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Biomedical subjects

V Castranova

Publications and source records attributed to V Castranova.

At least 163 records · Page 9Linked to original sources
PubMed1984

Relative effects of asbestos and wollastonite on alveolar macrophages.

Rabbit alveolar macrophages were exposed in culture to chrysotile asbestos, wollastonite, or latex, and the effects on various biochemical and physiological parameters related to cellular viability and fibrogenicity were determined. Exposure of alveolar macrophages to asbestos, wollastonite, or latex for 3 d has no effect on oxygen consumption or cellular volume. However, treatment of alveolar macrophages with as little as 25 micrograms asbestos/ml for 1 d increases lysosomal enzyme release and decreases membrane integrity, i.e., decreases trypan blue exclusion and increases leakage of cytosolic enzymes. In contrast, exposure of alveolar macrophages to wollastonite or latex at 250 micrograms/ml does not induce lysosomal enzyme release or alter membrane integrity even after 3 d of exposure in culture. These data suggest that chrysotile asbestos damages rabbit alveolar macrophages, while wollastonite, a potential substitute for asbestos, is far less cytotoxic.

Animals↗
PubMed1984

Volume-responsive sodium and proton movements in dog red blood cells.

Shrinkage of dog red blood cells (RBC) activates a Na transport pathway that is Cl dependent, amiloride sensitive, and capable of conducting Na-proton counterflow. It is possible to establish transmembrane gradients for either Na or protons and to demonstrate that each cation species can drive reciprocal movements of the other. The nature of the coupling between Na and proton movements was investigated using the fluorescent probe diS-C3(5) and also by an indirect method in which K movements through valinomycin channels were used to draw inferences about the membrane potential. No evidence was found to suggest that the Na-proton pathway activated by shrinkage of dog RBC is a conductive one. By exclusion, it is presumed that the coupling between the counterflow of Na and protons is electroneutral. The volume-activated Na-proton fluxes in dog RBC have certain properties that distinguish them from similar transport pathways in other cell types.

Amiloride↗
PubMed1984

Direct measurement of hydrogen peroxide release from rat alveolar macrophages: artifactual effect of horseradish peroxidase.

Investigators disagree on the amount of hydrogen peroxide (H2O2) released by resting and stimulated alveolar macrophages. The method commonly used to measure H2O2 release involves horseradish peroxidase (HRP)-catalyzed oxidation of scopoletin by H2O2. We describe an artifact in this method that may explain the seemingly inconsistent data reported by other investigators. Release of H2O2 and luminol-catalyzed chemiluminescence are stimulated in rat alveolar macrophages by type II HRP at concentrations normally used in the HRP-scopoletin method. The amount of H2O2 released depends upon the length of time the cells are preincubated at 37.5 degrees C and the time at which type II HRP is added. After stimulation with type II HRP, the cells do not release additional H2O2 upon exposure to zymosan particles. Myeloperoxidase, an alternative catalyst to type II HRP, does not stimulate H2O2 release and, therefore, can be used to measure H2O2 release from rat alveolar macrophages. Using myeloperoxidase, resting H2O2 release is negligible; after zymosan stimulation, 6.14 (+/- 0.87) X 10(-6) nmoles/cell X 10 min is released. In addition, more pure HRP preparations (types VI, VII, VIII, and IX) do not stimulate alveolar macrophages to release H2O2 and can be used to monitor zymosan-induced H2O2 release. As our data indicate that type II HRP stimulates H2O2 release from rat and guinea pig alveolar macrophages, it is not the catalyst of choice for this assay. In conclusion, our data explain the conflicting results found in the literature and indicate that rat alveolar macrophages release minimal amounts of H2O2 at rest and can be stimulated by zymosan.

Animals↗
PubMed1983

Incorporation of [3H]palmitate into disaturated phosphatidylcholines in alveolar type II cells isolated by centrifugal elutriation.

