Biomedical subjects

R J Flower

Publications and source records attributed to R J Flower.

At least 55 records · Page 3Linked to original sources

PubMed1999

Dexamethasone inhibition of leucocyte adhesion to rat mesenteric postcapillary venules: role of intercellular adhesion molecule 1 and KC.

BACKGROUND: A previous study showed that the glucocorticoid dexamethasone, at doses of 100 microg/kg and above, inhibited leucocyte adhesion to rat mesenteric postcapillary venules activated with interleukin 1beta (IL-1beta), as assessed by videomicroscopy. AIMS: To identify whether the adhesion molecule, intercellular adhesion molecule 1 (ICAM-1), or the chemokine KC could be targeted by the steroid to mediate its antiadhesive effect. METHODS: Rat mesenteries were treated with IL-1beta (20 ng intraperitoneally) and the extent of leucocyte adhesion measured at two and four hours using intravital microscopy. Rats were treated with dexamethasone, and passively immunised against ICAM-1 or KC. Endogenous expression of these two mediators was validated by immunohistochemistry, ELISA, and the injection of specific radiolabelled antibodies. RESULTS: Dexamethasone greatly reduced IL-1beta induced leucocyte adhesion, endothelial expression of ICAM-1 in the postcapillary venule, and release of the mast cell derived chemokine KC. Injection of specific antibodies to the latter mediators was also extremely effective in downregulating (>80%) IL-1beta induced leucocyte adhesion. CONCLUSIONS: Induction by IL-1beta of endogenous ICAM-1 and KC contributes to leucocyte adhesion to inflamed mesenteric vessels. Without excluding other possible mediators, these data clearly show that dexamethasone interferes with ICAM-1 expression and KC release from mast cells, resulting in suppression of leucocyte accumulation in the bowel wall, which is a prominent feature of several gastrointestinal pathologies.

Animals↗

PubMed1999

Endotoxin induction of nitric oxide synthase and cyclooxygenase-2 in equine alveolar macrophages.

OBJECTIVE: To determine the amount of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) enzymes induced in vitro in equine alveolar macrophages in response to lipopolysaccharide (LPS). Sample Population-Alveolar macrophages obtained from 12 horses. PROCEDURE: Alveolar macrophages were collected by bronchoalveolar lavage from 12 horses and incubated for 6 hours with LPS (0.001 to 10 microg/ml) or vehicle. Total RNA was extracted and purified. After first-strand cDNA synthesis, mRNA induction was measured, using a polymerase chain reaction (PCR) technique for COX-2, iNOS, and glyceraldehyde 3-phosphate dehydrogenase. In a second study, cells were incubated with LPS or vehicle for 24 hours. Culture medium was assayed for COX-2 and iNOS activity by determining prostaglandin E2 (PGE2) and total nitrite concentrations, respectively. RESULTS: Lipopolysaccharide induces COX-2 and iNOS mRNA in equine alveolar macrophages. Sequencing revealed that PCR products for COX-2 and iNOS had a high degree of nucleotide homology with the human sequences (91% COX-2, 93% iNOS). Production of mRNA for COX-2 and iNOS was accompanied by induction of enzyme activity. Comparing PCR fragment production, expression of mRNA for iNOS appeared to be less than that for COX-2. Induction of COX-2, but not iNOS, was LPS-concentration dependent. Conclusion-Lipopolysaccharide induces COX-2 and iNOS in equine macrophages. CLINICAL RELEVANCE: The induction of iNOS and COX-2 by LPS in equine macrophages suggests these enzymes may be important in the pathophysiology of sepsis. Pharmacologic modulation of iNOS and COX-2 activity may represent a novel therapeutic target in the management of endotoxemia in horses.

Animals↗

PubMed1998

Promoting detachment of neutrophils adherent to murine postcapillary venules to control inflammation: effect of lipocortin 1.

