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A M Das

Publications and source records attributed to A M Das.

At least 37 records · Page 2Linked to original sources
PubMed1998

The role of lipocortin-1 in the inhibitory action of dexamethasone on eosinophil trafficking in cutaneous inflammatory reactions in the mouse.

1. The ability of glucocorticosteroids to inhibit tissue eosinophilia may be an important feature of their anti-inflammatory action in allergic diseases. Our previous work showed that an effect of dexamethasone on the release of eosinophils from the bone marrow could explain its inhibitory action on eosinophil accumulation in a mouse air-pouch model. Thus, it was unclear from that study whether dexamethasone could interfere with the process of eosinophil trafficking. In the present study, therefore, we used a newly developed mouse model to evaluate the effects of systemic treatment with dexamethasone on the recruitment of (111)In-labelled blood eosinophils to sites of cutaneous inflammation in the mouse and whether lipocortin-1 (LC-1) was involved. 2. The i.d. injection of ovalbumin (OVA) in sensitized mice induced a dose-dependent recruitment of (111)In-labelled blood eosinophils which peaked at 4 to 8 h after antigen challenge. Systemic treatment with dexamethasone (50 microg per mouse, 3 h after antigen) effectively inhibited (111)In-eosinophil recruitment in this reaction by 70 to 85%. Similarly, a 1 h pretreatment with dexamethasone significantly suppressed (111)In-eosinophil induced by platelet-activating factor (PAF), leukotriene B4(LTB4) and the chemokine macrophage inflammatory protein-1alpha (MIP-1alpha) by 40 to 70%. 3. Two experimental approaches were used to evaluate the role of LC-1: treatment with LC-1 fragment Ac2-26 and use of an anti-LC-1 antiserum. LC-1 fragment Ac2-26 (100 microg per mouse) failed to affect (111)In-eosinophil recruitment. Moreover, pretreatment of animals with an anti-LC-1 antiserum failed to reverse the inhibitory effects of dexamethasone on (111)In-eosinophil recruitment induced by MIP-1alpha and by antigen in sensitized mice. 4. In contrast, the LC-1 fragment significantly inhibited glycogen-induced neutrophil recruitment into the peritoneal cavity of mice. Furthermore, the anti-LC-1 antiserum reversed the inhibitory effects of dexamethasone on the glycogen-induced neutrophil recruitment. 5. Thus, our results suggest that dexamethasone can inhibit the recruitment of eosinophils in mouse skin independent of an action on the bone marrow. However, by use of two different approaches, we showed that LC-1 does not play a role in mediating the inhibitory action of dexamethasone on eosinophil migration into cutaneous inflammatory reactions in the mouse. These data add further support to a LC-1-independent action of dexamethasone on eosinophils in vivo.

Animals↗
PubMed1998

Investigation of the functional role played by the chemokine monocyte chemoattractant protein-1 in interleukin-1-induced murine peritonitis.

1. Intraperitoneal (i.p.) injection of murine recombinant IL-1beta (mrIL-1beta) produced a dose-dependent (0.5-50 ng) and time-related (0.5-2 h) secretion of murine monocyte chemoattractant protein-1 (mMCP-1; 3-4 ng per cavity) in the lavage fluids. MCP-1 mRNA could also be detected in the cell pellets by reverse transcriptase-polymerase chain reaction (RT-PCR). 2. MCP-1 levels were reduced by more than 90% by co-administration of IL-1 receptor antagonist (10 microg) (n=6, P<0.05). In contrast, an IL-1 mutant with low affinity for IL-1 receptor type I, termed yIL-1betadelta4 (50 ng), produced only a modest release of the chemokine. Treatment of mice with dexamethasone (DEX) (approximately 1 mg kg(-1) s.c.) reduced mrIL-1beta-induced mMCP-1 gene expression (apparent total inhibition) and protein release in the lavage fluids (approximately 40% reduction; n=10; P<0.05). Drastic reductions in the numbers of residential macrophages or mast cells did not modify the levels of mMCP-1 recovered in the lavage fluids. 3. Injection of mrIL-1beta produced neutrophil accumulation into the peritoneal cavities (maximal at 4 h with 1.42+/-0.15 x 10(6) cells per mouse). Co-injection of a specific polyclonal antibody against mMCP-1 reduced this process by more than 50% (n=6; P<0.05). In conclusion, we studied the mechanisms leading to the specific release of the CC chemokine mMCP-1 after in vivo administration of mrIL-1beta.

