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• W2332814109 abstract "A study of trypsin-dispersed chick liver cell cultures infected with the virulent Jones' Barn strain of Trichomonas gallinae was made by several cytochemical methods. A decline in RNA and RNP is apparent from the 8th hr following inoculation of parasites and becomes pronounced with time. The DNA seems unchanged during the entire experiment. No changes in RNA or DNA were observed in the controls. Significant decrease in glycogen in the epithelial cells becomes evident at 8 hr postinoculation, and after 16 hr this polysaccharide can no longer be detected cytochemically. The controls show no change in glycogen levels. The foregoing changes in the infected cultures are paralleled by an accumulation of lipids in all the cell types. The macrophages and to a lesser extent the epithelial cells contain certain PAS-positive sudanophilic granules, very likely glycolipoprotein in nature, which are probably associated with phagocytosis. The high pathogenicity of the Jones' Barn (JB) strain of Trichomonas gallinae for birds, especially pigeons (Stabler, 1954), mice (Honigberg, 1961; Frost and Honigberg, 1962), and cell cultures (Honigberg et al., 1964) has been well established. In trypsin-dispersed chick liver cell cultures, the parasites cause degeneration of the epithelial and fibroblast-like cells and stimulate activity of the macrophages. The flagellates are phagocytized by the macrophages, within which they multiply. Further, they appear to invade some of the epithelial and fibroblast-like cells (Honigberg et al., 1964). In view of these findings it was thought worthwhile to evaluate the effects of the JB strain on the cell cultures with the aid of cytochemical methods. The present study, which constitutes the first phase of the proposed investigation, deals with observations made on the nucleic acids, glycogen, other PAS-positive material, and lipids in normal and infected trypsin-dispersed chick liver cell cultures. A preliminary report of the study was given at the AIBS meetings at Boulder, Colorado in August 1964 (Abraham and Honigberg, 1964). MATERIALS AND METHODS The Jones' Barn strain, isolated originally by Dr. R. M. Stabler, was employed in this study. The Received for publication 15 February 1965. * This investigation was supported by Research Grant AI-00742 from NIAID, U. S. Public Health Service, and a Teachers Research Grant from the University of Massachusetts. tPermanent address: Osmania University, Hyderabad, India. details of the history of this strain are found in various reports of Stabler (see Stabler, 1954). In all experiments the December 1962 isolate was used (see Honigberg, 1961). The trypsin-dispersed liver cell cultures were prepared and the experiments set up according to the method described by Honigberg et al. (1964). Earle's BSS was used in preparation of the media and for washing cultures. In all instances 5 X 105 organisms from 24-hr trichomonad cultures were inoculated in 0.33 ml of CPLM medium without agar into the experimental cultures, while similar aliquots of fresh CPLM were added to the control cultures. The total volume of the medium in all cultures was 1 ml and the medium was adjusted to about pH 7.5 with the aid of 5% CO2 in air. The experimental and control preparations (on 9 to 10 by 22 mm cover glasses), at least two of each, were removed and fixed at intervals of 1, 2, 4, 6, 8, 10, 12, 16, and 20 hr. At each time interval the pH of the medium was also checked. The cell cultures were washed in warm Earle's BSS for at least 5 min before processing them for cytochemical studies. This proved helpful in removing debris accumulated from the medium which otherwise might have given false reactions with the dyes. For the study of the deoxyribonucleic and ribonucleic acids (DNA and RNA), some of the cells were fixed in a cold mixture of diethyl ether and absolute alcohol (1 : 1), stained according to the acridine-orange technique of Bertalanffy and Bertalanffy (1961), and examined under a fluorescence microscope (exciter filters Corning no. 5970 or 5840). May-Griinwald-Giemsa stain was employed for demonstration of deoxyriboand ribonucleoproteins (DNP and RNP), while Azure A-Schiff (Himes and Moriber, 1956) was used for DNA after fixation in Carnoy's fluid. Ribonuclease-treated preparations were employed as controls in the study of RNA. Glycogen and certain carbohydrate-containing materials were demonstrated by the periodic acidSchiff technique (PAS) of McManus (see Pearse, 1961) and counterstained with celestine blue and Meyer's hemalum, after fixation in cold acetic" @default.
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• W2332814109 title "Cytochemistry of Chick Liver Cell Cultures Infected with Trichomonas gallinae" @default.
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