Plasmodium berghei - infected red cells sorted according to DNA content.

A cell-sorting method is described for the analysis and separation of red blood cells in Plasmodium berghei-infected mouse blood based on their DNA content. This method involves a selective uptake of the bis-benzimidazole dye 33258 Hoechst, a DNA-binding dye, by red blood cells containing parasites. Infected blood is incubated at 37 degrees C with the dye then washed at 4 degrees C to remove unbound dye. Uninfected cells are then non-fluorescent at the characteristic wavelengths for 33258 Hoechst excitation and emission, whereas parasitized cells display fluorescence intensities in approximately direct proportion tothe number of parasite nuclei (i.e. amount of parasite DNA) within the cell and can be sorted accordingly. Providing cells were incubated in a complex nutrient medium during dye uptake at 37 degrees C, the sorted parasite-infected cells produced lethal P. berghei infections when injected into BALB/c mice. The dye-labelling technique is simple and sufficient red blood cells at various stages of infection can be collected for biochemical or immunochemical studies by cell sorting.     

Human neutrophil--mediated destruction of antibody sensitized herpes simplex virus type I infected cells

Human peripheral blood polymorphonuclear neutrophils (PMN) were tested for their ability to act as effector cells in antibody-dependent cell cytotoxicity (ADCC) against Herpes simplex virus (HSV) infected target cells sensitized with anti-HSV serum. The PMN from all 29 individuals tested could mediate ADCC in the presence of a standard human anit-HSV serum. Since PMN are prominent cells early in herpes lesions, it was hypothesized that because ADCC could represent an in vitro model for antiviral recovery, perhaps the efficacy of PMN at mediating ADCC might be impaired in those subjects to frequent recrudescent herpes. However, evidence for the hypothesis was not obtained since the PMN from individuals with frequent, infrequent, or unrecorded herpes labialis all showed approximately the same activity at mediating ADCC. Alternative ways in which PMN could be involved in antiviral recovery were discussed.     

Mechanisms of recovery from viral infections: destruction of infected cells by neutrophils and complement

Highly enriched populations of bovine neutrophils (PMN) were able to destroy herpesvirus-infected cells when in the presence of C. This mechanism of cytotoxicity was termed C-dependent neutrophil-mediated cytotoxicity (CDNC). To demonstrate CDNC required viable PMN, an active source of C, and a target cell expressing viral antigens. Noninfected cells were not susceptible to lysis. Several approaches were used to exclude the presence of antibody as an explanation for the cytotoxicity observed. In a comparison of the effectiveness of different cell types at mediating CDNC, PMN were more effective than macrophages, and lymphocytes were without activity. The results was discussed in terms of the possible in vivo significance of the neutrophil as a cell type capable of mediating recovery from infection, since if a mechanism similar to CDNC occurs in vivo, this could play a role in defense before the time when protective levels of antibody and immune cells are generated.     

Erythrocyte interaction during aggregation according to scanning electron microscope findings

Scanning electron microscopy was used to study red cell aggregation caused by intravascular injection of dextran (m. w. 500 000). Two types of aggregates were revealed. Type 1 was represented by aggregates of red cells with no substantial change in the form while type 2 by aggregates with the wrinkling of the surface of red cells and enlarged area of red cell surface contacts. Both types are prone to disaggregation after injection of rheopolyglucin.     

X-ray anatomy of the superior mesenteric artery according to intravital angiographic findings

Analysis of clinical angiograms made in 1,624 subjects has determined roentgenological differential signs in the upper mesenteric artery. They help to recognise the artery in roentgenograms of frontal and lateral projections regardless the method of contrasting. For the first time roentgenological data on this vessel are obtained concerning age, sex and habitus of the subjects examined. The data were calculated according to biometrical requirements. In women, the artery begins higher than in men. In the asthenics, the artery begins lower than in the normosthenics and still lower than in the hypersthenics. The diameter of the artery in man is wider than in women. At the age of 60 years, the arterial lumen is 10--15% wider than that in persons below 20 years of age.     