Animals↗
PubMed1998

Regulation of mitochondrial ATP synthase activity in human myocardium.

1. In previous studies regulation of the F1F0-ATPase of mitochondrial complex V (ATP synthase) has been demonstrated in rat cardiomyocytes, canine mycocardium and skeletal muscle from children. The aim of the present study was to examine regulation of ATP synthase in human myocardium in response to different metabolic states. 2. Biopsy material was obtained from 10 children undergoing cardiac surgery. Mitochondria in the post-nuclear supernatant were incubated under different metabolic conditions for 15 min and then broken by sonication. ATP synthase was measured spectrophotometrically using a coupled enzyme assay. 3. ATP synthase can be rapidly measured in sonicated preparations of heart mitochondria from children. We show that direct regulation at the level of ATP synthase occurs in these mitochondria. ATP synthase capacity is decreased in response to blocking of the respiratory chain by cyanide (mimicking anoxia) or uncoupling of mitochondria, falling to 76% and 66% of control values respectively. Upregulation of ATP synthase can be demonstrated in heart mitochondria when the calcium concentration in the incubation medium is increased to 5 microM (130% of control). 4. ATP synthase is actively regulated in heart mitochondria from children. The enzyme is upregulated in response to increased calcium. This transition may reflect the increased energy demand when cardiac workload is increased.

Calcium↗
PubMed1997

Eotaxin-induced eosinophil migration in the peritoneal cavity of ovalbumin-sensitized mice: mechanism of action.

Cell accumulation in response to i.p. administration of the C-C chemokine, eotaxin, was studied in vivo. OVA-sensitized mice, exhibiting blood eosinophilia, had greater eosinophil (Eø) accumulation in response to 500 ng of eotaxin at 6 h (vehicle-injected, 3.0 +/- 0.5 x 10(5); eotaxin-injected, 8.6 +/- 1.0 x 10(5)) than nonsensitized, eotaxin-injected mice (2.5 +/- 0.4 x 10(5)). A nonspecific neutrophil migration was observed in both vehicle- and eotaxin-injected cavities. The number of intact mast cells in the peritoneal lavages after eotaxin injection was significantly lower than that in vehicle-injected animals (0.8 +/- 0.3 x 10(4) vs 2.8 +/-0.6 x 10(4), respectively). When endogenous peritoneal mast cells were depleted with compound 48/80 before eotaxin administration, there was a 51% reduction in Eø accumulation. This suggests an important role for endogenous mast cells in mediating the actions of eotaxin. The potential role of mast cell mediators in the actions of eotaxin was also investigated. Pretreatment with histamine-H1 or serotonin antagonists reduced Eø migration in response to eotaxin by 50 to 65%. Further, following pretreatment with a specific mAb against TNF-alpha, only nonspecific neutrophil influx was attenuated. Using neutralizing mAbs, Eø migration was found to be dependent on the adhesion molecules P- and E-selectin and CD11b. Eø accumulation was also sensitive to dexamethasone, with doses as low as 0.2 mg/kg inducing 100% inhibition. This study provides useful insight into the mechanisms of action of eotaxin.

Animals↗
PubMed1997

Subacute treatment of rats with dexamethasone reduces ICAM-1 levels on circulating monocytes.