In this study we investigated, using intravital microscopy, how neutrophil extravasation across mouse mesenteric postcapillary venules is inhibited by the glucocorticoid-regulated protein lipocortin (LC; also termed annexin) 1. Intraperitoneal injection of 1 mg of zymosan into mice induced neutrophil rolling on the activated mesenteric endothelium followed by adhesion (maximal at 2 hr: 5-6 cells per 100-micrometers of vessel length) and emigration (maximal at 4 hr: 8-10 cells per high-powered field). Treatment of mice with human recombinant LC1 (2 mg/kg s.c.) or its mimetic peptide Ac2-26 (13 mg/kg s.c.) did not modify cell rolling but markedly reduced (>/=50%) the degree of neutrophil adhesion and emigration (P < 0.05). Intravenous treatment with peptide Ac2-26 (13 mg/kg) or recombinant human LC1 (0.7-2 mg/kg) promoted detachment of neutrophils adherent to the endothelium 2 hr after zymosan administration, with adherent cells detaching within 4.12 +/- 0.75 min and 2.36 +/- 0.31 min, respectively (n = 20-25 cells). Recruitment of newly adherent cells to the endothelium was unaffected. The structurally related protein LC5 was inactive in this assay, whereas a chimeric molecule constructed from the N terminus of LC1 (49 aa) attached to the core region of LC5 produced cell detachment with kinetics similar to LC1. Removal of adherent neutrophils from activated postcapillary endothelium is a novel pharmacological action, and it is at this site where LC1 and its mimetics operate to down-regulate this aspect of the host inflammatory response.

Amino Acid Sequence↗

PubMed1998

Blockade of the classical pathway of protein secretion does not affect the cellular exportation of lipocortin 1.

The mechanism by which lipocortin 1 (LC1) is extruded from cells in the brain and periphery in response to a glucocorticoid challenge is unknown. This study examined the influence of three inhibitors of the classical endoplasmic reticulum-Golgi pathway of protein secretion on the dexamethasone-induced (0.1 microM, 2-3 h) cellular exportation of LC1 in vitro in brain (cortex, hippocampus, hypothalamus), anterior pituitary tissue and peritoneal macrophages. In all instances, the steroid-induced exportation of LC1 was unaffected by brefeldin A (1.4 microM), monensin (10 microM) and nocodazole (3.3 microM); however, these drugs readily blocked the release of corticotrophin from pituitary tissue. These data suggest that LC1 is exported by a mechanism distinct from the classical pathway of protein secretion.

Animals↗

PubMed1998

Suppression by dexamethasone of inducible nitric oxide synthase protein expression in vivo: a possible role for lipocortin 1.

Western blot and densitometric analysis of organ homogenates from lipopolysaccharide (LPS)-treated rats (1-10 mg kg(-1), i.p.) exhibited a strong induction of inducible nitric oxide synthase (iNOS) expression seen at all the doses tested (1, 3, and 10 mg kg(-1), n = 3). In particular, 3 hr after challenge of rats with LPS, iNOS was detectable in the liver, kidney, aorta, spleen and lung. Dexamethasone (DEX) (0.1-1 mg kg(-1); -1 hr) dose-dependently reduced iNOS expression in lung homogenates after exposure to LPS (1 mg kg(-1); P < 0.05). A partial reversal of DEX-induced suppression of iNOS expression in lung homogenates 3 hr after challenge with LPS was observed in rats which received a specific anti-lipocortin 1 sheep serum (LCS3; 1 mL kg(-1) 24 hr prior to the steroid), with an inhibition of 35+/-8%, as compared to animals passively immunised with normal sheep serum where dexamethasone exhibited an inhibition of 60+/-7% (n = 4). Peritoneal macrophages collected from rats treated with LPS (1 mg kg(-1); 3 hr) and cultured for 16 hr, released significant amounts of nitrite (51+/-1 microM) into the cell supernatants; this was reduced (-70+/-6%) after pre-treatment with dexamethasone (0.3 mg kg(-1)) and this effect was neutralised if animals were passively immunised with LCS3 (P < 0.01; n = 4). Thus lipocortin 1 mediates, at least in part, the inhibitory action exerted by dexamethasone on both iNOS protein expression in lung and iNOS activity (as measured by nitrite release) in primary peritoneal cells of rats.

