Alterations in cell surface galactosyltransferase activity during limb chondrogenesis in brachypod mutant mouse embryos

The autosomal mutation brachypod (bpH/bpH) in the mouse affects the development of precartilage mesenchymal condensation in the limb-bud. We have previously shown that this defect is localized to the expression of terminal N-acetylglucosamine (GlcNAc) glycoproteins in the plasma membrane (Elmer and Wright, '83). The present study is focused on cell surface galactosyltransferase (GalTase), an ectoenzyme that transfers galactose to its GlcNAc substrate. Purified plasma membrane preparations derived from wild-type (+/+), heterozygote (+/bpH) and brachypod (bpH/bpH) embryonic mouse limb cells were assayed for GalTase activity during in vitro and in utero chondrogenesis using High-Performance Liquid Chromatography (HPLC). On embryonic day E12, prior to overt expression of the mutant gene, no significant difference in GalTase activity was observed. By the third day in culture, all major chondrogenic elements of the autopod were present in +/+ and +/bpH embryos, whereas the mutant autopods were markedly deficient in staining and appeared consistently shorter. The accumulation of alcianophilic cartilage matrix in the wild-type was accompanied by a 29% increase in GalTase activity, which reflected the net change (29%) observed during development from days E12 to E13 in utero. The GalTase activity for the in utero E13 mutant (13%) was significantly different from control. In culture, day E12 mutant autopods actually decreased in their GalTase level by 3 days so that the activity was reduced to only 57% of the wild-type. Though GalTase activity in the heterozygote showed an intermediate expression, optical image analysis did not reveal consistent differences in cartilage development when compared to +/+, arguing against a gene-dosage effect at the gross anatomical level. These data indicate that an increase in plasma membrane GalTase activity is a natural developmental event that occurs during limb-bud chondrogenesis and a decrease in GalTase activity contributes to the dysmorphogenesis in brachypod limb-buds.     

Metabolism of Some Anionic Tallow-based Detergents by Sewage Microorganisms

A method in which the test detergent was the sole source of carbon was used to study the metabolism of several tallow-based detergents. These were tallow alcohol sulfates, long-chain ether alcohol sulfates, and esters of alpha-sulfo fatty acids. Sodium p-(1-methylundecyl)benzenesulfonate (LAS) was used as a reference material. The alcohol sulfates were the most rapidly and completely metabolized (96 to 99%), and one ether alcohol sulfate was 94% degraded. The other compounds were metabolized to the extent of 61 to 87%; LAS was 80% degraded. Except for the alcohol sulfates, loss of methylene blue activity (MBAS) occurred long before the chemical oxygen demand (COD) values had reached a minimum; with the alcohol sulfates, MBAS and COD decreased simultaneously.     

Toxicity of some goldenrods

This study was performed to determine the biological activity in mice of eight species of Solidago (goldenrod) and to determine some of the chemical groups present in these species. Biologically active substances were present in all of the species tested. Tests of six species were positive for alkaloids. All but one of the species contained demonstrable saponins. All of the species contained tannins, whereas six or seven species were positive for flavonoids. Quaternary bases were not found in any of the species tested. A comparison of the biological activity in mice of these species of Solidago, extracts of which were injected intraperitoneally, showed their descending order of activity to be: (1)S. flexicaulis L., (2)S. hispida Muhl., (3)S. juncea Ait., (4)S. serotina Ait., (5)S. canadensis L., (6)S. rugosa Ait., (7)S. uliginosa Nutt., and (8)S. squarrosa Muhl.     

Synergy of human neutrophils with fluconazole in killing Candida species

The killing of Candida species by human neutrophils in a long-term 24-h assay and possible synergy with fluconazole (FCZ) for killing was investigated. The test medium (TM) consisted of RPMI-1640, penicillin and streptomycin (P/S), and 10% fresh autologous serum. TM alone was highly fungistatic for Candida species compared to TM without serum. When neutrophils were cocultured in TM with Candida species for 24 h the inoculum colony-forming units (CFU) were always significantly reduced (killing) by 58 to 99%. FCZ was tested over a range of 1–500 g/ml, and though almost always fungistatic itself, it synergized with neutrophils for significantly increased killing of C. albicans (isolate Sh27) (P<0.01) and C. albicans (isolate 94-20) (P<0.05). Killing of non-albicans Candida species was so efficient in the absence of FCZ that demonstration of synergy with FCZ was difficult.     

Study of the role of iron in the anticryptococcal activity of human serum and fluconazole

Anticryptococcal activity of human serum and apotransferrin in RPMI 1640 was studied in vitro. The effects of varying concentrations of FeCl₃ on this activity was investigated. Possible synergy of serum and apotransferrin with fluconazole was also measured. The fungistatic activity of human serum, whether lyophilized, stored at 4 °C, fresh frozen or purchased from commercial sources vs. Cryptococcus neoformans was comparable. There was no significant loss of fungistatic activity after freezing and thawing the serum up to 10 times. The fungistatic activity of human serum was similar when tested in different tissue culture media with the exception of Medium 199. The addition of apotransferrin (2.0 or 0.2 mg/ml) to RPMI 1640 had an inhibitory effect on cryptococcal growth. This effect was reversed by 20 M of FeCl₃ at both apotransferrin concentrations. By contrast, addition of FeCl₃ to human serum and RPMI 1640 did not reverse inhibition of growth. Fluconazole synergized with the human serum preparations described, but not with pooled commercial serum, for fungicidal activity. Synergistic activity of fluconazole and human serum was not affected by the addition of FeCl₃. Apotransferrin did not show any synergistic fungicidal activity with fluconazole.     