We report for the first time that in vivo treatment with dexamethasone (DEX) reduces levels of intercellular adhesion molecule-1 (ICAM-1) expression on rat circulating unstimulated monocytes (-55%) and peritoneal macrophages (-26%). This effect was present following sub-acute (5 days) treatment with a low dose (0.1 mg/kg per day), but not after single administration of a high dose (1 mg/kg, -2 h), of the steroid. Both acute and sub-acute treatment with DEX failed to modify either basal or up-regulated CD11b expression on peripheral blood monocytes and neutrophils, elastase release from neutrophils, and beta-glucuronidase release from cultured macrophages. The lack of alteration of CD11b expression on circulating leukocytes suggests that the effect of DEX on ICAM-1 expression is secondary to gene repression rather than a non-specific blockade of cell differentiation. These data promote the concept that different dose-regimens with glucocorticoids affect distinct molecular targets and indicate that clinically-related protocols of DEX may reveal new mechanism(s) of action.

Animals↗
PubMed1997

A novel murine model of allergic inflammation to study the effect of dexamethasone on eosinophil recruitment.

1. We have developed a novel model of allergen-induced eosinophil into mouse air-pouches following sensitization and challenge with ovalbumin (Ova). This model was used to investigate the mechanism(s) underlying the anti-inflammatory action of the glucocorticoid hormone dexamethasone (Dex). 2. Injection of 10 micrograms Ova into 6-day-old dorsal air-pouches of mice sensitized to the same antigen provoked an intense cell accumulation as early as 6 h post-challenge (0.08 +/- 0.03 and 4.0 +/- 1.0 x 10(5) leucocytes in saline and Ova-treated air-pouches, respectively), maximal at 24 h (0.02 +/- 0.01 and 6.0 +/- 0.8 x 10(5) leucocytes in saline and Ova-treated air-pouches, respectively) and persisted up to 48 h. At the 24 h time-point, the cellular infiltrate consisted of 37% eosinophils, 18% neutrophils and 45% mononuclear cells, as assessed by histological examination. The same ratio of eosinophil/neutrophil was obtained by fluorescence-activated cell sorting (FACS) analysis, since 72% of the polymorphonuclear (PMN) population was positive for very-late antigen-4 (VLA-4) expression. 3. Subcutaneous (s.c.) administration of Dex (50 or 100 micrograms per mouse, -1 h) inhibited eosinophil accumulation into Ova challenged air-pouches by about 70% (P < 0.05) and 75% (P < 0.05), respectively, when compared to controls. Cell accumulation measured at 48 h after Ova injection was also significantly reduced (-75%) by Dex administration at the 24 h time-point (n = 12, P < 0.05). 4. Eosinophil numbers in the bone marrow and blood were quantitated. We found that the sensitization protocol induced a 3 fold increase in eosinophil numbers in the bone marrow (naive mice: 2.7 +/- 0.3 x 10(5); sensitized mice: 8.7 +/- 1.7 x 10(5), P < 0.05) and blood (naive mice: 0.5 +/- 0.2 x 10(5); sensitized mice: 1.5 +/- 0.3 x 10(5), P < 0.01). However, 24 h following Ova challenge, the eosinophil numbers in the bone marrow had dropped (3.7 +/- 0.8 x 10(5) with no change in the circulating pool, suggesting an equilibrium within the eosinophil pools had been reached. 5. Dex administration provoked a profound eosinopaenia in the blood of naive (5.2 +/- 1.5 to 0.9 +/- 0.6 x 10(4)) and sensitized mice (1.5 +/- 0.3 to 0.08 +/- 0.02 x 10(5)) at 4 h. This effect was reversed within 24 h. Dex also inhibited the release of eosinophils from the bone marrow in response to Ova challenge. 6. We show for the first time that express the steroid-inducible protein lipocortin 1 (LC1). FACS analysis of eosinophils emigrated into the Ova challenged air-pouches revealed detectable LC1-like immunoreactivity (373 x 10(3)). These data were also substantiated by LC1 detection in circulating eosinophils of interleukin-5 transgenic mice (strain: CBA/Ca). However, s.c. injection of Dex (50 micrograms) did not alter LC1 levels in blood eosinophils, such that 235 +/- 21 x 10(3) LC1-like molecules per cell were measured after vehicle treatment (n = 5), and 224 +/- 8 x 10(3) LC1-like molecules per cell were associated with this cell type 1 h after steroid treatment (n = 5, not significant). Finally, resident eosinophils (in the pleural cavity) were found to have much higher LC1 levels than that found in the blood circulation (2 fold increase, P < 0.05). 7. Passive immunization of mice against LC1 with a validated antiserum (termed LCS3) and protocol failed to modify the anti-migratory activity exerted by Dex towards eosinophil extravasation into Ova-challenged air-pouches. The steroid (50 micrograms s.c., -1 h) produced a similar degree of inhibition of eosinophil accumulation both in control animals (treated with a non-immune sheep serum) the LCS3-treated mice (-56% and 59%, respectively, n = 15-21, not significant). 8. In conclusion, the air-pouch provides a novel and convenient cavity to study allergen-induced cell recruitment which is sensitive to glucocorticoid hormone treatment. The effect of Dex on eosinophil distribution in these experimental conditions has been studied in detail and

