ALBERT

Description: BACKGROUND: The alkaline phosphatase (AP) substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-(3H)-quinazolinone (ELF((R))-97 for enzyme-labeled fluorescence) has been found useful for the histochemical detection of endogenous AP activity and AP-tagged proteins and oligonucleotide probes. In this study, we evaluated its effectiveness at detecting endogenous AP activity by flow cytometry. METHODS: The ELF-97 phosphatase substrate was used to detect endogenous AP activity in UMR-106 rat osteosarcoma cells and primary cultures of chick chondrocytes. Cells were labeled with the ELF-97 reagent and analyzed by flow cytometry using an argon ultraviolet (UV) laser. For comparison purposes, cells were also assayed for AP using a Fast Red Violet LB azo dye assay previously described for use in detecting AP activity by flow cytometry. RESULTS: The ELF-97 phosphatase substrate effectively detected endogenous AP activity in UMR-106 cells, with over 95% of the resulting fluorescent signal resulting from AP-specific activity (as determined by levamisole inhibition of AP activity). In contrast, less than 70% of the fluorescent signal from the Fast Red Violet LB (FRV) assay was AP-dependent, reflecting the high intrinsic fluorescence of the unreacted components. The ELF-97 phosphatase assay was also able to detect very low AP activity in chick chondrocytes that was undetectable by the azo dye method. CONCLUSIONS: The ELF-97 phosphatase assay was able to detect endogenous AP activity in fixed mammalian and avian cells by flow cytometry with superior sensitivity to previously described assays. This work also shows the applicability of ELF-97 to flow cytometry, supplementing its previously demonstrated histochemical applications. Copyright 1999 Wiley-Liss, Inc.

Description: We present an overview of the Long Duration Exposure Facility (LDEF) induced activation measurements. The LDEF, which was gravity-gradient stabilized, was exposed to the low Earth orbit (LEO) radiation environment over a 5.8 year period. Retrieved activation samples and structural components from the spacecraft were analyzed with low and ultra-low background HPGe gamma spectrometry at several national facilities. This allowed a very sensitive measurement of long-lived radionuclides produced by proton- and neutron-induced reactions in the time-dependent, non-isotropic LEO environment. A summary of major findings from this study is given that consists of directionally dependent activation, depth profiles, thermal neutron activation, and surface beryllium-7 deposition from the upper atmosphere. We also describe a database of these measurements that has been prepared for use in testing radiation environmental models and spacecraft design.

Description: HIV-1 infection is often complicated by central nervous system (CNS) dysfunction. Degenerative neuronal changes as well as neuronal loss have been documented in individuals with AIDS. Feline immunodeficiency virus (FIV) infection of cats provides a model for both the immune and the central nervous system manifestations of HIV infection of humans. In this study we have examined neurons in the frontal cortex of feline immunodeficiency virus-infected cats and controls for immunoreactivity with SMI 32, an antibody recognizing a non-phosphorylated epitope on neurofilaments. We noted a significant increase in the number of immunoreactive pyramidal cells in infected animals compared to controls. The changes seen in the neuronal cytoskeleton as a consequence of the inoculation with FIV were similar to those seen in humans undergoing the normal aging process as well as those suffering from neurological diseases, including Alzheimer's and dementia pugilistica. The changes we noted in the feline brain were also similar to that reported in animals with traumatic injuries or with spontaneously occurring or induced motor neuron diseases, suggesting that the increase in reactivity represents a deleterious effect of FIV on the central nervous system.

Description: The primate cingulate gyrus contains multiple cortical areas that can be distinguished by several neurochemical features, including the distribution of neurofilament protein-enriched pyramidal neurons. In addition, connectivity and functional properties indicate that there are multiple motor areas in the cortex lining the cingulate sulcus. These motor areas were targeted for analysis of potential interactions among regional specialization, connectivity, and cellular characteristics such as neurochemical profile and morphology. Specifically, intracortical injections of retrogradely transported dyes and intracellular injection were combined with immunocytochemistry to investigate neurons projecting from the cingulate motor areas to the putative forelimb region of the primary motor cortex, area M1. Two separate groups of neurons projecting to area M1 emanated from the cingulate sulcus, one anterior and one posterior, both of which furnished commissural and ipsilateral connections with area M1. The primary difference between the two populations was laminar origin, with the anterior projection originating largely in deep layers, and the posterior projection taking origin equally in superficial and deep layers. With regard to cellular morphology, the anterior projection exhibited more morphologic diversity than the posterior projection. Commissural projections from both anterior and posterior fields originated largely in layer VI. Neurofilament protein distribution was a reliable tool for localizing the two projections and for discriminating between them. Comparable proportions of the two sets of projection neurons contained neurofilament protein, although the density and distribution of the total population of neurofilament protein-enriched neurons was very different in the two subareas of origin. Within a projection, the participating neurons exhibited a high degree of morphologic heterogeneity, and no correlation was observed between somatodendritic morphology and neurofilament protein content. Thus, although the neurons that provide the anterior and posterior cingulate motor projections to area M1 differ morphologically and in laminar origin, their neurochemical profiles are similar with respect to neurofilament protein. This suggests that neurochemical phenotype may be a more important unifying feature for corticocortical projections than morphology.

Description: The restoration of body fluid balance following dehydration induced by exercise will occur through regulatory responses which stimulate ingestion of water and sodium ions. A number of different afferent signalling systems are necessary to generate appropriate thirst or sodium appetite. The primary sensory information of naturally occurring thirst is derived from receptors sensing cell volume and the volume of the extracellular fluid compartment. Sensory information from the oropharyngeal region is also an important determinant of thirst. The interaction of these various afferent signalling systems within the central nervous system determines the extent of fluid replacement following dehydration.

