Comparative Study of Cultured Burkitt Tumor Cells by Immunofluorescence, Autoradiography, and Electron Microscopy

Cultured Burkitt cells were examined by immunofluorescence, autoradiography, and electron microscopy in an effort to identify the stainable cells with those harboring herpes-type virus particles. Immediately after a 2-hr pulse of (3)H-thymidine, from 30 to 60% of the cells revealed heavy nuclear labeling. In most cases the grains were evenly dispersed, but in about 3 to 5% the grains showed a focal distribution and occasionally they extended into the cytoplasm. Such nuclear foci were rarely seen at 8 hr after the pulse. When the analysis was restricted to preselected immunofluorescent cells, up to 80% showed label at 8 hr and cytoplasmic grains were prominent. To reduce cellular deoxyribonucleic acid (DNA) synthesis, cells were X-irradiated with 3,000 to 6,000 R, and the isotope pulse was applied 1, 4, or 7 days later. Whereas the total number of labeled cells decreased in roughly twofold steps at the respective intervals (from 40 to 10%), the incorporation of (3)H-thymidine into fluorescent cells was not affected by X irradiation. In each series, about 70% of the fluorescent cells contained label when they were examined at 24 and 48 hr after the pulse, whereas at 8 and 72 hr fewer were positive. At the earlier intervals, unlabeled fluorescent cells most likely represented cells which had completed viral DNA synthesis prior to the pulse; at the later intervals, unlabeled fluorescent cells were probably cells which commenced viral replication after the pulse. These data support the conclusion that the immunofluorescent cells are the ones which harbor virus, and also confirm the expectation that the virus is a DNA virus from a member of the herpes group. This conclusion was firmly established by sectioning and electron microscopic examination of individual fluorescent cells, all of which contained numerous virus particles, whereas the nonstained cells prepared in a similar manner were free of them.     

Inhibition of Swarming of Proteus by Sodium Tetradecyl Sulfate, β-Phenethyl Alcohol,and p-Nitrophenylglycerine

The effects of sodium tetradecyl sulfate (STS), β-phenethyl alcohol (PEA), and p-nitrophenylglycerol (PNPG) on motility, swarming, flagellation, and growth of Proteus were examined. Growth-inhibitory concentrations (GIC) and swarming-inhibitory concentrations (SIC) were determined. A characterization of the swarming-inhibitory efficacy of these compounds was based on their GIC/SIC ratio and their concentration inhibition curves. Using the homologous series of sodium alkyl sulfates as a standard reference, we showed that PNPG was more effective than STS, which was the most effective of the homologous series. PEA was less effective than sodium decyl sulfate but more effective than sodium octyl sulfate. Motility tests in liquid medium and electron microscope investigations indicated that the modes of action of the three compounds, all of which effectively inhibit the swarming of Proteus, are different. Whereas STS and PEA inhibit swarming by inhibition of motility, PNPG seems to act on the swarming mechanism sensu strictori, without impairment of motility. STS immobilizes by inhibition of flagellum formation or by some lytic action on the flagella already synthesized. PEA acts by impairing flagellar function, but leaves the flagella morphologically intact.     

Electron microscopic observations on isolated mitochondrial membranes,prepared by various histological methods

Summary The pictures of isolated mitochondrial membranes, as seen on the electron-microscope, depend very much on the method of specimen preparation. Subunits of linear dimensions of about 25 m, (electron transport particles) are observed in carbon-replicas of the membranes and in specimens treated with trypsin or pepsin (0.02% for 30 mins) and shadowed with platinum. A three-layered structure of the unit membrane is seen in sections of specimens fixed with osmium tetroxide or formalin followed by post-fixation with osmium tetroxide. But fixation with potassium permanganate or with formalin, followed by post-fixation with potassium permanganate reveals an electron-dense globular structural element in the unit membrane. An electron-transparent ultrastructural element of the unit membrane is observed after treatment with trypsin (0.2% for 5 mins) and fixation with osmium tetroxide. Unsectioned specimens treated with 0.02% trypsin for 30 mins show a honeycomb-like structure of the membrane. Thus, part of the results appear to support the concept of a mosaic-like structure of the unit membrane, whereas other results are in agreement with the classical concept of a three-layered structure.The authors wish to express their gratitude to Dr. Sina Rosenthal, Department of Physiological Chemistry, Humboldt University, Berlin, who prepared the isolated membranes, to Mr. E. Fischer, Head Technician of the Department of Electron Microscopy, Greifswald University, who took most of the electron micrographs, to Mr. G. Bartsch, Department of Electron Microscopy, Greifswald University, and especially to Prof. W. Bargmann and to Doz. E. Lindner, Department of Anatomy, Kiel University, for many valuable suggestions.     

