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Biomedical subjects

M F Tsan

Publications and source records attributed to M F Tsan.

At least 55 records · Page 3Linked to original sources
PubMed1990

Tracheal insufflation of tumor necrosis factor protects rats against oxygen toxicity.

Tracheal insufflation of tumor necrosis factor (TNF; 5 micrograms or 1.2 x 10(5) U) markedly enhanced the survival of adult rats exposed to 100% O2: 12 of 17 rats (71%) survived for greater than 11 days, whereas 30 of 30 control (Hanks' balanced salt solution) insufflated rats (100%) died within 3 days of O2 exposure. Insufflation of gamma-interferon (5 micrograms) or intraperitoneal injection of up to 40 micrograms TNF did not afford any protection. At 55 h after O2 exposure, TNF-insufflated rats showed less pulmonary edema, as determined by the extravascular lung water content-to-bloodless lung dry weigh ratio and less alveolar capillary leak as determined by the protein content in the bronchoalveolar lavage fluid, than control insufflated rats similarly exposed. This protection against O2 toxicity by TNF insufflation was associated with increased lung superoxide dismutase, catalase, and glutathione peroxidase activities. The enhancement of lung antioxidant enzyme activities was noted at 55 h of O2 exposure, when control animals began to die of O2 toxicity. This temporal relationship suggests that TNF-induced increase in antioxidant enzyme activities contributes, at least in part, to the observed protection.

Animals↗
PubMed1989

Phallacidin prevents thrombin-induced increases in endothelial permeability to albumin.

Calf pulmonary artery endothelial monolayers cultured on polycarbonate filters were utilized to study 125I-labeled albumin permeability and actin filament distribution in response to thrombin challenge. Thirty-minute exposure to alpha-thrombin (10(-7) M) significantly increased albumin clearance rates. These changes were associated with marked alterations in actin filament distribution, resulting in loss of peripheral actin bands and an increase in the number of cytoplasmic stress fibers. Because the actin peripheral filaments are thought to play an important role in junctional stability, we postulated that stabilization of actin filaments should protect against thrombin-induced barrier disruptions. Pretreatment of cells with 0.3 microM 7-nitrobenz-2-oxa-1,3-diazole (NBD)-phallacidin, a specific actin-stabilizing agent, prevented the changes in actin filament distribution and markedly attenuated the increase in albumin permeability. Because of the potential toxicity of phallatoxins, we evaluated the effects of pretreatment on cell viability and growth parameters. There were no differences in viability, seeding efficiency, or doubling times in cells treated with 0.3 microM NBD-phallacidin in comparison to controls. Our data support the hypothesis that actin filaments, particularly peripheral bands, contribute significantly to the maintenance of barrier function in cultured endothelial cells.

Actins↗
PubMed1989

Protein kinase inhibitor prevents pulmonary edema in response to H2O2.

We investigated the effect of H2O2 (92 microM) in isolated guinea pig lungs perfused with a buffered Ringer solution. Pulmonary arterial pressure (Ppa), pulmonary capillary pressure (Ppc), and change in lung weight (delta W) were recorded at 0 min and at 15, 30, and 60 min after the H2O2. The capillary filtration coefficient (Kfc) was measured at 0 and 30 min. The perfusion of H2O2 increased the Ppa, Ppc, delta W, and Kfc. The thromboxane synthetase inhibitor Dazoxiben, or the vasodilator papaverine, prevented the increases in Ppa and Ppc. The protein kinase C (PKC) inhibitor H7 [1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride] prevented the increases in Ppa, Ppc, delta W, and Kfc, whereas the inactive isoquinoline HA1004 [N-(2-guanidinoethyl)-5-isoquinolinesulfonamide hydrochloride] had little effect on the H2O2 response. H2O2 increased the number of stress fibers and disrupted the peripheral band of cultured confluent endothelial cells, changes that were prevented with pretreatment with H7. PKC may mediate the increases in vascular permeability and pulmonary edema that occur in response to H2O2.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Modulation of endothelial GSH concentrations: effect of exogenous GSH and GSH monoethyl ester.