Description: This study quantified and compared the cross-sectional and longitudinal influence of age, self-report physical activity (SR-PA), and body composition (%fat) on the decline of maximal aerobic power (VO2peak) of women. The cross-sectional sample consisted of 409 healthy women, ages 20-64 yr. The 43 women of the longitudinal sample were from the same population and examined twice, the mean time between tests was 3.7 (+/-2.2) yr. Peak oxygen uptake was determined by indirect calorimetry during a maximal treadmill test. The zero-order correlation of -0.742 between VO2peak and %fat was significantly (P 〈 0.05) higher then the SR-PA (r = 0.626) and age correlations (r = -0.633). Linear regression defined the cross-sectional age-related decline in VO2peak at 0.537 ml.kg-1.min-1.yr-1. Multiple regression analysis (R = 0.851) showed that adding %fat and SR-PA and their interaction to the regression model reduced the age regression weight of -0.537, to -0.265 ml.kg-1.min-1.yr-1. Statistically controlling for time differences between tests, general linear models analysis showed that longitudinal changes in aerobic power were due to independent changes in %fat and SR-PA, confirming the cross-sectional results. These findings are consistent with men's data from the same lab showing that about 50% of the cross-sectional age-related decline in VO2peak was due to %fat and SR-PA.

Description: The Biomass Production Chamber at John F. Kennedy Space Center is a closed plant growth chamber facility that can be used to monitor the level of biogenic emissions from large populations of plants throughout their entire growth cycle. The head space atmosphere of a 26-day-old lettuce (Lactuca sativa cv. Waldmann's Green) stand was repeatedly sampled and emissions identified and quantified using GC-mass spectrometry. Concentrations of dimethyl sulphide, carbon disulphide, alpha-pinene, furan and 2-methylfuran were not significantly different throughout the day; whereas, isoprene showed significant differences in concentration between samples collected in light and dark periods. Volatile organic compounds from the atmosphere of wheat (Triticum aestivum cv. Yecora Rojo) were analysed and quantified from planting to maturity. Volatile plant-derived compounds included 1-butanol, 2-ethyl-1-hexanol, nonanal, benzaldehyde, tetramethylurea, tetramethylthiourea, 2-methylfuran and 3-methylfuran. Concentrations of volatiles were determined during seedling establishment, vegetative growth, anthesis, grain fill and senescence and found to vary depending on the developmental stage. Atmospheric concentrations of benzaldehyde and nonanal were highest during anthesis, 2-methylfuran and 3-methylfuran concentrations were greatest during grain fill, and the concentration of the tetramethylurea peaked during senescence.

Description: Transforming growth factor-beta1 (TGF-beta) is secreted in a latent form consisting of mature TGF-beta noncovalently associated with its amino-terminal propeptide, which is called latency associated peptide (LAP). Biological activity depends upon the release of TGF-beta from the latent complex following extracellular activation, which appears to be the key regulatory mechanism controlling TGF-beta action. We have identified two events associated with latent TGF-beta (LTGF-beta) activation in vivo: increased immunoreactivity of certain antibodies that specifically detect TGF-beta concomitant with decreased immunoreactivity of antibodies to LAP. Macrophages stimulated in vitro with interferon-gamma and lipopolysaccharide reportedly activate LTGF-beta via cell membrane-bound protease activity. We show through dual immunostaining of paraformaldehyde-fixed macrophages that such physiological TGF-beta activation is accompanied by a loss of LAP immunoreactivity with concomitant revelation of TGF-beta epitopes. The induction of TGF-beta immunoreactivity colocalized with immunoreactive betaglycan/RIII in activated macrophages, suggesting that LTGF-beta activation occurs on the cell surface. Confocal microscopy of metabolically active macrophages incubated with antibodies to TGF-beta and betaglycan/RIII prior to fixation supported the localization of activation to the cell surface. The ability to specifically detect and localize LTGF-beta activation provides an important tool for studies of its regulation.

Description: Tomato (Lycopersicon esculentum Mill.) cvs. Red Robin (RR) and Reimann Philipp (RP) were grown hydroponically for 105 d with a 12 h photoperiod, 26 C/22 C thermoperiod, and 500 micromol/ sq m/s PPF at either 400, 1200, 5000, or 10,000 micromol/mol (0.04, 0.12, 0.50, 1.00 kPa) CO2. Harvested fruits were analyzed for proximate composition, total dietary fiber, nitrate, and elemental composition. No trends were apparent with regard to CO2 effects on proximate composition, with fruit from all treatments and both cultivars averiging 18.9 % protein, 3.6 % fat, 10.2 % ash, and 67.2 % carbohydrate. In comparison, average values for field-grown fruit are 16.6 % protein, 3.8 % fat, 8.1 % ash, and 71.5 % carbohydrate (Duke and Atchely, 1986). Total dietary fiber was highest at 10,000 micromol/mol (28.4 % and 22.6 % for RR and RP) and lowest at 1000 micromol/mol (18.2 % and 15.9 % for RR and RP), but showed no overall trend in response to CO2. Nitrate values ranged from 0.19 % to 0.35 % and showed no trend with regard to CO2. K, Mg, and P concentrations showed no trend in response to CO2, but Ca levels increased from 198 and 956 ppm in RR and RP at 400 micromol/mol, to 2537 and 2825 ppm at 10,000 micromol/mol. This increase in Ca caused an increase in fruit Ca/P ratios from 0.07 and 0.37 for RR and RP at 400 micromol/molto 0.99 and 1.23 for RR and RP at 10,000 micromol/mol suggesting that more dietary Ca should be available from high CO2-grown fruit.

Description: No abstract available