Muscle cells enhance resistance to pro-inflammatory cytokine-induced cartilage destruction

Pro-inflammatory cytokines IL-1β and TNFα play important roles in the manifestation of arthritis by disrupting the anabolic and catabolic activities of the chondrocytes. We observed a novel mechanism of cartilage regulation by which muscle cells diminish the response of chondrocytes to IL-1β and TNFα. We found that chondrocytes cocultured with muscle cells or cultured in muscle cell-conditioned medium significantly enhanced the expression of cartilage matrix proteins (collagen II and collagen IX) and resisted IL-1β and TNFα-induced cartilage damage. Our data suggest that this effect is achieved by inhibiting the expression of key components of the signaling pathways of pro-inflammatory cytokines (including NFκB, ESE-1, Cox-2, and GADD45β), leading to attenuated expression of cartilage-degrading enzymes (MMPs and ADAMTS4). Therefore, our work unveils a potential role of muscle in regulating cartilage homeostasis and response to pro-inflammatory stimuli, and provides insights on designing treatment strategies for joint degenerative diseases such as arthritis.     

Increased or decreased immunogenicity of tumor-associated antigen according to the amount of virus-associated antigen in rat tumor cells infected with friend virus

Summary The immunogenicity of tumor-associated antigen (TAA) in 3-methylcholanthrene-induced rat fibrosarcoma KMT-17 cells was investigated with particular reference to the amount of virus-associated antigen (VAA) produced on the cell surface after artificial infection of tumor cells with Friend murine leukemia virus (FV). To compare the TAA immunogenicity of FV-infected KMT-17 tumor cells (FV-KMT-17) with non-infected KMT-17 cells, rats were immunized with the above two tumor-line cells, and the growth of original non-infected KMT-17 cells was observed. The result showed that TAA immunogenicity was not always paralleled by the amount of VAA newly produced. The amount of VAA increased in proportion to the number of passage generations of FV-KMT-17 cells through FV-tolerant rats that had been neonatally injected with Friend virus. The TAA immunogenicity of FV-KMT-17 cells after two passages was lower than that of non-infected KMT-17 cells. High TAA immunogenicity was observed in FV-KMT-17 tumor cells of the fourth passage generation; it then weakened gradually after subsequent passages of FV-KMT-17 tumor cells, and finally dropped to a level lower than the immunogenicity of the original non-infected tumor cells. However, such variations of immunogenicity were never observed in FV-tolerant rats. These observations suggest that TAA immunogenicity definitely increases when an appropriate amount of VAA is expressed on the tumor cell surface and decreases when a relatively low or excessive amount of VAA is expressed. We discuss the mechanisms leading to the above findings. Abbreviations used in this paper are: FV, Friend murine leukemia virus; TAA, KMT-17 tumor-associated antigen; VAA, FV-associated antigen     

Reassortment of cells according to position in mouse morulae

Sixteen-cell mouse morulae were disaggregated and blastomeres originally occupying outer or inner positions were separated. Outer, inner, or unsorted populations of blastomeres were labeled with either trinitrobenzene sulphonic acid (TNP) or fluorescein isothiocyanate (FITC) and individual blastomeres aggregated to unlabelled partially decompact eight- to ten-cell morulae. After up to 6 h in culture, the positions of the labelled blastomeres within the aggregates were examined. The combined results demonstrated that between 86 and 92% of outer cells remained on the surface of the aggregate and flattened into extensive polygonal shapes, whereas 76-77% of the inner cells had become engulfed by the host morula cells and retained their initial spherical shape. Using unsorted cells, 33-37% were internalised, which is compatible with the most recent estimates of the presence of six to eight inner cells at the 16-cell stage. The possibility that differential adhesiveness of the outer and inner cells is involved in the allocation of cells to the trophectoderm and inner cell mass of the blastocyst is discussed.     

Separation of lymphoid-line cells according to volume and density

Velocity sedimentation and isopycnic centrifugation was used to separate exponentially growing RAJI lymphoid line cells according to size and density. Mixtures of Ficoll, Isopaque and tissue culture medium were used as gradient media. These media had a constant pH, were isotonic, and did not have any significant harmful effect on the cells. The observed variation in cell size paralleled the progression of the cells through the cell cycle, as assessed by thymidine incorporation and impulse cytophotometric determination of DNA contents. Differences in cell density did not reflect the cell cycle phase. No correlation could be established between cell size and density. Velocity sedimentation could be used to obtain cell populations which were relatively pure according to cell cycle phase and growing synchronously for at least 24 h.