We studied the effects of exogenous glutathione (GSH) and GSH monoethyl ester (GSH-MEE) on the enhancement of endothelial GSH concentrations. The preparation of GSH-MEE used contained 91% GSH-MEE, approximately 9% GSH diethyl ester (GSH-DEE) and a trace amount of GSH. Both GSH and GSH-MEE markedly stimulated the intracellular concentrations of GSH in endothelial cells. GSH-MEE was more potent than GSH. The enhancement of endothelial GSH concentration by exogenous GSH was completely inhibited by buthionine sulfoximine (BSO), a potent inhibitor of gamma-glutamylcysteine synthase, or acivicin (AT-125), an inhibitor of gamma-glutamyl transpeptidase, suggesting that it was due to the extracellular breakdown and subsequent intracellular resynthesis of GSH. In contrast, the effect of GSH-MEE was largely resistant to BSO and acivicin, suggesting that it was primarily due to transport of GSH-MEE followed by intracellular hydrolysis. The GSH-MEE preparation, which contained 9% GSH-DEE, at concentrations of 2 mM or higher caused vacuolization of endothelial cells. The enhancement of GSH concentrations by exogenous GSH, but not by GSH-MEE, protected endothelial cells against H2O2-induced injury.

Animals↗
PubMed1989

Relation between myocardial glutathione content and extent of ischemia-reperfusion injury.

The relation between the extent of myocardial injury sustained during reperfusion and total glutathione (GSH) content in the ischemic myocardium was examined in anesthetized open-chest pigs subjected to coronary occlusion for 45 minutes and reperfusion for 2 hours. In pigs infused with saline during reperfusion (n = 6) there was a decrease in myocardial GSH content from 380 +/- 48 micrograms/g in normally perfused myocardium to 182 +/- 36 micrograms/g in the ischemic reperfused myocardium (p less than 0.02). Myocardial infarct size (expressed as a percentage of the ischemic area) was 12.5 +/- 0.8%. There was a delay of recovery of contractile function before returning to 60% of preocclusion value. In pigs pretreated with buthionine sulfoximine (BSO) (n = 5), an inhibitor of cellular GSH synthesis, there was reduction in GSH content to 215 +/- 25 micrograms/gm in normally perfused myocardium and to 77 +/- 8 micrograms/gm in the ischemic reperfused myocardium. The extent of injury was greater as evidenced by an increase in infarct size to 30.4 +/- 4.0% (p less than 0.001), severe destructive changes in subepicardial ultrastructure, which were absent in saline-infused pigs, and persistence of dyskinesia throughout reperfusion. In pigs infused with glutathione intravenously (0.8 gm/kg) at a rate of 6.5 mg/kg/min (n = 6), 5 minutes before and continuously during reperfusion, there was an increase in GSH content to 582 +/- 67 micrograms/g in normally perfused myocardium and to 312 +/- 80 micrograms/g in ischemic reperfused myocardium. The increase in myocardial GSH was associated with a reduction in infarct size to 7.5 +/- 1.3% (p less than 0.05, compared with saline-infused pigs) and an early recovery of contractile function of the ischemic myocardium. GSH infusion into pigs pretreated with BSO (n = 4) failed to increase myocardial GSH content and failed to reduce the extent of myocardial injury. Thus, the extent of myocardial injury sustained during reperfusion is very dependent on the effectiveness of its antioxidant defenses. Markedly increased susceptibility to injury occurs when the GSH content in the ischemic myocardium becomes depleted.

Animals↗
PubMed1988

L-2-oxothiazolidine-4-carboxylate protects cultured endothelial cells against hyperoxia-induced injury.

When bovine pulmonary artery endothelial monolayers were exposed to hyperoxia (95% O2 and 5% Co2), they responded by selectively elevating the intracellular concentration of glutathione without affecting the activities of glutathione peroxidase or glutathione reductase, L-2-Oxothiazolidine-4-carboxylate, an intracellular cysteine-delivering agent, further enhanced the intracellular concentration of glutathione in oxygen-exposed endothelial cells and protected them from the lethal effect of hyperoxia. In contrast, buthionine sulfoximine, a potent inhibitor of gamma-glutamylcysteine synthetase, reduced the glutathione concentration and rendered the cells more sensitive to the toxic effect of oxygen. Both L-2-oxothiazolidine-4-carboxylate and buthionine sulfoximine had no effect on the activities of glutathione peroxidase or glutathione reductase. Our results suggest that L-2-oxothiazolidine-4-carboxylate may have the potential of preventing oxygen toxicity.

