Autoradiographic studies with fatty acids and some other lipids: A review

In this review, we have mainly included studies in which whole-body autoradiography was used. In lipid research, most studies have been done with fatty acids. These studies showed some common characteristics in the pattern of tissue distribution. A major uptake was seen in the brown fat, liver and adrenal cortex but also to some extent in other tissues with a high metabolic activity or high cell turn-over, e.g. the gastric and intestinal mucosa, diaphragm, kidney cortex and bone marrow. Low levels of radioactivity were generally found in the brain, testes, thymus, white fat, skeletal muscles, lungs and spleen. Most fatty acids showed some specific features, e.g the strong uptake of erucic, arachidonic and docosahexaenoic acid in myocardium and of eicosapentaenoic acid in the adrenal cortex. Studies with PGE1 and LTC3 showed that the liver and kidney and to a lesser degree the lungs were the major sites of metabolism. The distribution of free cholesterol and triolein emulsion labelled in the fatty acid moieties did show some similarities with respect to the general pattern found with most fatty acids. Specific for cholesterol was a very strong uptake in the adrenal cortex. There was also a significant uptake in the spleen whereas the uptake in the brown fat was not as marked as for most fatty acids. Specific for triolein was a marked uptake in the spleen and myocardium, in fed animals also in the white adipose tissue. These studies show that whole-body autoradiography can give much valuable information of the uptake and distribution of lipids that would be rather difficult to obtain with conventional methods. Combined with electron-microscopy, autoradiography can be used to study cellular and even subcellular distribution, and thus given further data on the metabolism of lipids in the body.     

High-performance liquid chromatographic identification of beta-lactam antibiotics produced by dermatophytes

Thirteen of 48 dermatophyte isolates were found by bioassay to produce beta-lactam antibiotics and seven produced other antibiotics. Estimation and detection of specific beta-lactams in culture broths by derivatization and HPLC was only possible following concentration and extraction procedures. Analysis of the concentrated broths demonstrated the production of penicillin X and penicillin G by two Trichophyton mentagrophytes strains and by one Microsporum canis strain; one further T. mentagrophytes strain produced only penicillin X. Additions of the beta-lactam side-chain precursors, phenylacetic acid and phenoxyacetic acid, to fermentation media failed to increase the antibiotic titres.     

Critical roles for juvenile hormone and its esterase+ in the prepupa of Trichoplusia ni

In the caterpillar Trichoplusia ni (Lepidoptera: Noctuidae) it has been demonstrated by allatectomy that the appearance of juvenile hormone during the prepupal stage is crucial for the successful larval-pupal ecdysis of most larvae. Application of juvenile hormone or juvenile hormone esterase inhibitors at key times disrupted normal development as well. Thus the subsequent disappearance of juvenile hormone is regulated by degradation by juvenile hormone esterase in addition to a hypothetical reduction in biosynthesis. This reduction in juvenile hormone titer in the prepupa is just as critical for normal development as was its previous appearance. These observations on the critical role of juvenile hormone in the prepupa are in contrast to observations in some other species. For instance, in the case of Manduca sexta (Lepidoptera: Sphingidae), juvenile hormone is considered only supplementary to the action of prothoracicotropic hormone in the postwandering stage and primarily is required for normal pupal development. It thus appears that even within the Lepidoptera the role of juvenile hormone in prepupal development can vary dramatically.     

Flavonoids of the Hydrangeaceae Dumortier

Fourteen species representing nine genera of the Hydrangeaceae Dumortier were surveyed for their flavonoid pigments. All taxa exhibited profiles based upon common flavonols. Myricetin was seen in two genera: Jamesia and Decumaria. Jamesia was further distinguished by the absence of kaempferol or its glycosides. A complex array of 3-O-mono-, 3-O-di- and 3-O-triglycosides was observed, although not all species had all levels of glycosylation. Decumaria barbara was unique within the species studied in its possession of 3,7-di- and 3,7-triglycosides. The overall pattern of flavonol glycosides observed for the Hydrangeaceae closely resembles that found in herbaceous genera of Saxifragaceae. The comparatively low frequency of myricetin contrasts with its high occurrence in herbaceous genera.     

Mapping parathyroid hormone, β-globin,insulin, and LDH-A genes within the human chromosome 11 short arm by spot blotting sorted chromosomes

Summary Rearranged human chromosomes carrying segments of chromosome 11 were separated from the normal chromosome 11 by high-resolution chromosome sorting. Sorted chromosomes were tested with parathyroid hormone, -globin, insulin, and LDH-A gene-specific probes to determine the genes carried by each chromosome segment. Based on the gene content and karyotypes of these abnormal chromosomes, the parathyroid hormone, -globin, insulin, and LDH-A genes and the unique restriction fragment ADJ-762 are all located on the terminal band of the short arm of human chromosome 11 (band 11p15), with LDH-A proximal to the other loci.     