In order to study synthesis of pulmonary surfactant materials, we measured incorporation of [3H]palmitate into disaturated phosphatidylcholines (PC) in alveolar type II cells isolated by centrifugal elutriation. The time course for this process is not linear and, at high external palmitate levels (1 mM), incorporation is maximal in 4-5 h. Incorporation is dependent on extracellular palmitate with a Vmax (at 1 mM) of 1.66 nmol palmitate incorporated into disaturated PC/4.2 X 10(5) cells per 2 h and a K1/2 of 0.1 mM palmitate. Addition of an optimal amount of extracellular choline (0.05 mM) increases Vmax and decreases K1/2 for palmitate. Incorporation of palmitate is dependent upon cell number, inhibited by extracellular Ca2+ and stimulated by external Mg2+. Cholinergic and beta-adrenergic agonists do not increase incorporation. Pulmonary lavage fluid inhibits incorporation of palmitate into disaturated PC, suggesting there is negative feedback involved. Disaturated PC which has been recently synthesized (i.e., over a 2 h period) is broken down intracellularly by type II cells when they are suspended in palmitate-free medium. These results indicate that (1) several factors, such as substrate levels, cell number, Ca2+, Mg2+ and amount of surfactant present, are involved in the regulation of palmitate incorporation into disaturated PC; (2) disaturated PC which has been recently synthesized may be broken down by type II cells; and (3) surfactant synthesis in freshly isolated cells differs slightly from that reported by other investigators in type II cells maintained in primary cell culture.

Animals↗
PubMed1983

Ascorbate uptake by isolated rat alveolar macrophages and type II cells.

Studies were conducted to measure intracellular ascorbate content and to characterize ascorbate uptake in three fractions of isolated rat pneumocytes (i.e., alveolar macrophages, alveolar type II epithelial cells, and another fraction of small pneumocytes that contains neither macrophages nor type II cells). When cells are incubated in medium containing 0.1 mM ascorbate (i.e., the concentration normally found in plasma), intracellular ascorbate concentrations are 3.2 mM in alveolar macrophages and type II cells and 0.9 mM in other lung cells; ascorbate influx is 1.5 nmol . 10(7) cells-1 . h-1 for alveolar macrophages, 0.24 nmol . 10(7) cells-1 . h-1 for type II cells, and very slow in other pneumocytes. Ascorbate influx displays saturation kinetics in both alveolar macrophages (K1/2 = 2 mM; Vmax = 32.2 nmol . 10(7) cells-1 . h-1) and type II cells (K1/2 = 5 mM; Vmax = 14.2 nmol . 10(7) cells-1 . h-1). After correction for differences in the membrane surface areas of these two types of lung cells, the rates for maximum ascorbate influx (Vmax) are similar in alveolar macrophages and type II cells. In addition, ascorbate uptake by alveolar macrophages and type II cells is dependent on metabolic activity and extracellular sodium. In contrast, ascorbate uptake in other lung cells does not exhibit saturation kinetics and is not dependent on metabolism or sodium. Thus alveolar macrophages and type II cells possess an energy-dependent cotransport system for ascorbate and sodium influx. The high ascorbate content and the existence of a specialized transport mechanism for ascorbate uptake may explain the relative resistance of alveolar macrophages and type II cells to oxidant injury.

Animals↗
PubMed1983

Transmembrane potential of isolated rat alveolar type II cells.

