigated for its effects on human neutrophil‐mediated killing of asexual blood forms of Plasmodium falciparum. Pretreatment of neutrophils with human recombinant‐GM‐CSF markedly increased the parasite killing (measured by a radiometric assay), in the presence of normal serum (containing complement), immune serum (IS), purified IgG (from IS) or heat inactivated IS. GM‐CSF pretreatment also enhanced phagocytosis of the parasite by neutrophils and the expression of CR3, FcγRII and FcγRIII receptors. Treatment of neutrophils with a combination of GM‐CSF and TNF resulted in a synergistic increase in phagocytosis and killing of the parasite. The findings suggest that GM‐CSF is likely to form part of the cytokine network responsible for regulating the antiparasitic activity of the neutrophil in malaria.

Opsonization and Phagocytosis of Plasmodium falciparum Merozoites Measured by Flow Cytometry

ABSTRACT A flow cytometric phagocytosis assay was established to investigate the role of anti-merozoite antibody, complement, and cytokines on the phagocytosis of Plasmodium falciparum merozoites by human neutrophils. This assay involved allowing fluorescein isothiocyanate-labeled merozoites to interact with phagocytes and analysis of the cells on a FACScan with Lysis II software. To differentiate the proportion of neutrophil surface-bound merozoites from the merozoites ingested by neutrophils, the fluorescence of bound merozoites was quenched by adding trypan blue. The data showed that sera from malaria-immune individuals in the Solomon Islands and Papua New Guinea promoted merozoite engulfment by neutrophils. The cytokines tumor necrosis factor alpha, gamma interferon, granulocyte-macrophage colony-stimulating factor, and interleukin-1β significantly increased the amount and the rate of merozoite phagocytosis by neutrophils. Optimum merozoite phagocytosis occurred when both cytokines and anti-malarial antibody were present.

Augmentation of the human monocyte/macrophage chemiluminescence response during short‐term exposure to interferon‐gamma and tumour necrosis factor‐alpha

The effects of short‐term (30 min) pre‐incubation of human monocytes and macrophages (3‐day cultured monocytes) with leucocyte‐derived human interferon‐gamma (IFN‐γ) and recombinant human tumour necrosis factor‐alpha (rTNF‐α) were examined. Pre‐incubation of either monocytes or macrophages with rTNF‐α or IFN‐γ (100 U/5 × 105 cells) augmented their respiratory burst to formyl‐L‐methionyl‐L‐leucyl‐L‐phenylalanine (fMLP), measured by the luminol‐ and lucigenin‐dependent chemiluminescence assay. In addition, both cell types showed a burst of respiratory activity in the presence of rTNF‐α or IFN‐γ only. The effects of IFN‐γ were removed by adsorption with an anti‐ IFN‐γ monoclonal antibody and those of rTNF‐α were abolished by heating at 100°C, or by the addition of anti‐TNF‐α monoclonal antibody. The results demonstrate that both IFN‐γ and rTNF‐α are stimulators of monocytes and macrophages, and rapidly alter the capacity of the cells to respond to fMLP, which binds to cell surface receptors.

Extraction of intraerythrocytic malarial parasites by phagocytic cells.

Phagocytosis is an intricate process adopted by some unicellular organisms as a feeding behaviour. It has developed in the tissues of multicellular organisms, both vertebrates and invertebrates, as a defence system to confine and eliminate foreign matter and, in this manner, protect the host against infection. During evolutionary development, phagocytic cells have evolved to show greater specificity. Lakshmi Kumaratilake, Antonio Ferrante, Jaliya Kumaratilake and Anthony Allison here describe a unique mechanism used by phogocytic leukocytes to engulf intra-erythrocytic malarial parasites.

SYNTHESIS OF HYDROPEROXIDE AND PERKETAL DERIVATIVES OF POLYUNSATURATED FATTY ACIDS AS POTENTIAL ANTIMALARIAL AGENTS

Hydroperoxide derivatives of beta-oxa-substituted polyunsaturated fatty acids were prepared by 15-lipoxygenase catalysed oxidation and perketal derivatives of fatty acid hydroperoxides were synthesized. The perketals are more stable than their parent fatty acid hydroperoxides, but less active as antimalarial agents in the in vitro growth inhibition of Plasmodium falciparum

Degradation of human cartilage by monocytes

The effect of human peripheral blood monocytes on the degradation of human articular cartilage was studied in vitro using a radiometric assay to detect proteoglycan breakdown. The results showed that proteoglycan breakdown was increased by 60% after a 20 h exposure to monocytes (p less than 0.001).