Animals↗

PubMed1998

Endogenous monocyte chemoattractant protein-1 recruits monocytes in the zymosan peritonitis model.

The role of monocyte chemoattractant protein-1 (MCP-1) in the recruitment of blood-derived monocytes in a model of zymosan peritoneal inflammation was investigated. After zymosan injection (1 mg) a rapid influx of polymorphonuclear leukocytes (PMN) and monocytes into the peritoneal cavity associated with mouse MCP-1 (JE) gene activation and protein secretion in the exudates occurred. MCP-1 production (maximal at 4 h) preceded the accumulation of monocytes (F4/80-positive cells, maximally recovered between 16 and 24 h). Treatment of mice with a single injection of anti-mouse MCP-1 antibody inhibited 16-h monocyte accumulation by approximately 40%, however, a significant decrease in the number of PMN was also measured. Finally, intraperitoneal injection of murine recombinant MCP-1 (1 microg) produced a selective accumulation of monocytes (F4/80-positive cells) into the peritoneal cavity. In conclusion, we show the novel existence of a strict relationship between MCP-1 production and leukocyte accumulation in this model of acute inflammation.

Acute Disease↗

PubMed1998

Resident mast cells are important for eotaxin-induced eosinophil accumulation in vivo.

Eotaxin administration intraperitoneally, but not into dorsal air-pouches, of ovalbumin-sensitized mice exhibiting blood eosinophilia induced a threefold increase in eosinophil (E phi s) infiltration. Transfer of 1 x 10(6) mixed peritoneal cavity cells (PCC), containing 3.5 to 4.5 x 10(4) mast cells (MC), from donor mice to air-pouches of sensitized (but not unsensitized) recipient mice, established an E phi infiltration to eotaxin (vehicle, 0.86 +/- 0.27 x 10(6); eotaxin, 1.63 +/- 0.16 x 10(6) E phi s/air-pouch). Neutrophil numbers were also increased. When MC-depleted (-93%) PCC were injected into air-pouches of recipient animals, E phi infiltration was not supported (-52%). Injection of macrophage-depleted (-99%) PCC into air-pouches elicited a full E phi response to eotaxin but not neutrophil infiltration (-81%). Systemic dexamethasone treatment of recipient mice reduced E phi accumulation; treatment of donor mice only reduced neutrophil accumulation. Our study points to a crucial role for MC in E phi recruitment by eotaxin.

Animals↗

PubMed1998

IL-6 stimulates annexin 1 expression and translocation and suggests a new biological role as class II acute phase protein.

Annexin 1 (Ax 1), a protein whose synthesis and secretion are induced during the inflammatory response, has been proposed as a mediator of the anti-inflammatory action of glucocorticoids. To gain insight into a broader role of Ax 1 during the inflammatory response, the authors have investigated how pro-inflammatory cytokines [interleukin 1 (IL-1), IL-6 and tumour necrosis factor alpha (TNF-alpha)] affect Ax 1 expression and regulation at transcriptional and translational levels. The authors show that induction of the Ax 1 protein and its translocation to the cell membrane are stimulated by interleukin 6. However neither IL-1 nor TNF-alpha display these effects. Analysis of 5'-deletion mutants and the full length Ax 1 promoter fused to a luciferase reporter gene using transient transfections of human lung adenocarcinoma A 549 cells identified a unique 30 bp region of the Ax 1 promoter as critical for the responsiveness of the reporter gene to IL-6 and dexamethasone. Gel retardation and supershift assays showed that IL-6 stimulation is mediated by a C/EBP beta-like transcriptional factor. These data suggest that Ax 1 may participate in host defence as a new acute class II phase protein.

Acute-Phase Proteins↗

PubMed1998

Inhibitory effect of peptides derived from the N-terminus of lipocortin 1 on arachidonic acid release and proliferation in the A549 cell line: identification of E-Q-E-Y-V as a crucial component.

