No abstract

The life cycle of Leishmania involves two distinct stages of the parasite: the flagellated promastigotes live extracellularly in the gut of insect Phlebotomes, and upon transmission to the vertebrate host they are taken up by macrophages and transform into nonflagellated amastigotes. The amastigotes are obligatory parasites of macrophages and lodge within modified phagolysosomes termed parasitophorous vacuoles (1, 2). Amastigotes obtained from tissues of infected animals readily infect primary macrophage cultures, macrophage lines, or Sticker sarcoma cells (3-6).In a variety of in vitro models oxygen metabolites produced by macrophages stimulated by immune complexes, particles, or membrane-active drugs have been related to the killing of intracellular microorganisms by the phagocytes (7-9). The key metabolite is the superoxide anion (O2-), derived from the univalent reduction of molecular oxygen. 02-can undergo dismutation to hydrogen peroxide (H202) and can lead to the generation of other toxic oxygen rnetabolites such as the hydroxyl radical (OH.) (I0, I I). It has also been shown that a series of electron carriers, including phenazines, thiazines, and quinones, when in contact with living cells such as Escherichia coli, can be reduced to auto-oxidizable intermediates. Upon reoxydation, these intermediates can generate 02-and H202 (12). For this reason we chose to investigate the effect of 5-methylphenazinium methyl sulfate (phenazine methosulfate, PMS) ~ and other potential redox cycling agents on Leishmania m. amazonensis. We report here that intracellular amastigotes are killed by drug concentrations that do not appear to be toxic to the host phagocytes and that low concentrations of the drugs also inhibit the growth or kill Leishmania promastigotes in culture.

A previous study of the characteristics of the lesion produced by poliomyelitis virus in vitro in fixed and stained preparations of human adult fibroblasts indicated that an msinophilic area, which becomes soon more precisely outlined, appeared in the perinuclear cytoplasm between 15 and 20 hours after infection( 1). As this eosinophilic mass increased in size in the center of the cell, the nucleus and the cytoplasm, which became increasingly basophilic in content were pushed to the periphery. A t the next stage, the cell progressively emptied itself of its cytoplasm. We have since observed a similar lesion in fixed and stained preparations of fibroblasts derived from human embryonic muscle and of human and monkey kidney epithelial cells, infected with poliomyelitis virus. The increasing basophilia of the virus-infwted cell was found to be associated with a strongly positive and generalized Gomori alkaline phosphatase reaction. The purpose of the present communication is to describe the evolution of the cellular lesion as it was observed by phase contrast cinematography wikhout fixation or staining.Materials and methods. Strains of fibroblasts derived from human ,tonsils were maintained by serial passage in roller tubes for 2 to 3 months before they were used. These cells are 3 to 4 times as large as the average skin-muscle fibroblasts, regular in shape, and remarkably resistant to nonspecific degeneration. Their susceptibility to poliomyelitis virus was reported in a previous communication (2 ) . For examination with the phase contrast microscope explants were prepared by the hanging drop technic. The medium consisted of chicken plasma, human serum free of antibodies for the viruses that were used, bovine amniotic fluid, Hanks' solution and traces of chick embryo extract. The MEFl strain of Type 2 virus and the Mahoney strain of Type 1 poliomyelitis virus were used and it may be noted that the lesions were the same with both. Before transplantation to the hanging drop the cells were mixed with virus suspensions having a titer of 1k5 and left at room temperature for 30 minutes. Observations were carried out at 36-37OC and at 30-31°C. The evolution of the lesion was slower at 30-31°C but otherwise the same at both temperatures. The lower temperature was used more often because the control cells remained in better condition at 30-31°C. The observations were recorded with a Zeiss; Winkel phase contrast microscope at magnifications of 240X, 540X and 9OOX, using a 1OOV tungsten, ribbon lamp for the light source. The moving picture system of the camera was the one designed by J.

Developing eosinophils from the bone marrow of a patient with acute "eosinophilic" leukemia were characterized by electron microscopy. It was suggested that the first sequential step in granule formation occurred at the level of the endoplasmic reticulum without actual participation of the Golgi complex. Progressive densification of the former profiles, presumably mediated by Golgi vesicles, resulted in the formation of dense immature granules. Ultrastructural observations of the "leukemic" eosinophils which were generally arrested at an intermediate stage of maturation revealed also large vacuoles containing sequestered immature granules, without any indication of phagocytic activity. Morphological evidence that has been accumulated indicates that the membrane of these vacuoles fused with the cell membrane, thus being in contact with the extracellular space. These profiles strongly suggested that granules and/or granule-associated material were secreted by developing bone marrow eosinophils.