Die Feinstruktur von Milz und Leber bei experimenteller Amyloidose

Zusammenfassung Experimentell erzeugtes Amyloid besteht aus einer faserigen Komponente und einer homogenen Kittsubstanz. Darin eingelagert sieht man regelmäßig Thrombocyten. Die faserige Komponente des Amyloids besitzt eine andere Struktur als Reticulum- und Kollagenfasern und verhält sich nach Kontrastierung mit Schwermetallsalzen anders als diese. Mit großer Wahrscheinlichkeit handelt es sich bei den Fasern um die Eiweißkomponente der amyloiden Substanz. Das Amyloid stellt ein rein zwischenzelliges Differenzierungsprodukt dar, es tritt niemals intrazellulär auf.Während der amyloiderzeugenden Behandlung kommt es zu einer plasmazellulären Transformation von Zellen in der roten Pulpa der Milz. Vergleichbare Erscheinungen zeigen sich auch in der Leber. Die Amyloiddepots liegen häufig in unmittelbarer Nachbarschaft von Plasmazellen. Das Cytoplasma, insbesondere Ergastoplasmalamellen von Plasmazellen stehen oft mit dem Amyloid ohne Zwischenschaltung von Zellmembranen in unmittelbarem Kontakt. Diese Befunde sprechen für eine Beteiligung von Plasmazellen bei der Amyloidentstehung. Insbesondere können sie das regelmäßige Auftreten von Antigen-Antikörper-komplexen im Amyloid verständlich machen.Die Untersuchung wurde mit Hilfe der Deutschen Forschungsgemeinschaft durchgeführt.Herrn Prof. Dr. E. Letterer zum 30. 6. 60 gewidmet.     

Gibberellin and sympodial branching inTilia

U r?stově na za?átku fyziologického odpo?inku inhibovaných rostlinTilia platyphyllos Scop. aTilia cordata Mill. se vlivem giberelové kyseliny (GA₃) zachoval terminální pupen jako náznak monopodiálního větvení, co? lze vylo?it jako fylogenetickou rekapitulaci. P?itom se ukázalo, ?e po?et normálně na letorostech zachovávaných list? je p?edur?en ji? v pupenech.     

Stephanoscyphus (Scyphozoa,Coronatae) und seine direkte Abstammung von den fossilen Conulata

The scyphopolypStephanoscyphus Allman 1874 represents the polyp generation of the scyphomedusan order Coronatae. Though this polyp has been known for more than a hundred years its general morphology, systematics, and evolution have been inadequately described. Participating in the International Indian Ocean Expedition, 1964 to 1965, on board of the German research vessel “Meteor”, the author was able to collect a sufficient supply of livingStephanoscyphus off the coasts of South Arabia and East Africa. For the first time, it was possible to rear these polyps in laboratory cultures. A thorough investigation of morphology, developmental history and behaviour based on longterm observations of the living polyps gave clear indications thatStephanoscyphus directly descended from the fossil group of Conulata, the scyphozoan nature of which has been affirmed byKiderlen (1937) andKnight (1937). The main feature whichStephanoscyphus has in common with the Conulata is the possession of a periderm tube. The characteristics found in a detailed investigation of the periderm tube conform well with those found in the periderm of the Conulata except for the closure of the aperture by triangular flaps which are absent inStephanoscyphus. The soft body contains primitive features as well. Hence it must be concluded finally that the type of organization which the fossil ancestors exhibited has survived inStephanoscyphus and that the Coronatae represent the most basic group of all living Scyphozoa. On the other hand, the results give strong support for the scyphozoan nature of the Conulata, the organization and life history of which have been elucidated by the observations of the living representatives ofStephanoscyphus.     

Luciferin derivatives in bioluminescence-enhanced enzyme immunoassays

Ultrasensitive bioluminescence immunoassays for the determination of peptides and proteins (illustrated with human urinary kallikrein, bradykinin and the determination of human urinary kallikrein antibody titres) have been developed. The usable ranges of the standard curves are from 5 pg to 5000 pg per litre. The relative intra-assay coefficients of variation of the tests were between 2% and 6%, and the inter-assay coefficients of variation between 4% and 12%.     

Extensive changes in cytokeratin expression patterns in pathologically affected human gingiva

The stratified squamous epithelium of the oral gingiva and the hard palate is characterized by a tissue architecture and a cytoskeletal composition similar to, although not identical with, that of the epidermis and fundamentally different from that of the adjacent non-masticatory oral mucosa. Using immunocytochemistry with antibodies specific for individual cytokeratins, in situ hybridization and Northern blots of RNA with riboprobes specific for individual cytokeratin mRNAs, and gel electrophoresis of cytoskeletal proteins of microdissected biopsy tissue samples, we show changes in the pattern of expression of cytokeratins and their corresponding mRNAs in pathologically altered oral gingiva. Besides a frequently, although not consistently, observed increase in the number of cells producing cytokeratins 4 and 13 (which are normally found as abundant components in the sulcular epithelium and the alveolar mucosa but not in the oral gingiva) and a reduction in the number of cells producing cytokeratins 1, 10 and 11, the most extensive change was noted for cytokeratin 19, a frequent cytokeratin in diverse one-layered and complex epithelia. While in normal oral gingiva cytokeratin 19 is restricted to certain, sparsely scattered cells of --or near--the basal cell layer, probably neuroendocrine (Merkel) cells, in altered tissue of inflamed samples it can appear in larger regions of the basal cell layer(s) and, in apparently more advanced stages, also in a variable number of suprabasal cells. Specifically, our in situ hybridization experiments show that this altered suprabasal cytokeratin 19 expression is more extended at the mRNA than at the protein level, indicating that cytokeratin 19 mRNA synthesis may be a relatively early event during the alteration. These changes in cytokeratin expression under an external pathological influence are discussed in relation to other factors known to contribute to the expression of certain cytokeratins and with respect to changes occurring during dysplasia and malignant transformation of oral epithelia.