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W Jessup

Publications and source records attributed to W Jessup.

87 records · Page 5Linked to original sources

Membrane proteins are critical targets in free radical mediated cytolysis.

The hypothesis that proteins are critical targets in free radical mediated cytolysis was tested using U937 mononuclear phagocytes as targets and iron together with hydrogen peroxide to generate radicals. Those conditions which, after a lag of approx. 30 min, led to drastic lysis were also associated with very rapid membrane depolarisation. Conversely, when the early membrane depolarisation was prevented (by the addition of chelator and catalase), so was lysis. A similar correlation between early membrane depolarisation and subsequent lysis was also observed when the cells were exposed to a toxin from Actinobacillus actinomycetemcomitans. Those conditions of radical attack which led to lysis normally caused substantial lipid peroxidation. However, depolarisation and subsequent lysis were not prevented even when lipid peroxidation was completely suppressed by exogenous antioxidant. ATP levels were not grossly affected within the critical first 30 min period. These data exclude lipids and ATP as the target for lytic damage. We argue therefore that proteins are probably amongst the primary targets in cytolysis by radicals.

Adenosine Triphosphate↗

Effects of exogenous amines on mammalian cells, with particular reference to membrane flow.

We have reviewed the evidence that amines accumulate in intracellular vesicles of low pH, such as lysosomes and endosomes. There is consequent elevation of intravesicular pH, and inhibition of receptor-ligand dissociation often results from this pH change. We have argued that the capacity for fusion of such vesicles is also reduced by the high pH. We suggest that the variety of effects of amines on membrane flow and macromolecular transport we describe are at least partly due to such reduced fusion (Figs. 1 and 2). We propose that an internal low pH may facilitate heterologous vesicle-vesicle and vesicle-plasma membrane fusion. There is some evidence that clathrin can accelerate phospholipid vesicle fusion in vitro at low pH (Blumenthal et al., 1983) but no direct evidence on the role of intravesicular pH. This idea is consistent not only with the preceding discussion, but also with the fact that the intracellular membrane-bound compartments least involved in fusion events (e.g. mitochondria) are of neutral or alkaline internal pH. Membrane fusion is certainly required for the formation of vesicles at the periphery of the Golgi apparatus, and possibly earlier in the transport and processing of biosynthetic products in the Golgi (Bergeron et al., 1982). Thus the accumulation of amines in the Golgi may be responsible for several effects on the flow of macromolecules along their translocation pathways. The status of the plasma membrane in this view is complex. It might be argued that the pH dictating the fusion step in endocytosis is that of the extracellular fluid, in which case the inhibitory effects of amines on this process are not explained. However, the rapidity of acidification of the newly formed endocytic vesicles allows the possibility that plasma membrane invaginations might temporarily sequester areas which are of lower pH than that of the bulk extracellular fluid even before fusion, since the proton pumping enzyme(s) are probably present on the plasma membrane. Were this the case, then an acid pH could again be a factor determining membrane fusion at the plasma membrane. The inhibition of endocytosis by weak bases thus may again reflect elevation of pH in a sequestered compartment. From the data on the dependence of response on the concentration of amines, we anticipate that most responses involving membrane flow will be biphasic, with inhibitory effects at low amine concentration, giving way to stimulatory ones at higher concentrations. We suggest that the reported dichotomy between different amines in intracellular membrane fusion systems (D'Arcy Hart, 1982) may result from this concentration dependence.(ABSTRACT TRUNCATED AT 400 WORDS)

Amines↗

Inhibition of some spontaneous secretory processes in macrophages and fibroblasts by ammonium chloride.

Ammonium chloride inhibits the spontaneous secretion of lysozyme by mouse macrophages and the murine macrophage-like cell line P388D1 and the spontaneous secretion of lysosomal enzymes by P388D1 cells, normal human fibroblasts and the hypersecretory mucolipidosis II (I-cell) fibroblasts. NH4Cl can also inhibit protein synthesis, but this is shown to occur mainly at higher concentrations, or after longer periods of exposure, than are needed for the inhibition of the spontaneous secretory processes. It is confirmed that this amine can also interfere with the continuous endocytosis of fluid in the murine cell types. The nature of the inhibitions is discussed.

Ammonium Chloride↗

Lysosomal hydrolases in macrophages exposed to swainsonine.

Swainsonine reversibly inhibits macrophage lysosomal acid alpha-mannosidase in vitro. When supplied to cultured cells for periods of up to 24 h, swainsonine penetrates the cells and produces a dose- and time-dependent inhibition of cellular alpha-mannosidase. Exposure of macrophages to swainsonine for 24 h, followed by continued incubation in the absence of this agent, produces elevated cellular activity of alpha-mannosidase, relative to unexposed controls; prolonged incubation of macrophage cultures with swainsonine for 1-2 weeks results also in significant increases in cell protein, lactate dehydrogenase activity and in that of another lysosomal enzyme, beta-hexosaminidase.

Alkaloids↗

Endocytosis and intracellular protein degradation in cystic fibrosis fibroblasts.

Normal rates of pinocytosis of [3H]sucrose were measured in cystic fibrosis fibroblasts, and were not affected by the addition of cystic fibrosis serum. Bulk protein degradation (a significant proportion of which occurs intralysosomally following autophagy) and its regulation by growth state was apparently identical in normal and cystic fibrosis cultures.

Cell Line↗

Lysosomal enzyme secretion by cystic fibrosis fibroblasts is normal.

Lysosomal enzyme secretion by skin fibroblasts derived from cystic fibrosis homozygotes and normal donors of the same range of age, sex and culture passage number were compared. No differences were detected between these groups in their rates of spontaneous lysosomal enzyme release. Incubation of either normal or cystic fibrosis fibroblasts with cystic fibrosis serum did not alter these secretion rates. The lysosomal enzyme secretion of both normal and cystic fibrosis fibroblasts responded similarly to incubation with NH4Cl, monensin, A23187, trifluoperazine or Cytochalasin B. It was concluded that lysosomal enzyme hypersecretion, which has previously been suggested to occur in cystic fibrosis, is unlikely to provide a reliable diagnostic test for the disease.

Acetylglucosaminidase↗

The effect of weak bases on lysosomal enzyme secretion by mononuclear phagocytes.

NH4Cl induces a dose-dependent secretion of lysosomal enzymes by mouse peritoneal macrophages and human peripheral blood monocytes. The mechanism of NH4Cl-stimulated hexosaminidase release is distinct from that initiated by the inflammatory stimulus, zymosan. The spontaneous lysosomal secretion of the continuous murine macrophage-like cell line, P388D1, is inhibited by up to 50% in the presence of NH4Cl and other weak bases.

Ammonium Chloride↗

Spontaneous lysosomal enzyme secretion by a murine macrophage-like cell line.

Lysosomal enzyme secretion by the murine macrophage-like cell line, P388D1, was compared with that of normal peritoneal macrophages. Unlike macrophages, lysosomal hydrolase secretion by P388D1 cells occurred spontaneously in vitro and was not further stimulated by the presentation of inflammatory agents such as zymosan and asbestos.

Animals↗