Cells, Cultured↗
PubMed1988

Hyperoxia causes increased albumin permeability of cultured endothelial monolayers.

When confluent calf pulmonary arterial endothelial monolayers cultured on polycarbonate micropore membranes were exposed to hyperoxia (95% O2) for 3 days, endothelial cells became enlarged, and their permeability to 125I-labeled albumin was markedly increased. Similar changes were not observed when endothelial monolayers were exposed to hyperoxia for 1 or 2 days. Cell counting and acridine orange staining of endothelial monolayers revealed that the hyperoxia-induced increase in albumin permeability was not associated with a denuding injury or loss of cells from the monolayers. Vimentin filament staining of O2-exposed monolayers showed thickening of the perinuclear vimentin coil in some cells. Rhodamine-phalloidin staining demonstrated that hyperoxia caused a progressive alteration in the actin distribution. Two days after O2 exposure, peripheral actin bands became thinner, whereas the number of cytoplasmic stress fibers was increased. Three days after O2 exposure, peripheral actin bands of most cells were disrupted or absent. Because peripheral actin bands play an important role in maintaining the integrity of endothelial monolayers, disruption of peripheral bands by hyperoxia may in part be responsible for the observed change in permeability.

Actins↗
PubMed1988

Direct staining and visualization of endothelial monolayers cultured on synthetic polycarbonate filters.

Endothelial and epithelial cells cultured on synthetic filter supports have been used to study permeability and transport under various experimental conditions. However, because of the non-transparent nature of these filters, morphological studies using light microscopy are not possible. Presently, investigators circumvent this problem by using cells cultured on glass coverslips, extrapolating morphological data from a system clearly different from that used for functional studies. We describe here a useful technique for direct staining and visualization of cells grown on polycarbonate filter supports, using fluorochrome probes and fluorescence microscopy. We have utilized acridine orange, rhodamine phalloidin, and an anti-vimentin monoclonal antibody to provide information about cell shape, monolayer configuration, and cytoskeletal protein distribution in cultured calf pulmonary artery endothelial cell monolayers. Comparison staining of coverslip and filter preparations revealed a number of clear differences in these parameters. This technique should enable investigators to perform the necessary studies to obtain direct correlations between functional and morphological data.

Acridine Orange↗
PubMed1988

Red blood cells protect endothelial cells against H2O2-mediated but not hyperoxia-induced damage.

We studied the effect of intact red blood cells on the exogenous H2O2-mediated damage as well as on the hyperoxia-induced injury of cultured endothelial cells. Red blood cells protected endothelial cells against H2O2-mediated injury efficiently, but had no effect on the hyperoxia-induced damage. Failure of red blood cells to protect endothelial cells against hyperoxia-induced injury was not due to hemolysis. Furthermore, hyperoxia-exposed red blood cells were still capable of protecting endothelial cells against H2O2-mediated damage.

Animals↗
PubMed1988

Effect of hyperoxia on the cytoarchitecture of cultured endothelial cells.

When confluent pulmonary artery endothelial cells in culture were exposed to hyperoxia (95% O2 and 5% CO2), they became enlarged and mean corpuscular volume increased 30-35%. Rhodamine-phalloidin staining of actin filaments demonstrated that hyperoxia was associated with a progressive alteration in the actin distribution. Three days after oxygen exposure, the number and thickness of cytoplasmic stress fibers were increased, while the peripheral bands were disrupted or absent. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the amount of filamentous actin was increased in oxygen-exposed cells, while the total actin content remained unchanged, suggesting that oxygen exposure shifted the equilibrium from G actin to F actin.

Actins↗
PubMed1987

Should platelets be radiolabelled in plasma medium?