Animals↗
PubMed1995

Lung eosinophilia is dependent on IL-5 and the adhesion molecules CD18 and VLA-4, in a guinea-pig model.

Blood and tissue-eosinophilia is a characteristic feature of a number of disease states. In experimental animals, the intravenous injection of parasitic larvae induces a profound eosinophilia that can be mimicked by the intravenous injection of Sephadex particles. In the present study, this procedure was used to investigate the mechanisms involved in the development of lung eosinophilia in a guinea-pig model. Intravenous administration of Sephadex particles to guinea-pigs resulted in a significant increase in the influx of eosinophils in the airways and in lung tissue eosinophil peroxidase (EPO) activity (at t = 24 hr). An anti-interleukin-5 (IL-5) monoclonal antibody (mAb) totally inhibited the eosinophilia in the airways and significantly reduced the lung tissue EPO activity. The concomitant accumulation of neutrophils and mononuclear cells, however, was not affected by this treatment. Monoclonal antibodies to VLA-4 and CD18 caused 58% and 62% suppression of eosinophilia in the bronchoalveolar lavage (BAL), respectively, whilst having no effect on lung tissue EPO activity. Co-administration of the two mAb resulted in total inhibition of eosinophil accumulation into BAL and significant suppression of lung tissue EPO activity (55% inhibition). This procedure also resulted in 72% inhibition of mononuclear cell influx and 68% inhibition of neutrophil influx in the BAL, the latter effect being entirely due to the actions of the anti-CD18 mAb. The results of this study indicate for the first time a requirement for IL-5 in the development of lung eosinophilia in this model. Further, it is clear that both the molecules VLA-4 and CD18 contribute to the development of this response and that maximal inhibition of lung eosinophilia is achieved only when the two adhesion pathways are simultaneously blocked.

Animals↗
PubMed1994

Absence of cytochrome c oxidase activity in a boy with dysfunction of renal tubules, brain and muscle.

We report on a boy who developed proximal renal tubular acidosis with loss of carnitine at the age of about 6 months. A few months later he began to suffer from progressive muscular weakness and neurological disturbances. Blood biochemistry showed elevated lactate and beta-hydroxybutyrate with increased lactate/pyruvate and beta-hydroxybutyrate/acetoacetate ratios. A high urinary excretion of lactate and citric acid cycle intermediates was found. These results indicated a defect of the mitochondrial respiratory chain. Analysis of biopsy material from skeletal muscle revealed low activities of all respiratory chain complexes. In muscle and fibroblasts cytochrome c-oxidase (complex IV) was absent. Despite high dose multi-vitamin therapy the boy died at the age of 30 months from central respiratory failure. At autopsy the neuropathological diagnosis of Leigh disease was made.

