Microtubules and cyclic amp in human leukocytes: on the order of things

We have shown previously that the β-adrenergic agonist isoproterenol (2μM) and the phosphodiesterase inhibitor isobutylmethylxanthine (1 mM) produce a much greater increase in cyclic AMP in human leukocytes that have been pretreated with colchicine (or with other agents that affect microtubule assembly) than in control leukocytes. The effects of colchicines were both time- and dose-dependant. These and other data suggested that the generation of cyclic AMP is normally restricted by an intact system of cytoplasmic microtubules. If so, then the same time and dose dependencies might apply to other colchicines-induced changes in leukocyte function. We have now assayed the distribution of concanavalin A (Con A)-receptor complexes on the leukocyte membrane, taking into account that leukocytes competent to assemble microtubules show a uniform distribution of surface- bound Con A whereas microtubule-deficient cells accumulate Con A in surface caps. We have found that the effect of colchicine on capping is also both time- and dose dependent, and that the dose-response relationships conform to those required to increase cyclic AMP levels. These findings provide further evidence that both colchicine-induced Con-A capping and colchicine- induced cyclic AMP generation depend upon the relaxation of constraints normally imposed by cytoplasmic microtubules upon the plasma membrane, which limit, respectively, lateral mobility of the lectin-receptor complexes, and expression of hormone-sensitive adenylate cyclase. Moreover, colchicine-induced Con-A cap formation is not affected even by very large changes in leukocyte cyclic AMP levels. Thus, elevated cyclic AMP levels do not appear to promote the dissolution of microtubules; rather, the dissolution of microtubules permits the generation of increased amounts of cyclic AMP.     

The effect of bromodeoxyuridine and dibutyryl cyclic AMP on the induction of placental alkaline phosphate in human choriocarcinoma cells.

The effect of 5-bromo-2'-deoxyuridine (BrdUrd) and dibutyryl cyclic AMP (Bt2cAMP) on the expression of the placental isoenzyme of human alkaline phosphatase was examined in BeWo choriocarcinoma cells. By using a combination of specific immunoprecipitation and polyacrylamide-gel electrophoresis of cells labelled either metabolically with [35S]methionine or cell-surface-labelled with 125I, both BrdUrd (5 micrograms/ml) and 1 mM-Bt2cAMP were shown to result in the enhanced accumulation of a specific protein. This protein has immunochemical identity and co-electrophoreses with placental alkaline phosphatase in two-dimensional gels. These results clearly demonstrate that the induction of placental alkaline phosphatase activity in choriocarcinoma cells treated with these agents is a consequence of the accumulation of specific enzyme protein rather than of altered catalytic activity.     

The oxygen tolerance of sulfate-reducing bacteria isolated from North Sea waters

Five strains of sulfate-reducing bacteria, previously isolated from North Sea waters and identified asDesulfovibrio vulgaris, were investigated for their abilities to survive in aerobic natural seawater. Viable organisms of all strains were recoverable after exposure to oxygen for more than 72 h. The level of the protective enzymes superoxide dismutase and catalase detectable in these strains and the low rates at which oxygen was reduced probably account at least in part for their considerable abilities to survive in aerobic environments. The autoxidizable nature of cytochromec ₃ and KCN-inhibitable cytochromec oxidase activity present in these bacteria are postulated to act as possible oxygen-scavenging mechanisms analogous to the activity of NADH oxidase present in certain other strict anaerobes.     

Cyclosporin A in cadaveric organ transplantation.

The use of cyclosporin A (CyA) with a protocol designed to avoid the effects of nephrotoxicity resulted in a one-year survival of 86% in recipients of renal allografts from unmatched cadaveric donors. The drug also controlled rejection of liver and pancreatic allografts. It was possible to change patients initially treated with CyA to azathioprine and corticosteroids and vice versa, thus enlarging the potential value of CyA in organ allografting. Of 34 recipients of renal allografts, 29 were currently receiving only CyA as immunosuppressive treatment. Twelve patients never required any adjuvant steroid treatment. These results suggest that CyA is an effective immunosuppressant, and if used with care side effects need not be severe.     

A study of altitudinal zonation in the montane forest belt of Mt. Elgon,Kenya/Uganda

Altitudinal changes in the vegetation of the Montane Forest Belt of Mt. Elgon, East Africa, were investigated by analysis of aerial photographs and by sample plots along two transects, one on a relatively wet, and the other on a relatively dry aspect.Classifications for both tree and herb/shrub plots show that the forests along the two transects become increasingly similar floristically with altitude. The total number of tree species recorded is greater for the transect on the moister aspect. The mean altitudinal range for tree species increases on both transects with altitude, while that for herb/shrub species remains constant. It is suggested that the upper altitudinal limits of many species are determined principally by temperature-related factors, while the degree to which species extend their ranges downwards is much influenced by competition.Structural heterogeneity is particularly well marked at altitudes corresponding to the bamboo zone on the wetter slopes and, even in the absence of bamboo, tree density here is much reduced. it is suggested that thicket-forming species, including bamboo, have enhanced competitive ability in intermediate altitude montane forests.Previously advanced classificatory schemes for montane forest vegetation in eastern Africa are examined in the light of the findings.Nomenclature follows Agnew (1974) and Dale & Greenway (1961) with minor modifications after Chapman & White (1970), Mabberley (1973) and Pennington & Styles (1975).The work was funded by a grant from the Natural Environment Research Council (U.K.). We are grateful to the Government of Kenya and to the Forest Department, Kenya, for permission to work on Mt. Elgon. Dr. A. D. Q. Agnew and Miss C. Kabuye very kindly identified some of the plant specimens. The illustrations were drawn by Mr. J. Shaw and the manuseript was critically read by Dr. D. Riley.     