The effects of nutrient addition and pH manipulation in bag experiments on the phytoplankton of a small acidic lake in West Virginia,USA

In situ bag experiments were performed during summer and autumn in a small acidic lake, Tibbs Run Lake, West Virginia, USA. The objective was to evaluate phytoplankton responses to pH manipulation and nutrient addition. Increasing the pH from below 4.5 to over 6.3 resulted in great declines in phytoplankton biovolume. There was also a succession from dinoflagellates (Peridinium inconspicuum to small chlorophytes. The trend was more rapid where phosphorus (P) additions were made along with pH enhancement. During summer, P limitation was indicated, while nitrogen (N) appeared to limit production in autumn. In both seasons, nutrient additions greatly altered the phytoplankton composition in high pH treatments, but had no discernable effects at (the natural) low pH. A low pH, P addition treatment in autumn was the single exception. When N was subsequently added, phytoplankton composition changed dramatically, probably because the proceeding P additions caused severe secondary N-limitation. In general, however, the results supported the view that phytoplankton compositional responses to nutrient additions are suppressed in low pH, relative to high pH lake water.     

Electron paramagnetic resonance studies of membrane proteins in hepatic microsomes

Hepatic microsomal membranes, prepared under various conditions that yield either ‘intact’ or ‘disrupted’ microsomal vesicles, have been labeled via the sulfhydryl groups of intrinsic membrane proteins using nitroxide analogs of $\text{N-}\text{ethylmaleimide}$. Electron paramagnetic resonance spectra revealed the presence of two dominant classes of bound label corresponding to differing degrees of immobilization, the ratio of which were quantitated using a parameter designated the ‘$\text{W/S}$’ ratio. For latent microsomes, the value of this parameter was determined to be $\text{0.65 ± 0.02}$ and was influenced by factors such as label/protein ratio, incubation period, nitroxide structure, temperature and pH. The $\text{W/S}$ ratio was also sensitive to the degree of membrane integrity as revealed by the latency of mannose 6-phosphate activity of glucose-6-phosphohydrolase. In addition, membrane disruption resulted in a corresponding decrease in the order parameter for nitroxide-labeled fatty acids intercalated within the lipid bilayer. The $\text{W/S}$ ratio was observed to be dependent upon the method of microsome preparation yielding values of $\text{1.02 ± 0.02}$ for ‘hypertonically disrupted’ vesicles and $\text{1.28 ± 0.02}$ for ‘mechanically disrupted’ vesicles. Microsomal marker enzymes such as cytochrome $\text{P-450}$ and FAD-containing monooxygenase retained significant levels of functionally following nitroxide incorporation.     

Biliary excretion of warfarin metabolites and their metabolism by rat gut flora.

The bile was determined to be the major excretory route for ¹⁴C-warfarin in the rat with approximately 10% of the administered dose excreted within 5 hours after injection. Relatively little radioactivity appeared in the faces, indicating that considerable enterohepatic recycling was taking place. Less than 4% of the radioactivity in the bile could be extracted with organic solvents. Following incubation of the bile with β-glucuronidase or rat gut flora, however, the bulk of the radioactivity was extractable. The extent of gut flora-mediated hydrolysis of these polar biliary warfarin conjugates was approximately the same as obtained with β-glucuronidase. Approximately $\text{1}\text{3}$ of the gut flora-hydrolyzable conjugates were labile and subject to nonenzymatic hydrolysis at 37 C for 24 h. Chromatographic examination of the extracts from β-glucuronidase-treated bile revealed the presence of warfarin and 4′-, 6-, 7-, and 8-hydroxywarfarin. Warfarin and 7-hydroxywarfarin were the most abundant metabolites in the extracts.     

Hip Fracture,Skeletal Fragility,Osteoporosis and Hormonal Deprivation in Elderly Women

Fractures of the hip have been shown to have a significant personal and societal impact in Western countries; this impact is largely borne by elderly women, and represents a substantial health care commitment in modern society. For many people a fracture of the proximal end of the femur represents a preterminal event of considerable cost, both in economic loss and psychosocial well-being. These fractures are generally recognized as a clinical complication of osteoporosis, and are one index of general skeletal fragility which is also manifested in fractures of the vertebrate and of the distal radius (Colles fracture).There is increasing evidence that hormonal deprivation in elderly women is directly related to loss of skeletal integrity and consequent fragility. There is also increasing evidence that hormonal substitution is effective in preventing this structural loss and fragility. Unfortunately, a therapeutic dilemma has arisen in that the preparation that seems to give optimal protection, conjugated estrogens, has also been reported to cause an increased incidence of endometrial carcinoma. The search for a preparation or dosage regimen of estrogens which simultaneously prevents skeletal atrophy and fragility and avoids the increased risk of malignancy must be a long-term goal.