Oxygenation of malignant tumors after localized microwave hyperthermia

Summary The oxyhemoglobin saturation (HbO₂) of single red blood cells within tumor microvessels (diameter: 3–12 µm) of DS-Carcinosarcoma was studied using a cryophotometric micromethod. In untreated control tumors (mean tissue temperature approx. 35° C) the measured values scattered over the whole saturation range from zero to 100 sat.%, the mean being 51 sat.%. Upon heating at 40° C for 30 min, the oxygenation of the tumor tissue significantly improved as compared with control conditions. After 40° C-hyperthermia a mean oxyhemoglobin saturation of 66 sat.% was obtained. In contradistinction to this, after 43° C-hyperthermia the tumor oxygenation was significantly lower and reached a mean HbO₂ saturation value of 47 sat.%. A further temperature rise to 45° C caused the oxygenation to drop drastically (mean oxyhemoglobin saturation value: 24 sat.%). This is due to a severe restriction of nutritive blood flow.The changes in tumor oxygenation after hyperthermia seem to be predominantly mediated through changes in tumor blood flow, including tumor microcirculation, which showed a similar temperature dependence. Metabolic effects probably play a minor role in the oxyhemoglobin saturation distribution within tumor microvessels.Supported by the Deutsche Forschungsgemeinschaft (Va 57/2-1). Presented in part at the International Symposium on Biomedical Thermology, June 30 to July 4, 1981, Strasbourg, France     

Storage of embryonically transcribed poly(A) RNA and its utilization during metamorphosis of the hydroidHydractinia echinata

Summary The formation of tentacles and stolons during metamorphosis is severely disturbed if inhibitors of mRNA metabolism are applied during certain phases of development. The periods of sensitivity to -amanitin are late gastrulation and the disk stage of metamorphosis. A cordycepin sensitive phase exists during the first hour of metamorphosis. In all drug sensitive phases an enhanced poly(A) synthesis is found indicating increased mRNA metabolism in these stages. Pulse-chase experiments show that planula larvae store a poly(A)-rich RNA population sedimenting between 28–18s. These long living molecules are of embryonic origin, are located in RNP particles and are degraded during metamorphosis. The particles in question appear to be stored mainly in interstitial cells. In early metamorphosis no uridine is incorporated but labelled poly(A) is added to preexisting molecules.     

Crystal and molecular structure of the sulfhydryl protease calotropin DI at 3·2 Å resolution

The three-dimensional structure of the sulfhydryl protease calotropin DI from the madar plant, Calotropis gigantea, has been determined at 3·2 Å resolution using the multiple isomorphous replacement method with five heavy atom derivatives. A Fourier synthesis based on protein phases with a mean figure of merit of 0·857 was used for model building. The polypeptide backbone of calotropin DI is folded to form two distinct lobes, one of which is comprised mainly of α-helices, while the other is characterized by a system of all antiparallel pleated sheets. The overall molecular architecture closely resembles those found in the sulfhydryl proteases papain and actinidin.Despite the unknown amino acid sequence of calotropin DI a number of residues around its active center could be identified. These amino acid side-chains were found in a similar arrangement as the corresponding ones in papain and actinidin. The polypeptide chain between residues 1 and 18 of calotropin DI folds in a unique manner, providing a possible explanation for the unusual inability of calotropin DI to hydrolyze those synthetic substrates that papain and actinidin act upon.     

Isolation of an Hfr donor of Pseudomonas aeruginosa PAO by insertion of the plasmid RP1 into the tryptophan synthase gene

Summary A derivative of the IncP-1 plasmid RP1, temperature-sensitive for maintenance, was inserted into the Pseudomonas aeruginosa chromosome by selection for a plasmid marker (carbenicillin resistance) at nonppermissive temperature. In one strain, PAO 1000, the plasmid was stably integrated in the trpA, B gene cluster mapped at 27 min, as shown by the following evidence. (i) Trp⁺ transductants lost all plasmid markers. (ii) Cleared lysates of PAO 1000 showed no plasmid band typical of the autonomous RP1 in agarose gel electrophoresis. (iii) No transfer of carbenicillin resistance by PAO 1000 was detectable. (iv) PAO 1000 mobilised the chromosome from an origin at, or very near, the plasmid insertion site with high frequency (recovery of proximal markers 10^–3 per donor). Matings on the plate with and without interruption of conjugation showed that chromosome transfer was unidirectional. (v) Recombinants from PAO 1000-mediated crosses did not inherit plasmid markers or the trpA, B mutation. A derivative of PAO 1000 was obtained which had lost the Hfr property and all plasmid markers except carbenicillin resistance. This strain (PAO 1001), when carrying the autonomous RP1 plasmid, was capable of unidirectional chromosome mobilisation like PAO 1000, but with 50-fold lower efficiency. We propose that integration of the temperature-sensitive RP1 plasmid in PAO 1000 occurred via transposition of Tnl, the element specifying carbenicillin resistance.     