The subcellular distribution of adenylate and guanylate cyclases in murine lymphoid cells.

Membrane vesicles can be prepared from murine lymphoid cells by nitrogen cavitation and fractionated by sedimentation through nonlinear sucrose density gradients. Two subpopulations of membrane vesicles, PMI and PMII, can be distinguished on the basis of sedimentation rate. The subcellular distribution of adenylate and guanylate cyclases in these membrane subpopulations have been compared with the distribution of a number of marker enzymes. Approximately 20-30% of the total adenylate and guanylate cyclase activity is located at the top of the sucrose gradient (soluble enzyme), the remainder of the activity being distributed in the PMI and PMII fractions (membrane-bound enzyme). More than 90% of the 5'-nucleotidase and NADH oxidase activities detected in lymphoid cell homogenates are located in PMI and PMII fractions, whereas succinate cytochrome c reductase activity is detected only in the PMII fractions. In addition, beta-galactosidase activity is distributed in the soluble and PMII fractions of the sucrose density gradients. On the basis of the fractionation patterns of these various enzyme activities, it appears that PMI fractions contain vesicles of plasma membrane and endoplasmic reticulum, whereas PMII fractions contain mitochondria, lysomes, and plasma membrane vesicles. Approximately 30-40% of the adenylate and guanylate cyclase activities in PMII can be converted to a PMI-like form following dialysis and resedimentation through a second nonlinear sucrose gradient. Adenylate and guanulate cyclases can be distinguished on the basis of sensitivity to nonionic detergents.     

Indoleacetic Acid synthesis in soybean cotyledon callus tissue

Growth of an auxin-requiring soybean cotyledon callus tissue (Glycine max L., Merr. var. Acme) was promoted by tryptophan, tryptamine, indole, indoleacetamide and, to a very slight degree, anthranilic acid. When tryptophan-3-¹⁴C was supplied in the growth medium, labeled indoleacetic acid (IAA) was found in both the tissue and the medium. Medium, from which the cells had been removed, was also found to convert labeled tryptophan to IAA. Soybean callus contained 0.044 μmole/g free tryptophan, but this is apparently not available for conversion to IAA. These results suggest that while exogenously supplied trytophan could elevate a specific internal pool where IAA synthesis occurs some of the growth on a tryptophan medium can be accounted for by external conversion.     

Metabolism of Indole-3-Acetic Acid: II. Oxindole Pathway in Parthenocissus tricuspidata Crown-Gall Tissue Cultures

An indoleacetic acid oxidase preparation from an acetone powder of Parthenocissus tricuspidata crown-gall tissue has been examined. An intermediate in the reaction is 3-hydroxymethyloxindole and nonenzymic conversion of it to 3-methyleneoxindole was observed. Neither reaction mixtures nor 3-methyleneoxindole have any auxin-like activity in Avena or wheat coleoptile bioassays. In vivo studies show that although 53% decarboxylation of indoleacetic acid was observed in 48 hours, only a small amount of 3-methyloxindole could be recovered from the medium. The other decarboxylated products remain to be identified but are not 3-hydroxymethyloxindole or 3-methyleneoxindole.     

Macrophage plasminogen activator: modulation of enzyme production by anti-inflammatory steroids, mitotic inhibitors, and cyclic nucleotides.

Plasminogen activator production by cultured mouse peritoneal macrophages can be modulated in vitro by low concentrations of various pharmacologically active molecules. Glucocorticoid hormones and their synthetic derivatives, as well as cholera toxin, colchicine, and vinblastine markedly inhibit production of this enzyme without affecting other important macrophage functions. The effect of glucocorticoids is of particular interest, both because their relative in vivo anti-inflammatory potencies correlate exactly with their effect on plasminogen activator production in culture and because this effect occurs at near physiological concentrations. In view of the correlations established in other systems between plasminogen activator production and cell migration, we have also examined the age of the macrophages in thioglycollate-induced exudates. Confirming the results of Van Furth and Cohn (1968), we have found that the majority of these cells are young, having recently replicated and arrived in the peritoneal cavity. Using a fibrinagar overlay technique which allowed us to determine the production of plasminogen activator by individual cells. we have found that the majority of these cells produce the enzyme. The potential roles of plasminogen activator in monocyte migration and the relationship of this enzyme to the anti-inflammatory effect of gluccorticoids are correlated and emphasized.     

Streptokinase in central retinal vein occlusion: a controlled clinical trial.

Forty patients with central retinal vein occlusion were allocated at random either to a treatment group given streptokinase followed by anticoagulatns or to a control group given no specific treatment. The two groups, which were each of 20 patients, were broadly similar in respect of clinical and laboratory values and similar in their initial visual acuity. At follow-up ("final" vision) the visual acuity in the treated group was significantly better than in the untreated group. Only one treated patient developed thrombotic glaucoma compared with four controls. Streptokinase may, however, have been responsible for vitreous haemorrhage (and permanent loss of vision) in three patients and hence probably has only a limited role in the treatment of central retinal vein occlusion.