Type II cells were isolated from rat lungs by elastase digestion and purified by centrifugal elutriation. The fluorescent dye, Di-S-C3(5), was used as a probe to monitor transmembrane potential (Em) of cells suspended in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered medium. With this technique, the Em of type II cells was estimated to be -27 +/- 2 mV. This resting Em is very close to the equilibrium potential for chloride (-21 mV), which suggests that chloride is passively distributed in type II cells. The resting Em of type II cells is more dependent on the extracellular concentration of potassium (K+) than on external sodium (Na+); i.e., the membrane depolarizes as external sodium is replaced by potassium, suggesting that in unstimulated type II cells the membrane is more permeable to potassium than to sodium. In addition, the resting potential appears to be due, in part, to the activity of a ouabain-sensitive, Na-K pump, which acts to hyperpolarize type II cells. Addition of a membrane perturbant, phorbol myristate acetate (PMA, 10 micrograms/ml), to a type II cell suspension results in an increase in oxygen consumption and membrane depolarization. Both of these responses are sodium dependent and thus appear to be linked to a PMA-induced increase in sodium permeability.

Animals↗
PubMed1983

Transport properties of isolated type II alveolar epithelial cells.

Type II cells are granular cells located in the alveolar epithelium. In addition to the synthesis and secretion of surfactant, these pneumocytes exhibit several other interesting properties. Although type II cells possess a high permeability to sodium, they maintain a low free intracellular sodium concentration by the presence of a Na-K pump. The activity of the Na-K pump is high and can result in substantial net movement of solute and water. Therefore, type II cells may employ this pumping capacity to play a significant role in the transepithelial transport of water. Type II cells are also relatively resistant to oxidant damage and play a role in the regeneration of the alveolar epithelium after oxidant injury. Ascorbate is a known antioxidant that is accumulated by type II cells via a specialized transport system for the uptake of ascorbate and sodium. The presence of this specialized transport system in type II cells and alveolar macrophages may explain in part why these cells are more resistant to oxidant injury than other pneumocytes.

Animals↗
PubMed1982

Inhibition by nonsteroidal antiinflammatory drugs of luminol-dependent human-granulocyte chemiluminescence and [3H]FMLP binding. Effect of sulindac sulfide, indomethacin metabolite, and optical enantiomers (+) and (-) MK830.

A system is described to evaluate for nonsteroidal antiinflammatory drugs by means of luminol-dependent human-granulocyte chemiluminescence (CL) is described. The CL is produced using either opsonized zymosan (yeast cells) or the soluble chemotactic peptide f-Met-Leu-Phe as the perturbant of the granulocyte membrane. Using either system, the following drug effects 2 x 10(-5) M were noted: only sulindac sulfide, and not sulindac sulfone or sulindac, displayed marked inhibition of chemiluminescence, following the in vivo data regarding inflammatory effects. The 5-OH indomethacin metabolite was likewise inactive as an inhibitor of CL mirroring in vivo effects. MK(+)410, MK(-)830 and MK835 all showed approximately 50% inhibition of CL, displaying deviation from in vivo data. MK(+)830 markedly stimulated CL, 4-6 times the control (without drug), which is clearly different from its enantiomer, MK(-)830. The reasons for this behavior are unclear. However, receptor binding studies with [3H]FMLP were accomplished in the presence and absence of the various drugs at 2 x 10(-5) M that were effective inhibitors of chemiluminescence (CL). Indomethacin, MK(-)830 and MK(+)410 had equivalent percent control binding and percent control CL. Sulindac sulfide and MK(+)835 both had higher percent control binding than percent control CL, with MK(+)835 displaying apparent increased numbers of available receptors relative to control. MK(+)830, which produces large increases in CL, produced a minor effect on percent control binding. A direct relationship between binding and CL does not exist with each drug. Chemiluminescence is dependent on ion movement and oxidative metabolism and is a secondary event to agonist-receptor occupation.

Anti-Inflammatory Agents↗
PubMed1982

Volcanic ash: toxicity to isolated lung cells.

Samples of volcanic ash from Mount St. Helens were collected from Spokane, Washington, after the major eruption of May 18, 1980. The toxicity of ash to the lung was estimated by monitoring the effects of in vitro and in vivo exposure on various physiological parameters of isolated lung cells. Volcanic ash had little effect on O2 consumption of rabbit type II pneumocytes, O2 consumption or superoxide release of resting rat alveolar macrophages, or membrane integrity of rat alveolar macrophages. Ash also caused no significant lipid peroxidation in rat lung microsomes. However, volcanic ash did inhibit superoxide anion release from zymosan-stimulated rat alveolar macrophages. Since superoxide is an antibacterial substance, this result suggests that exposure to volcanic ash may adversely affect the ability of alveolar macrophages to protect the lung from infection.