Because of the low labelling efficiency of platelets with In-111 oxine in plasma containing media, most investigators have labelled platelets in plasma-free media. However, labelling of platelets with In-111 oxine in plasma-free media may result in either: irreversible damage leading to rapid clearance from the circulation and permanent hepatic sequestration; or reversible damage with transient hepatic sequestration without affecting the in vivo survival. Since human platelets can be labelled with In-111 oxine in plasma in sufficient efficiency for routine clinical use, they should be labelled in plasma whenever possible. The use of newer In-111 chelates, tropolone and mercaptopyridine-N-oxide, which label platelets with high efficiency in plasma as well as in plasma-free media, may offer another alternative.

Animals↗
PubMed1987

Cyclosporin-A for the treatment of pure red cell aplasia in a patient with chronic lymphocytic leukemia.

A 62-year-old man with B-cell chronic lymphocytic leukemia had three separate episodes of pure red cell aplasia (PRCA). The last episode was treated with cyclosporin-A (CyA) and prednisone. After the patient was on the therapy for 2 weeks, erythropoietic recovery was observed and with continued therapy the hematocrit (Hct) became normal. The PRCA remission was associated with a fall in the blood lymphocyte count, and a reduction in the spleen and lymph node size and bone marrow lymphocyte density. At diagnosis of PRCA the blood T-cells bearing IgG Fc receptors (T gamma cells) were increased, and the marrow contained very few or no late-stage erythroid progenitors. After remission of PRCA the T gamma cell fraction decreased, and the marrow erythroid progenitor's number became normal. We speculate that therapy with CyA and prednisone inhibited the production of interleukins-1 and -2 from monocytes and T-cells, respectively, and was responsible for the reduction of the T gamma cell fraction and B-cell leukemic mass in this patient. Further, we believe that normalization of T gamma cells in association with the therapy was responsible for the PRCA remission.

Cyclosporins↗
PubMed1987

Modulation of intracellular glutathione concentrations alters lymphocyte activation and proliferation.

Glutathione (GSH) has been implicated in lymphocyte activation and differentiation, as well as in protection from radiation damage. Since [3H]thymidine ([3H]TdR) at high concentrations in the nucleus causes radiation damage to the cells, it is important to rule out the possibility that changes in [3H]TdR uptake by mitogen-activated lymphocytes are not caused by 3H-induced cell injury following alterations in intracellular GSH concentration. In this study, flow-cytometric analysis of cell cycle was used to measure lymphocyte activation. Intracellular GSH levels were enhanced using 2-L-oxothiazolidine-4-carboxylate (OTC) and 2-mercaptoethanol (2ME), which deliver cysteine intracellularly, and suppressed by buthionine sulfoximine (BSO) which inhibits gamma-glutamylcysteine synthetase. Enhancement of intracellular GSH concentrations in lymphocytes with 2-oxothiazolidine-4-carboxylate or 2-mercaptoethanol augments mitogen-induced lymphocyte activation, and proliferation, while suppression of intracellular GSH levels by buthionine sulfoximine inhibits the progression of cellular proliferation--but not activation, as measured by flow cytometry. There was a linear relationship between intracellular GSH concentration and conA-activated cells by flow cytometry and between GSH concentration and [3H]TdR incorporation as measured at 24 h. We conclude that alterations of intracellular GSH concentrations may be one way to modulate lymphocyte activation and differentiation.

Acridine Orange↗
PubMed1987

Effects of chelates and incubation media on platelet labeling with indium-111.

We studied the effects of various [111In]chelates and incubation media on labeling efficiency (LE) and in vivo survival of platelets. High LE of human and rabbit platelets in plasma were obtained with [111In]tropolone and [111In]mercaptopyridine-N-oxide. Indium-111 oxine in plasma resulted in a moderate LE and required a longer incubation time, while [111In]oxine sulfate had low LE and inconsistent labeling. High LE for all forms of [111In]chelates were achieved in labeling media free of plasma. However, in vivo platelet survival in rabbits was markedly reduced when platelets were labeled in the absence of plasma.

Animals↗
PubMed1987

Glutathione and lymphocyte activation: a function of ageing and auto-immune disease.

