Biomedical subjects
Publications and source records attributed to L Manuelidis.
Ferritin-Ricin II and Ferritin-Concanavalin A bound to budding as well as mature C-type viral particles. No differencies in binding between the viral coat and adjacent plasma membrane were detected with either lectin conjugate. Aggregation of viral particles by lectin conjugates was observed, and linking of virus to the plasma membrane resulted in phagocytosis of viral particles.
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The ganglioside level and pattern of human gliomas in monolayer cultures were examined. These gliomas revealed morphological variations that correlated with several features of ganglioside analysis. Glioblastoma lines TC 178 and TC 501 that morphologically had changed during extended subculture revealed reduced amounts and a simplified pattern of gangliosides with almost total loss of the characteristic brain complex gangliosides. In contrast, two glioblastoma lines TC 526 and TC 593, as well as the oligodendroglioma line TC 620 showed brain-like gangliosides and the cells in these cultures had maintained their characteristic morphology observed during early subcultures. The possibility that altered ganglioside levels occur in conjunction with morphological changes after propagation in vitro is discussed.
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Human glioblastoma cells in long-term monolayer culture showed an even distribution of intramembrane particles (IMP) on all surfaces of the plasma membrane; junctional complexes were rarely observed and rectilinear arrays were not seen. Cells treated with Con A-ferritin and Ricin II-ferritin showed an even distribution of lectin receptors and under conditions used no capping occurred. Lectin-ferritin complexes were taken up into pinocytotic vesicles. Cleaved preparations of Ricin II-ferritin treated cells showed no change in the distribution of IMP.
Cholera toxin coupled to peroxidase yielded a highly specific ultrastructural marker of plasma membrane monosialogangliosides. Studies with cultures of brain and brain tumors suggested that long-term culture of tissue in monolayers results in eventual loss of surface monosialogangliosides.
The chromosomal localization of satellite DNA in two tissue culture lines derived -rom malignant mouse CNS tumors was investigated by in situ hybridization of 3H single-stranded satellite DNA purified by isopynic centrifugation in alkaline CSC1. Both tumors were glioblastomas originally induced by a methylcholanthrene implantation into the cerebrum of C3H mice; both displayed aneuploid chromosomal constitutions. One of these glioblastomas (TC 541) revealed labelling only of centromeric portions of the chromosomes even in cells containing greater than 200 chromosomes and thus it had a pattern of satellite distribution comparable to that of normal cells. The other glioblastoma (TC 509), that produced C-type particles and had a decrease in satellite DNA, displayed interstitial and telomeric label in some chromosomes in addition to labelling of the centromeres. "Hoechst 33258" fluorescence showed some interstitial and telomeric bright bands as well as centromeric bright regions, though to be consistent with in situ studies. The localization of satellite DNA to the chromosome arms and its possible relation to C-type virus is discussed.
Cholera toxin was coupled to peroxidase to yield a highly specific marker for GM1 gangliosides. Study of embryonic brain cells in culture revealed intense binding of cholera-peroxidase by plasma membranes of both neurons and glial cells. In contrast, long-term monolayer glioblastoma cultures, including one producing C-type virus, revealed virtually no labelling of their plasma membranes. Such cells were shown to be capable of incorporating exogenously applied GM1 into their plasma membranes. Studies with fixed brain and synaptosomal fractions were in accord with results on embryonic brain cells in culture, and autoradiographic findings with 125I cholera supported observations made utilizing cholera-peroxidase. From our studies there is some indication that long-term propagation in vitro alters the plasma membrane GM1.
Chromaffin cells in organotypic cultures of adrenal glands from 19 day rat embryos developed a marked increase in the volume of their rough endoplasmic reticulum (RER) and enlargement of nucleoli in response to 1 mM dibutyryl cyclic AMP (dbcAMP) treatment for 17 days in vitro. 0.1 mM dbcAMP had a similar but less dramatic effect. When dbcAMP was removed from the medium after 11 days in vitro, cells became more like control explants treated with no additives, suggesting the effects of dbcAMP were reversible. ACTH which selectively stimulates adrenal cortical cells had no effect on RER morphology of chromaffin cells. The 'neuronal-like' configuration of dbcAMP treated chromaffin cells is discussed.
The transmission and serial propagation of Creutzfeldt-Jakob disease from man to guinea pigs are reported. The latency, symptomatology, and morphology of the infection during the first four passages are presented. The incubation period between the first and subsequent passages was halved. One hundred percent take, morbidity, and mortality were achieved in all inoculated animals. All guinea pigs developed a subacute spongiform virus encephalopathy with marked neuronal destruction in the cerebral cortex and subcortical grey structures. The neuronal loss resulted in cerebral atrophy and hydrocephalus ex vacuo.
The quantitative preservation of satellite NA was studied in several central nervous system (CNS) neoplasms; four tumor lines deriveo from 3-methylcholanthrene implantation into the CNS of mice were compared with brain and tissue cultures of normal mouse cells by analytical centrifugation in cesium chlorie. Three tumors showed no detectable difference from normal cells; nuclear and whole cell preparations were comparable. Only a glioblastoma line proucing C-type particles (TC509) revealed a significant difference from normal cells and exhibited a decrease of approximately 20% in satellite DNA or 2% of the total DNA on repeated examination for 1 year. C-type RNA virus may be related to relative decreases in satellite DNA observed in TC509.
Human DNA digested with Hae III showed multiple repeats of a 170 base pair fragment. The most prominent band was the 340 base pair dimer, estimated to be 0.8% of the entire genome. Eco R1 and Hha I yielded fragments with similar electrophoretic mobility to the Hae III dimer. In each case this band was markedly enriched in DNA reassociating at a 0t of less than or equal to 1. Hybridization of the Hae III dimer to gels eluted on to filters demonstrated that the multiple Hae III fragments and Eco R1 fragments contained compatible sequences. These sequences may comprise a distinct subclass of DNA.
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Cells from a human glioblastoma (TC 526) maintained in tissue culture for ten years had a mean membrane potential of 27 +/- 0.9 mV at an external potassium concentration [Ko] of 5.3 mM. When [Ko] was varied between 2.5 and 5.3 mM, membrane potential changes were close to those predicted by the Nernst equation. At higher [Ko], the Nernstian slope was approached only in the presence of 10(-5) M ouabain, which did not affect membrane potential at a [Ko] of 5.3 mM. An electrogenic sodium pump activated by high [Ko] could explain these findings; such a mechanism has been demonstrated in other tissues.
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