1. The ability of the glucocorticoid-induced protein lipocortin 1 (LC1) to inhibit arachidonic acid release and cell proliferation in A549 cells may be mimicked by a sequence taken from the N-terminal, LC1(13-25) (FIENEEQEYVQTV). We have now synthesized and tested for biological activity a library of 25 smaller peptides derived from this sequence. 2. Peptides were tested in two assays: A549 cells were prelabelled with tritiated arachidonic acid and thapsigargin (50 nM) and EGF (10 nM) used to stimulate the release of this fatty acid. Cell proliferation was determined by counting cell numbers following 3 day incubation with these peptides, or controls. 3. Many of the peptides were highly insoluble but could be more readily dissolved in aqueous solution in the presence of commercial liposomes or phosphatidyl serine (5 microM). Since neither of these agents alone had any effect on arachidonic acid release or cell proliferation, all peptides were tested in the presence of 5 microM phosphatidyl serine. Under these conditions LC1(13-25) was active in both assay systems with an IC40 of 40.7 and 57.0 microM respectively. 4. Deletion of amino acids from the C-terminus of the peptide progressively diminished (2-3 fold) the molar potency of LC1(13-25) in both assays: after the removal of Val22 biological activity was virtually undetectable or very weak (< 30% of LC1[13-25]). 5. Removal of amino acids from the N-terminus also lead to a progressive reduction (3-5 fold) in the molar potency of the peptides and biological activity became undetectable, or very weak, after the removal of Glu18. 6. All active peptides contained the core sequence EQEYV(Glu-Gln-Glu-Tyr-Val) which seems to represent a crucial component of the pharmacophore, although this sequence on its own was inactive and the shortest peptide with significant activity was LC1(18-25) (EQEYVQTV). 7. Methoxylation of Tyr21 abolished the ability of LC1(18-25) to inhibit cell proliferation and arachidonic acid release. A cyclized version of LC1(18-25) was also tested and found to be inactive. 8. LC1(18-25) (178 microM) inhibits cPLA2 activation in A549 cells as judged by a band-shift assay, whereas equimolar concentrations of an inactive peptide LC1(19-25) were without effect in this assay system. 9. Several possible mechanisms whereby these peptides act are discussed in the light of LC1 biology and of the effect of glucocorticoids on cell function.

Amino Acid Sequence↗

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

Role of inducible nitric oxide synthase in the regulation of neutrophil migration in zymosan-induced inflammation.

In the present study, by comparing the responses in wild-type mice and mice lacking the inducible (or type 2) nitric oxide synthase (iNOS), we investigated the role played by iNOS in the regulation of polymorphonuclear granulocyte (PMN) accumulation and chemokine production in the mouse peritoneal cavity in response to administration of zymosan (0.2 mg). Zymosan injection induced the production of nitric oxide, and triggered a time-dependent PMN immigration into the peritoneal cavity. This response was associated with increases in the level of the chemokines macrophage inflammatory protein (MIP)-1alpha, MIP-2, monocyte chemo-attractant protein (MCP)-1 and cytokine-induced neutrophil chemo-attractant (KC), as measured in the peritoneal cavities. Injection of zymosan also induced a time-dependent increase in the production of the anti-inflammatory cytokine interleukin-10 (IL-10) in the peritoneal cavity. When comparing the response between wild-type and iNOS knockout (KO) mice, we observed that the low-level PMN accumulation measured at 1 hr was slightly but significantly increased in the absence of functional iNOS. On the other hand, the delayed response (2-4 hr after zymosan) of PMN accumulation was suppressed in the iNOS KO mice. The early enhancement of PMN infiltration in the iNOS-deficient mice was associated with increased peritoneal levels of MIP-2, KC and IL-10 proteins. The delayed suppression of PMN infiltration was associated with reduced MIP-2 and IL-10 levels in the peritoneal cavity. The lack of iNOS did not affect the release of MIP-1alpha and MCP-1 at any of the time-points studied. The current data demonstrate that iNOS regulates the production of certain CXC (but not CC) proinflammatory chemokines, the production of IL-10 and exerts a biphasic regulatory effect on PMN accumulation in zymosan-induced acute inflammation.