A decline in tissue and serum of glutathione (GSH) content and GSH-metabolizing enzymes with age has been implicated in the increasing susceptibility to carcinogens, disease and drugs which occurs with advanced age. Immunological senescence has been directly associated with increased incidence of cancer and infection with age. The auto-immune diseases of rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) demonstrate depressed T-cell function together with B-cell hyperactivity. In addition, RA and SLE are chronic inflammatory conditions which have been associated with low serum and erythrocyte GSH concentrations when compared to normal. We hypothesized that augmentation of intracellular GSH concentrations in lymphocytes may enhance immune function in depressed immune states. Our data, using murine animal models for ageing (C57BL/6J) and the RA/SLE-like auto-immune diseases of the MRL/lpr mouse, indicate that intracellular glutathione of splenic lymphocytes does not decline with age or with a chronic inflammatory auto-immune disease. In contrast, immune responsiveness in splenic lymphocytes does decline. We can, however, augment both intracellular GSH concentrations and the immune response of splenic lymphocytes from animals of all ages as well as in those animals with the SLE-like auto-immune disease.

Aging↗
PubMed1986

Organ distribution and fate of human platelets: studies of asplenic and splenomegalic patients.

Little is known about the organ distribution and fate of human platelets. We investigated the kinetics, organ distribution, and fate of autologous 111In-oxine-labeled platelets in 12 normal volunteers, four asplenic subjects, and four patients with splenomegaly. The initial recovery of infused 111In-platelets from the circulation was 97.8 +/- 9.8% (means +/- SD) for asplenic subjects and 26.3 +/- 5.9% for splenomegalic patients as compared to 59.2 +/- 9.3% for normal controls. The mean platelet survival times as derived from the multiple-hit model were 9.2 +/- 1.0 days for asplenics and 6.2 +/- 0.6 days for splenomegalic subjects (8.4 +/- 0.8 days for normals). At 30 min postinfusion, 79.4 +/- 19.2% of the infused 111In-platelets pooled in the spleen of splenomegalic subjects and 42.7 +/- 12.2% in normal controls. There was 7.1 +/- 2.0, 12.6 +/- 3.7, and 29.3 +/- 8.4% pooling in the liver of splenomegalic, normal, and asplenic subjects, respectively. At 10 days postinfusion, 37 and 24% of the 111In-platelets were sequestered in the spleen and liver of normal control subjects, respectively. Similar figures for splenomegalic subjects were 71 and 14%, respectively. In asplenic subjects, 89% was sequestered in the liver. We conclude that spleen and liver are the primary sites of platelet destruction, accounting for 61% of infused 111In-platelets in normal volunteers and 85% in splenomegalics, while the liver is the primary site of platelet destruction, accounting for 89% in asplenic subjects.

Aged↗
PubMed1986

Enhancement of intracellular glutathione promotes lymphocyte activation by mitogen.

We have examined the effect of chemically modulating intracellular glutathione (GSH) levels on murine lymphocyte activation. Lymphocyte activation was determined by the induction of polyamine synthesis (ornithine decarboxylase (ODC) induction) and DNA synthesis ([3H]thymidine([3H]Tdr) incorporation). Intracellular GSH levels were enhanced using L-2-oxothiazolidine-4-carboxylate (OTC), which delivers cysteine intracellularly, and suppressed by buthionine sulfoximine (BSO), which inhibits gamma-glutamylcysteine synthetase. In addition, the thiol 2-mercaptoethanol (2-ME) was tested for its ability to augment intracellular GSH levels. Our results indicate that both OTC and 2-ME enhance GSH concentrations and [3H]Tdr incorporation in resting and mitogen (concanavalin A)-stimulated cells. The induction of ODC by concanavalin A (Con A) was augmented by the addition of OTC or 2-ME. The GSH concentration of Con A-stimulated cells was reduced when compared to resting cells; however, it was markedly enhanced by OTC or 2-ME. The stimulatory effects of 2-ME on GSH concentrations, [3H]Tdr incorporation, and ODC induction in both resting and Con A-stimulated cells were much more potent than those of OTC. In contrast, BSO suppressed intracellular GSH and [3H]Tdr incorporation in resting and Con A-stimulated cells. BSO also inhibited the promotion of intracellular GSH concentrations and [3H]Tdr uptake by OTC or 2-ME. However, BSO did not affect the induction of ODC by Con A or its enhancement by OTC or 2-ME. We conclude that enhancement of intracellular GSH concentration results in an increased lymphocyte response to mitogen stimulation.

Animals↗