Classification of amphipod compound eyes — The fine structure of the ommatidial units (Crustacea,Amphipoda)

Summary The ultrastructure of the compound eyes of 13 amphipod species has been investigated. An amphipod type of compound eye can be characterized by the constellation and consistency of a number of morphological features, most of which are also found in other compound eyes. The amphipod eye falls into four sub-categories (types). The ampeliscid type has a tripartite aberrant lens eye; the lysianassid type has a reduced or no dioptric apparatus and a hypertrophied rhabdom; the hyperid type possesses a large number of ommatidial units with long crystalline cones and dark instead of reflecting accessory pigment; and finally, the gammarid type can be interpreted as a generalized amphipod type. The lysianassid type is adapted to low light intensities and demonstrates convergent development with the compound eyes of other deep-sea crustaceans. The ampeliscid type is more similar to the gammarid type. The type characterization of the amphipod compound eye might well serve as a basis and incentive for functional studies also revealing adaptational mechanisms.This paper is dedicated to Professor Erik Dahl on his 65th birthday and retirement from the Chair of Structural Zoology, Department of Zoology, University of LundThe investigation has been supported by grants from the Swedish Natural Science Research Council (Grants 2760-009 and 009-43). Our thanks are due to the staffs of the marine biological stations in Espegrend (Norway) and Kristineberg (Sweden) and of the research vessel Jean Charcot, Brest, France. The skilled technical assistance of Mrs. Rita Wallén and Miss Maria Walles is gratefully acknowledged     

Modulation of the G0 to S phase transit time by insulin: Potential involvement of protein phosphorylation

BHK fibroblasts can be growth arrested by incubation in low serum (0.1%) medium. Growth is initiated by incubating cells in high serum (10%) medium. We have found that if the quiscent cells in low serum medium are incubated with insulin, the G₀ to S transit time is decreased by two to six hours when serum (10%) is added back to the culture. The effect of insulin treatment of quiescent cells on the cellular phosphoprotein profile was examined. It was found that insulin stimulated the phosphorylation of a 96,000 dalton cytosol protein. This protein is also intensely phosphorylated in proliferating cells and may be one of the critical intracellular events to occur when a cell initiates growth.     

Effect of dietary phosphate intake on phosphate transport by isolated rat renal brush-border vesicles

Renal brush-border membrane vesicles isolated from rats kept for 6-8 weeks on a low-phosphate diet (0.15% of dry matter) showed a markedly faster Na(+)-dependent phosphate uptake than did membrane vesicles isolated from animals kept on a high-phosphate diet (2% of dry matter). Phosphate-uptake rate by brush-border membrane vesicles isolated from animals on a low-phosphate diet remained significantly increased after acute parathyroidectomy. Dietary adaptation was also observed in animals that had been parathyroidectomized before exposure to the different diets. In animals on the low-phosphate diet parathyrin administration inhibited phosphate uptake by brush-border vesicles only if the animals were repleted with P(i) (5ml of 20mm-NaH(2)PO(4)) 1h before being killed. After acute phosphate loading and parathyrin administration the difference in the transport rate between the two dietary groups remained statistically significant. The results suggest that the adaptation of proximal-tubule phosphate transport to dietary intake of phosphate is reflected in the Na(+)/phosphate co-transport system located in the luminal membrane of the proximal-tubule cell. Since the dietary effects on phosphate transport by brush-border membranes are only partially reversed by acute changes in parathyrin concentration and are also observed in chronically parathyroidectomized animals, the adaptation of the Na(+)/phosphate co-transport system to dietary phosphate intake seems to involve an additional mechanism independent of parathyrin.     

Metabolic activation and hepatotoxicity. Toxicity of bromobenzene in hepatocytes isolated from phenobarbital-and diethylmaleate-treated rats.

Hepatocytes freshly isolated from diethylmaleate-treated rats exhibited a markedly decreased concentration of reduced glutathione (GSH) which increased to the level present in hepatocytes from nontreated rats upon incubation in a complete medium. When bromobenzene was present in the medium, however, this increase in GSH concentration upon incubation was reversed and a further decrease occurred that resulted in GSH depletion and cell death. This was prevented by metyrapone, an inhibitor of the cytochrome P-450-linked metabolism of bromobenzene. Bromobenzene metabolism in hepatocytes was accompanied by a fraction of metabolites covalently binding to cellular proteins. The size of this fraction, relative to the amount of total metabolites, was increased in hepatocytes isolated from diethylmaleate-treated rats and in hepatocytes from phenobarbital-treated rats incubated with bromobenzene in the presence of 1,2-epoxy-3,3,3-trichloropropane, an inhibitor of microsomal epoxide hydrase which, however, also acted as a GSH-depleting agent. In addition, the metabolism of bromobenzene by hepatocytes was associated with a marked decrease in various coenzyme levels, including coenzyme A, NAD(H), and NADP(H). Cysteine and cysteamine inhibited the formation of protein-bound metabolites of bromobenzene in microsomes, but did not prevent bromobenzene toxicity in hepatocytes when added at higher concentrations to the incubation medium (containing 0.4 mm cysteine). Methionine, on the other hand, did not cause a significant effect on bromobenzene metabolism in microsomes and prevented toxicity in hepatocytes, presumably by stimulating GSH synthesis and thereby decreasing the amount of reactive metabolites available for interaction with other cellular nucleophiles. It is concluded that, in contrast to hepatocytes with normal levels of GSH, hepatocytes from diethylmaleate-treated rats were sensitive to bromobenzene toxicity under our incubation conditions. In this system, bromobenzene metabolism led to GSH depletion and was associated with a progressive decrease in coenzyme A and nicotinamide nucleotide levels and a moderate increase in the formation of metabolites covalently bound to protein. Methionine was a potent protective agent which probably acted by enhanced GSH synthesis via the formation of cystathionine.