Acidosis, Renal Tubular↗
PubMed1994

Regulation of the mitochondrial ATP-synthase in human fibroblasts.

The F1F0-ATPase activity of mitochondrial complex V can be rapidly measured in sonicated preparations of monolayer skin fibroblast cultures from children. We show that direct regulation at the level of ATP-synthase occurs in these cell preparations. ATP-synthase capacity is decreased in response to blocking of the respiratory chain by cyanide (mimicking anoxia) or uncoupling of mitochondria. ATP-synthase capacity falls to 60% and 35% of control, respectively. Up-regulation of ATP-synthase can be demonstrated in fibroblasts exposed to 4 mmol/l calcium (127% of control). Mitochondrial recovery was unchanged under the different incubation conditions as judged by the activity of mitochondrial marker enzymes. We conclude that direct regulation at the level of ATP-synthase occurs in vivo in human fibroblasts. The naturally occurring inhibitor protein IF1 and the calcium binding inhibitor protein CaBI may be involved in this regulation of ATP-synthase.

Adolescent↗
PubMed1993

Field tests for rational drug use in twelve developing countries.

Increasing efforts are being made to improve drug-use practices and prescribing behaviour in developing countries. An essential tool for such work is an objective and standard method of assessment. We present here a set of drug-use indicators produced and tested in twelve developing countries. We describe practical applications, which include the use of indicators to increase awareness among prescribers in Malawi and Bangladesh, to identify priorities for action (eg, polypharmacy in Indonesia and Nigeria, overuse of injections in Uganda, Sudan, and Nigeria, and low percentage of patients who understood the dosage schedule in Malawi), and to quantify the impact of interventions in Yemen, Uganda, Sudan, and Zimbabwe.

Developing Countries↗
PubMed1993

Regulation of the mitochondrial ATP synthase is defective in rat heart during alcohol-induced cardiomyopathy.

Mitochondrial ATP synthase capacity was measured in cardiomyocytes from sucrose-fed (control) and alcohol-fed (cardiomyopathic) rats. Cells from alcohol-fed rats showed an ATP synthase capacity raised by 40% relative to the control values of 1.8 mumol/min per mg cell protein. Cells from control rats were able to increase their ATP synthase capacity by up to 80% in response to increased energy demand. In contrast, cells from alcohol-fed rats had lost the ability to up-regulate their ATP synthase. Cells from control rats maintained their internal ATP levels at 38 nmol/mg cell protein before and after an increased energy demand. In cells from alcohol-fed rats, ATP levels fell by 20% after 2 min of increased energy demand. It was concluded that the inability of cells from alcohol-fed rats to maintain ATP levels was due to their reduced ability to increase mitochondrial ATP synthase activity as energy demand is increased. Such ATP deficits may contribute to heart dysfunction in alcohol-induced cardiomyopathy.

ATP Synthetase Complexes↗
PubMed1992

Mitochondrial ATP synthase regulation in heart: defects in hypertension are restored after treatment with captopril.

Control of mitochondrial ATP synthase capacity was investigated in cultured cardiomyocytes from normotensive (Wistar-Kyoto) and spontaneously hypertensive rats. Cells from spontaneously hypertensive rats have a higher basal ATP synthase capacity than those from normotensives, but lack the normal up-regulation in response to an increased energy demand. After treatment of spontaneously hypertensive rats with captopril (60 mg/kg per day for 12 weeks), cellular hypertrophy characteristic of the hypertensives was abolished and the cardiomyocytes showed a normal ATP synthase capacity. Normal up-regulation of this enzyme was also restored. All cells showed a normal down-regulation of the synthase in response to cyanide. Experiments with the calcium antagonists, verapamil and ruthenium red, suggest that abnormal ATP synthase regulation observed in the untreated spontaneously hypertensive rats results from an alteration of Ca2+ handling in cardiac cells under chronic high workload, which is reversed by captopril treatment.

Animals↗