Animals↗

PubMed1998

Lipocortin 1 (annexin 1) in patches associated with the membrane of a lung adenocarcinoma cell line and in the cell cytoplasm.

Lipocortin 1 (annexin I) is a calcium- and phospholipid-binding annexin protein which can be externalised from cells despite the lack of a signal sequence. To determine its cellular distribution lipocortin 1 in A549 human lung adenocarcinoma cells was localised by light- and electron-microscopic immunocytochemistry and by cell fractionation and western blotting. Lipocortin 1 immunoreactivity is concentrated in prominent patches associated with the plasma membrane. The intensity of these patches varied with the confluence and duration of the culture and was not detectably diminished by an EDTA wash before fixation. Tubulin and cytokeratin 8 were colocalized with lipocortin 1 in the patches. Within the cells lipocortin 1 was distributed throughout the cytoplasm. Electron microscopy revealed prominent immunoreactivity along the plasma membrane with occasional large clusters of gold particles in contact with the membrane surface of the cells; within the cytoplasm the membrane of some vesicle/vacuole structures and some small electron-dense bodies was immunoreactive, but no immunogold particles were associated with the multilamellar bodies. Subcellular fractionation, extraction and western blotting showed that lipocortin 1 in the membrane pellet was present as two distinct fractions; one, intimately associated with the lipid bilayer, which behaved like an integral membrane protein and one loosely attached which behaved like a peripheral membrane protein. The results show that a substantial amounts of lipocortin 1 is concentrated in focal structures associated with and immediately beneath the plasma membrane. These might form part of the mechanism by which lipocortin 1 is released from the cells.

Adenocarcinoma↗

PubMed1998

Human annexin 1 is highly expressed during the differentiation of the epithelial cell line A 549: involvement of nuclear factor interleukin 6 in phorbol ester induction of annexin 1.

The role of annexin 1 (Ax 1) in cell differentiation was studied in the A 549 epithelial cell line, a human lung adenocarcinoma line, that responds to phorbol esters and glucocorticoids by induction of differentiated properties. Ax 1 has also been reported to be involved in the control of cell proliferation. We report that Ax 1 synthesis occurs upon phorbol 12-myristate 13-acetate (PMA) treatment of A 549 cells and its appearance is correlated with the presence of dipeptidyl peptidase IV, or CD26, a marker of epithelial cell differentiation. In addition, using transfection experiments and site-directed mutagenesis with the Ax 1 promoter coupled to a reporter gene, we report that a unique region of the Ax 1 promoter confers the response of the reporter gene to PMA and dexamethasone. This response to PMA and/or dexamethasone involves the induction of the synthesis and/or the activity of trans/cis-activating transcriptional factors. Furthermore, we have delineated the mechanism of the transcriptional activation of Ax 1 by PMA and the involvement of a specific transcription factor, nuclear factor interleukin 6 (C/EBP beta).

Adenocarcinoma↗

PubMed1997

Tamoxifen inhibits the release of arachidonic acid stimulated by thapsigargin in estrogen receptor-negative A549 cells.

