Incorporation of a charged amino acid into the membrane-spanning domain blocks cell surface transport but not membrane anchoring of a viral glycoprotein.

The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G protein) consists of a continuous stretch of 20 uncharged and mostly hydrophobic amino acids. We examined the effects of two mutations which change the amino acid sequence in this domain. These mutations were generated by oligonucleotide-directed mutagenesis of a cDNA clone encoding the G protein, and the altered G proteins were then expressed in animal cells. Replacement of an isoleucine residue in the center of this domain with a strongly polar but uncharged amino acid (glutamine) had no effect on membrane anchoring or transport of the protein to the cell surface. Replacement of this same isoleucine residue with a charged amino acid (arginine) generated a G protein that still spanned intracellular membranes but was not transported efficiently to the cell surface. The protein accumulated in the Golgi region in about 50% of the cells, and about 20% of the cells had detectable protein levels in a punctate pattern on the cell surface. In the remaining cells the protein accumulated in a vesicular pattern throughout the cytoplasm. Models which might explain the abnormal behavior of this protein are discussed.     

Rapid measurement of creatine kinase activity in a coronary care unit using a portable benchtop reflectance photometer.

Rapid measurements of plasma creatine kinase activity using an inexpensive benchtop reflectance photometer (Ames Seralyzer) and disposable reagent strips were evaluated in the laboratory and coronary care unit. The system proved simple to use and capable of yielding rapid (four minutes per analysis), precise (coefficient of variation less than 9%), and accurate results (correlation with routine method 0.995) when used by medical staff. Creatine kinase values were available 6.5-102 hours earlier than routine laboratory data, depending on the time and day of sampling, thereby facilitating appropriate and economic patient management. This instrument might be used to supplement the routine enzyme service for selected admissions, resulting in greatly improved availability of results and hence contributing to the early discharge of patients from intensive care facilities.     

Potentiation of lung vascular response to endotoxin by superoxide dismutase

We studied the effects of superoxide dismutase (SOD), an enzyme that converts superoxide into peroxide, on the cardiopulmonary response to endotoxin in sheep. Sheep (n = 18) were prepared for chronic measurement of cardiopulmonary variables, including lung lymph flow, by surgically implanting catheters under halothane anesthesia. Nine of the animals were studied before and after the administration of endotoxin (0.75 microgram/kg) with and without SOD. An additional nine animals received SOD without the lipopolysaccharide. Endotoxin produced an increase in lung lymph flow that was initially associated with a marked pulmonary arterial (PA) hypertension and reduced lymph-to-plasma protein ratio (L/P). The lymph flow remained elevated later in the response, but there was only a mild increase in PA pressure, and the L/P was normal. There was also a fall in blood neutrophils and in cardiac index. SOD increased this secondary elevation in lung lymph flow, and the corresponding L/P was greater than the preendotoxin value. The fall in neutrophil count, cardiac output, and the elevation in PA pressure seen with endotoxin were not affected by SOD. When administered in the absence of endotoxin, SOD produced no perceptible change in the cardiopulmonary and lymph values. We conclude that peroxide, hydroxyl ion, and/or other free radicals formed by the action of SOD must be responsible for a portion of the endotoxin response rather than superoxide itself.     

The cytosol origin of macromolecules extruded by cultured chick embryo fibroblast cells

The DNA and protein extruded by chick embryo fibroblast cells has been analysed by chromatography. A high proportion of the DNA is in the form of a protein-complex of size around 5 X 10(5) dalton. The patterns of the DNA and protein extruded into the supernatant are closely similar in many respects to those found in the cell cytosol. It is concluded that the macromolecular material extruded by cells in culture is of cytosol origin: a possible function in terms of "information" carriage is proposed.     

Different hormonal requirements for androgen-independent growth of normal and tumor epithelial cells from rat prostate

Summary The proliferation of isolated normal prostate epithelial cells from rat and man is androgen-independent and requires cholera toxin, insulin, dexamethasone, epidermal growth factor (EGF) and one or more polypeptide factors that are concentrated in bovine neural tissue. The active agents in the neural tissue extract are heparin-binding polypeptides (prostatropins), the predominant form of which has a molecular weight of 17400 and an acetylalanine at the aminoterminus. Prostatropins supported a half-maximal increase in normal prostate epithelial cell number at 50 picomolar. The proliferation of primary and serially-cultured epithelial cells from androgen-responsive Dunning R3327 rat prostate tumors was also androgen-independent, but exhibited dramatic alterations in response to hormones that stimulated normal cell proliferation. At low cell density, androgen-independent growth of isolated tumor-derived epithelial cells was independent on cholera toxin, was stimulated by dexamethasone, required insulin andeither EGFor prostatropin. The presence of either EGF or prostatropin masked the response to the other factor. In the absence of EGF, purified prostatropins supported a half-maximal increase in tumor cell number at 7 picomolar. Endogenous production of EGF-like and prostatropin-like factors or both was suggested by the reduced requirement for EGF and prostatropin at high prostate tumor cell density. These results suggest that anti-hormonal therapies against prostate tumor growth should be based on intervention with the activity of insulin (or insulin-like factors) or simultaneous intervention with both EGF and prostatropin (or their homologues). This work was supported by NIH grants CA 37589 and HL 33847, and grant 1718 from the Council for Tobacco Research. Editor’s Statement This paper is the first report of the comparison of the hormone requirements of primary cultures of normal and tumor prostate epithelial cells from the same system.     