Animals↗
PubMed1982

Ionic content and regulation of cellular volume in rat alveolar type II cells.

Alveolar type II epithelial cells were isolated by elastase digestion of rat lungs and purified by centrifugal elutriation. This method yields an enriched fraction of 1 x 10(7) cells/rat containing 85% pure type II pneumocytes. Purified type II cells exhibit a high rate of oxygen consumption, 215 nmol O2 . 10(6) cells-1 . h-1, which is unaffected by the addition of succinate. Type II cells contain 2 microliters H2O/10(7) cells and are approximately 325-330 micrometers 3 in volume. These pneumocytes contain 107 nmol K/l cell H2O and 70 mmol Cl/l cell H2O. In addition, type II cells have a high Na content, i.e., 156 mmol/l cell H2O. However, most of this Na is bound with only 33% being exchangeable. Therefore, the cytoplasmic concentration of free Na is 51 mmol/l cell H2O. Na uptake is very rapid and type II cells i.e., specific activity equilibrium is reached in 15 min with a half time of 5 min. In addition to this large passive transport of Na, type II cells exhibit a highly active Na-K pump involved in the regulation of cellular volume. Volume regulation is inhibited by a decrease in temperature (2 degrees C) and by the addition of ouabain (10(-4) M), Hg (10(-5) M), or Cu (10(-3) M).

Animals↗
PubMed1982

Multinucleation in alveolar macrophages from rats treated with chlorphentermine.

When rats were treated with chlorphentermine, a cationic amphiphilic drug, a phospholipid storage disorder developed in alveolar macrophages, the severity of which was directly proportional to the duration of treatment over a 4-week period. Concomitantly, a progressively greater percentage of the cells became multinucleated such that, after 4 weeks of drug treatment, 18 per cent of the cells contained more than one nucleus. Greater than 99 per cent of the macrophages from control rats had on nucleus per cell. Associated with the multinucleation was an increase in the DNA, RNA, and protein content of the macrophages and increases in the RNA to DNA and protein to RNA ratios relative to cells from untreated rats. Centrifugal elutriation was employed as a means to study the multinucleation as a function of increasing cell size. At each of the 4 weekly intervals studied, the percentage of cells with two or more nuclei increased as the cells became larger. An increase in multinucleation was also found in cells of any given size range as the time of treatment increased. Possible mechanisms responsible for the multinucleation phenomenon are discussed.

Animals↗
PubMed1981

Abnormal responses of granulocytes in chronic granulomatous disease.

Stimulation of normal granulocytes with chemotactic factor, phorbol myristate acetate, concanavalin A, and calcium ionophore results in rapid depolarization which precedes the 'respiratory burst'. Treatment of granulocytes in chronic granulomatous disease with these stimulants fails to generate chemiluminescence. This defect is associated with an absence of transmembrane potential shifts in response to treatment with chemotactic factor, phorbol myristate acetate, and concanavalin A while depolarization in response to A23187 is unaffected by this disease state.

Calcimycin↗
PubMed1981

Transmembrane potential changes associated with superoxide release from human granulocytes.

