Despite the evidence that functional malarial immunity is multifactorial, involving complex interactions of humoral and cellular immune responses, present knowledge of efferent mechanisms of cell-mediated immunity is limited. However, immunologic resolution of murine and primate malarias are often associated with intraerythrocytic retardation of parasite development resulting in moribund parasites known as crisis-forms. Human malaria immune serum from Sudan retards Plasmodium falciparum in vitro by a non-antibody substance termed crisis-form factor, CFF. Epidemologic evidence reveals that CFF is closely associated with clinical malaria immunity in Sudan, where serum CFF concentrations increase with malaria endemicity, and fluctuate with seasonal malaria transmission. Thus, CFF may be part of the acquired immunity to malaria, and a product of cell-mediated responses. It is the specific aim of this project to purify and characterize CFF; to determine its source and the stimulatory mechanism, or antigen associated with its production, and to determine how it retards parasite development. Purification of CFF from human immune serum will be conducted using standard column chromatographic procedures followed by high-performance liquid chromatography using FPLC. Polyclonal and monoclonal antibodies will be developed against purified CFF to be used for ultimate purification by affinity chromatography and for analysis of the cellular source and stimulatory cascade leading to CFF production. The relationship between CFF and known cytokines will be studied by comparing effects of huIFNGamma, Il 1,2,3, TNF and other known lymphokines and monokines on cultures of P. falciparum. Antiplasmodium activity of secretory products of human mononuclear cells stimulated in vitro with known activators will be compared to activity of CFF-containing human immune sera to determine the cell type responsible for CFF production, and as a prelude to investigation of the in vivo production of CFF and the possible role of malarial antigens in its production. The mode of action of CFF will be studied by monitoring incorporation of radiolabelled precursors of parasite protein and nucleic acid synthesis, glucose utilization, mitochondrial activity and other known parameters of parasite metabolism in highly synchronized cultures of P. falciparum.