Detection of noncontingent feedback in EMG biofeedback

Noncontingent feedback is frequently used as a placebo control procedure in biofeedback research. Researchers, however, have criticized this procedure for lacking credibility because of easy detection. The present study examined detection of false feedback in biofeedback with EMG. Contingent feedback (CF), truly random false feedback (FF), and controlled false feedback (CFF) groups were compared for changes in EMG levels, report of inaccurate feedback, and report of learning muscle activity reduction. The results indicated that FF procedures are easily detected; therefore, differences found between the FF and CF groups may be influenced by extraneous variables. The CFF group did not detect false feedback, but subjects reported some suspicions in later trials. With more trials, CFF may have also been detected. These results indicate a need for more attention to appropriate placebo control procedures in evaluating the parameters and efficacy of biofeedback.     

Turbine flowmeter vs. Fleisch pneumotachometer: a comparative study for exercise testing

The purpose of this study was to investigate the characteristics of a newly developed turbine flowmeter (Alpha Technologies, model VMM-2) for use in an exercise testing system by comparing its measurement of expiratory flow (VE), O2 uptake (VO2), and CO2 output (VCO2) with the Fleisch pneumotachometer. An IBM PC/AT-based breath-by-breath system was developed, with turbine flowmeter and dual-Fleisch pneumotachometers connected in series. A normal subject was tested twice at rest, 100-W, and 175-W of exercise. Expired gas of 24-32 breaths was collected in a Douglas bag. VE was within 4% accuracy for both flowmeter systems. The Fleisch pneumotachometer system had 5% accuracy for VO2 and VCO2 at rest and exercise. The turbine flowmeter system had up to 20% error for VO2 and VCO2 at rest. Errors decreased as work load increased. Visual observations of the flow curves revealed the turbine signal always lagged the Fleisch signal at the beginning of inspiration or expiration. At the end of inspiration or expiration, the turbine signal continued after the Fleisch signal had returned to zero. The "lag-before-start" and "spin-after-stop" effects of the turbine flowmeter resulted in larger than acceptable error for the VO2 and VCO2 measurements at low flow rates.     

Inspiratory neuron activity in the ventrolateral medulla of the dog

The purpose of this study was to describe the distribution and activity pattern of respiratory neurons located in the ventrolateral medulla (VLM) of the dog. Spike activity of 129 respiratory neurons was recorded in 23 ketamine-anesthetized spontaneously breathing dogs. Pontamine blue dye was used to mark the location of each neuron. Most VLM neurons displaying respiratory related spike patterns were located in a column related closely to ambigual and retroambigual nuclei. Both inspiratory and expiratory neurons were present with inspiratory units being grouped more rostrally. The predominant inspiratory neuron firing pattern was "late" inspiratory, although eight "early" types were located. All expiratory firing patterns were the late expiratory variety. Each neuron burst pattern was characterized by determining burst duration (BD), spikes per burst (S/B), peak frequency (PF), time to peak frequency (TPF), rate of rise to peak frequency (PF/TPF), and mean frequency. CO2-induced minute ventilation increases were associated with decreases in BD and TPF and increases in PF, S/B, and PF/TPF. In 11 experiments the relative influences of vagotomy and tracheal occlusion on late inspiratory units were compared. Tracheal occlusion increased late inspiratory BD and S/B but did not alter PF/TPF. Vagotomy increased BD and S/B beyond those obtained by tracheal occlusion and, in some neurons, decreased the PF/TPF. We conclude that the location of respiratory units in the VLM of the dog is similar to that in other species, the discharge pattern of VLM respiratory units is similar to those in cat VLM, and vagotomy and tracheal occlusion affect discharge patterns differently.     

Chromosome 8 breakpoint far 3'' of the c-myc oncogene in a Burkitt''s lymphoma 2;8 variant translocation is equivalent to the murine pvt-1 locus.

The 2;8 variant translocation of human Burkitt's lymphomas is closely related cytogenetically to the t(6;15) of murine plasmacytomas; both involve a reciprocal exchange between the Ig kappa locus and a band region indistinguishable from that bearing the c-myc oncogene. To define their molecular relationship, we have compared cloned chromosome 8 DNA from the t(2;8) breakpoint in the human Burkitt's lymphoma JBL2 with cloned DNA from the murine pvt-1 locus, the major chromosome 15 breakpoint region in murine t(6;15). DNA sequencing and Southern blot analysis shows that these two regions are homologous. Thus the t(2;8) in JBL2 is the molecular equivalent of many murine t(6;15). The murine pvt-1 locus lies an unknown distance 3' of c-myc; analysis of DNA from several tumours with c-myc amplification reveals that pvt-1 is co-amplified in at least one case, placing pvt-1 approximately 100-500 kb 3' of c-myc. The significance of these results with respect to the role of pvt-1 in tumorigenesis is discussed.     

Cell envelope phospholipid changes in a moderate halophile during phenotypic adaptation to altered salinity and osmotic stress

Abstract The increased content of negatively-charged phospholipids in membranes of Vibrio costicola grown at high salinities is mediated by increased phospholipid synthesis of phosphatidylglycerol relative to phosphatidylethanolamine. This phenomenon provides a system for investigating the factors involved in triggering and controlling haloadaptation in this moderately halophilic bacterium. We review recent experiments, which show that when subjected to sudden increases in external salinity, V. costicola senses both the absolute NaCl concentration and the magnitude of the salt shift. We show that the latter is sensed at least in part via osmotic pressure effects, since shift-up into sucrose-containing media triggers comparable changes in growth and in phospholipid composition and synthesis.