Free radical damage to cultured porcine aortic endothelial cells and lung fibroblasts: Modulation by culture conditions

Summary Culture conditions modulating cell damage from xanthine plus xanthine oxidase-derived partially reduced oxygen species were studied. Porcine thoracic aorta endothelial cells and porcine lung fibroblasts were maintained in monolayer culture. Cells were prelabeled with⁵¹Cr before xanthine plus xanthine oxidase exposure. Endothelial cells showed 30 to 100% more lysis than fibroblasts and thus seemed more sensitive to this oxidant stress. The effect of cell culture age, as indicated by population doubling level (PDL), was examined. Response of low PDL endothelial cells and fibroblasts subjected to oxidant stress was compared with the response of PDL 15 cells. Both low PDL endothelial cells and fibroblasts responded differently to the lytic effect of xanthine oxidase-derived free radicals than did higher PDL cells. Specific activities of the antioxidant enzymes catalase, managanese superoxide dismutase, copper-zinc superoxide dismutase, glutathione peroxidase, and glucose-6-phosphate dehydrogenase were measured in both low and high PDL fibroblasts and endothelial cells. Antioxidant enzyme specific activities could only partially explain the differences in response to oxidant stress between fibroblasts and endothelial cells and between low and high PDL cells. Cell culture medium composition modulated the rate of production, and relative proportions of xanthine plus xanthine oxidase-derived partially reduced species of oxygen, i.e. superoxide, hydrogen peroxide, and hydroxyl radical. Serum content of medium was important in modulating free radical generation; superoxide production rates decreased 32%, H₂O₂ became undetectable, and hydroxyl radical generation decreased 54% in the presence of 10% serum. The medium protein and iron content also modulated free radical generation. The data suggest that cell culture media constituents, cell type, and cell culture age greatly affect in vitro response of cells subjected to oxidant stress. Research supported by American Lung Association Fellowship Training Grant and Research Training Grant, the R. J. Reynolds Corporation, and National Institutes of Health Grants HL29784 and 1 HL 23805.     

Mixing patterns in Amazon lakes

The diel mixing patterns of two small floodplain lakes, Lago Jacaretinga in the Amazon drainage, and Lago Cristalino in the Rio Negro system, were investigated during both the high-water and low-water states of the Amazon River hydrograph. Measurements included temperature, oxygen, ammonia, phosphate, and chlorophyll. In both lakes thermal stratification developed during the day and was eroded at night. During the low-water period when the lakes were shallow, nocturnal circulation extended to the lake bottom, whereas when the lakes were deeper (greater than about 5 m), circulation did not reach the bottom and an anoxic hypolimnion developed. During the low-water period, percent of oxygen concentrations were relatively high but always less than saturation. Low oxygen concentrations were observed during the high-water period. At all times nocturnal mixing supplied a significant amount of oxygen to the lake ecosystems. Nighttime upward mixing of recycled nitrogen and phosphorus also appeared to be important nutrient sources for algal productivity.     

Catalytic properties of the resolved flavoprotein and cytochrome B components of the NADPH dependent O2−• generating oxidase from human neutrophils

The resolved flavoprotein and cytochrome b₅₅₉ components of the NADPH dependent $\text{O}{}_2{}^{\mathrm{?}}{}_{\mathrm{?}}$ generating oxidase from human neutrophils were the subject of further study. The resolved flavoprotein, depleted of cytochrome b₅₅₉, was reduced by NADPH under anaerobic conditions and reoxidized by oxygen. NADPH dependent $\text{O}{}_2{}^{\mathrm{?}}{}_{\mathrm{?}}$ generation by the resolved flavoprotein fraction was not detectable, however it was competent in the transfer of electrons from NADPH to artificial electron acceptors. The resolved cytochrome b₅₅₉, depleted of flavoprotein, demonstrated no measureable NADPH dependent $\text{O}{}_2{}^{\mathrm{?}}{}_{\mathrm{?}}$ generating activity and was not reduced by NADPH under anaerobic conditions. The dithionite reduced form of the resolved cytochrome b₅₅₉ was rapidly oxidized by oxygen, as was the cytochrome b₅₅₉ in the intact oxidase.     

Paramagnetic 1H and 13C NMR studies on cobalt-substituted human carbonic anhydrase I carboxymethylated at active site histidine-200: molecular basis for the changes in catalytic properties induced by the modification

Using bromo[1-13C]acetate to modify N tau of His-200 of human carbonic anhydrase isozyme I leads to the introduction of a useful 13C NMR probe into the active site. To complement our previous diamagnetic NMR studies with this probe, we have now succeeded in directly observing the paramagnetically perturbed resonance of the carboxylate in the cobalt-substituted modified enzyme above pH 8. In the pH range 8-10, the resonance undergoes a pH-dependent slow-exchange process, with the more alkaline form having a much smaller pseudocontact shift and a narrower line width. Below pH 8, the resonance apparently undergoes a very large paramagnetic downfield shift that was estimated by extrapolation. An ionization of approximate pK of 6 appears to control this process. Paramagnetic spin-relaxation studies on the resonance under conditions where it was directly observed yielded distance measurements between the carboxylate carbon and the active site cobalt ion. In inhibitor complexes, this distance was in the range of 5-7 A. In the absence of inhibitors, the distance was approximately 3.0-3.2 A at pH 7.9, consistent with the coordination of the carboxylate to the metal. However, at pH 10, the distance was increased to 4.8 A. These distance determinations were aided by relaxation measurements of a paramagnetically shifted proton resonance at 60-65 ppm downfield assigned by others to a proton of a ligand histidine of metal and confirmed by us to be 5.2 +/- 0.1 A from the metal. Our findings provide a molecular basis for the observed changes in catalytic properties that accompany the carboxymethylation.