Treatment of human granulocytes with concanavalin A, phorbol myristate acetate (PMA), N-formyl-methionyl-leucyl-phenylalanine (FMLP), and A23187 (a calcium ionophore) stimulates the release of superoxide anion and the generation of chemiluminescence. The fluorescent probe, Di-S-C3(5), has been used to monitor shifts in membrane potential in response to these stimulants which precede the secretion of superoxide. Concanavalin A, PMA, and FMLP induce a biphasic shift in transmembrane potential (Em), i.e., a rapid depolarization followed by a prolonged hyperpolarization. This depolarization is dependent on both external sodium and calcium while the hyperpolarization is inhibited by ouabain which blocks the electrogenic Na-K pump. In contrast, A23187 induces a rapid and prolonged depolarization. This monophasic shift in Em is dependent on external calcium. These results suggest that depolarization acts as a signal to initiate events associated with the "respiratory burst" of these phagocytes.

Calcimycin↗
PubMed1981

Transmembrane potential changes during phagocytosis in rat alveolar macrophages.

Studies were carried out to measure changes in the transmembrane potential of rat alveolar macrophages during exposure of the cells to zymosan particles or to the membrane perturbant, phorbol-12-myristate-13-acetate (PMA), and to determine if changes in membrane potential are related to superoxide anion release. Exposure of the cells to either zymosan or PMA leads to membrane depolarization, which precedes superoxide anion release. Furthermore, the magnitude of the depolarization is dependent upon the concentration of either zymosan or PMA. During exposure of the alveolar macrophages to increasing levels of zymosan, there is an increase in the amount of superoxide released as well as an increase in the magnitude of the depolarization. Incubation of the cells in medium containing 150 mM K+, a medium which causes membrane depolarization, leads to superoxide release from resting cells and a decrease in the amount of superoxide released from cells exposed to zymosan. These results indicate that release of superoxide anion from rat alveolar macrophages is related to membrane depolarization and suggest that the transmembrane potential change may act as a signal to initiate the phagocytotic responses of the cells.

Animals↗
PubMed1981

Ascorbate uptake by isolated rat lung cells.

Experiments were done to determine the intracellular concentration of ascorbate in isolated rat lung cells and the concentration in plasma and to study ascorbate influx in these cells. The intracellular ascorbate concentration was 2.25 mM and the plasma level was about 0.14 mM; i.e., the lung cell ascorbate concentration was about 16 times greater than the plasma level. When the cells were incubated in medium containing physiological levels of ascorbate (0.1 mM), influx increased linearly up to 60 min of incubation and was 0.54 +/- 0.04 nmol.10(7) cells-1.h-1. Influx was dependent on the extracellular ascorbate concentration. At concentrations ranging from 0.025 to 1 mM, uptake appeared to exhibit saturation kinetics with an apparent Km of 0.16 mM. At physiological levels of extracellular ascorbate (0.1 mM) at least 90% of the uptake appeared to be carrier mediated, and this influx was inhibited by various metabolic inhibitors. In addition, ascorbate influx was inhibited by ouabain and removal of extracellular sodium. These results suggest that lung cells contain a transport mechanism for ascorbate that is energy-dependent and that may be coupled to Na+ influx.

Animals↗
PubMed1981

Factors which affect superoxide anion release from rat alveolar macrophages.

In order to investigate some of the characteristics of superoxide anion release from alveolar macrophages, the effects of substances known to influence superoxide release from polymorphonuclear leukocytes (PMN) were studied in rat alveolar macrophages. There is a relatively small, but constant, amount of superoxide released from alveolar macrophages at rest. The amount released increases 5- to 6-fold and becomes maximal in about 20-30 min following exposure to unopsonized zymosan particles. The rate of superoxide release is maximal only 2 min after exposure of the cells to particles, i.e., long before particle uptake is complete. In addition to particles, release of superoxide anion can be stimulated by phorbol-12-myristate-13-acetate (PMA). Lectins and chemotactic factors, which stimulate release in PMN, have little or no effect in alveolar macrophages. Superoxide release during exposure to zymosan appears to be dependent upon extracellular Ca++. Also, the release mechanism can be affected by the addition of cyclic AMP or various protein modifiers to the medium. Since many of these findings differ from those reported by others for PMN, the control of superoxide anion release from alveolar macrophages and PMN is probably different.

Animals↗