In pre-labelled A549 cells the tumour promoter thapsigargin (50 nM) stimulates the release of [5,6,8,9,11,12,14,15-3H(N)]-arachidonic acid (3H-AA) by ca. 300% above basal levels. A549 cells are estrogen receptor negative (ER-), yet this stimulation by thapsigargin is inhibited in a dose-dependent manner by a 3 h pre-treatment with the anti-estrogen tamoxifen (1-20 microM). Moreover, the presence of excess (100 microM) estradiol does not reverse this effect of tamoxifen. Thapsigargin stimulated 3H-AA release is not inhibited over the same concentration range by 4 hydroxy-tamoxifen nor by the steroidal anti-estrogen ICI 164384. However, the steroidal anti-estrogen ICI 182780 inhibits thapsigargin stimulated 3H-AA release in a similar manner to tamoxifen and this effect is also not reversed by the presence of excess estradiol. Stimulation of 3H-AA release by EGF (10 nM), IL-1beta (1 ng ml-1) and bradykinin (100 nM) was unaffected by these concentrations of tamoxifen. Ionomycin (10 microM) stimulates 3H-AA release by ca. 700% and A23187 (10 microM) by ca. 300% above basal levels. Pre-treatment with tamoxifen (1-20 microM) inhibits 3H-AA release stimulated by both these agents and again the presence of excess estradiol does not reverse this effect. Unlike the effects of glucocorticoids on 3H-AA release in A549 cells the effects of tamoxifen are not reversed by neutralizing anti-bodies to lipocortin 1. Arachidonic acid release is central to cell proliferation in A549 cells and we propose that this action of tamoxifen could explain the anti-proliferative effect seen in these cells and could have important implications for control of cell proliferation of ER- cells in general.

Annexin A1↗

PubMed1997

Lipocortin 1 protects against splanchnic artery occlusion and reperfusion injury by affecting neutrophil migration.

Splanchnic artery occlusion and reperfusion (SAO/R) shock was induced in rats by clamping both the superior mesenteric artery and the celiac trunk for 45 min, followed by release of the clamp (60-min reperfusion). Following this reperfusion period, rats developed a fall in mean arterial blood pressure, associated with a significant increase in tissue myeloperoxidase (MPO) activity in the intestine and a marked histologic injury to the distal ileum. Treatment of rats with a lipocortin-1 (LC1)-derived N-terminal peptide, peptide Ac(2-26), dose-dependently (0.125-0.5 mg/kg s.c.) reduced the progressive fall in blood pressure and prevented the infiltration of neutrophils into the reperfused intestine (reduced MPO activity). The LC1 peptide also reduced the degree of ischemia/reperfusion injury in the bowel as evaluated by histologic examination. The glucocorticoid dexamethasone (0.1 mg/kg s.c., -1 h) also produced a marked improvement in SAO/R shock (i.e., maintained mean arterial blood pressure and reduced tissue MPO activity), and this was reversed by pretreatment with two different antisera raised against the LC1 pharmacophore. Peptide Ac(2-26) (0.5 mg/kg s.c., -30 min) reduced (>60%) the extent of IL-1beta-induced cell emigration and significantly attenuated (approximately 45%) the number of adherent leukocytes in the rat mesenteric vascular bed, as assessed by video microscopy. These results suggest that LC1 inhibits neutrophil migration and accumulation into reperfused tissues, thereby ameliorating the outcome of SAO/R shock.

Amino Acid Sequence↗

PubMed1997

Inhibition of poly (ADP-ribose) synthetase attenuates neutrophil recruitment and exerts antiinflammatory effects.

A cytotoxic cycle triggered by DNA single-strand breakage and poly (ADP-ribose) synthetase activation has been shown to contribute to the cellular injury during various forms of oxidant stress in vitro. The aim of this study was to investigate the role of poly (ADP-ribose) synthetase (PARS) in the process of neutrophil recruitment and in development of local and systemic inflammation. In pharmacological studies, PARS was inhibited by 3-aminobenzamide (10-20 mg/kg) in rats and mice. In other sets of studies, inflammatory responses in PARS-/- mice were compared with the responses in corresponding wild-type controls. Inhibition of PARS reduced neutrophil recruitment and reduced the extent of edema in zymosan- and carrageenan-triggered models of local inflammation. Moreover, inhibition of PARS prevented neutrophil recruitment, and reduced organ injury in rodent models of inflammation and multiple organ failure elicited by intraperitoneal injection of zymosan. Inhibition of PARS also reduced the extent of neutrophil emigration across murine mesenteric postcapillary venules. This reduction was due to an increased rate of adherent neutrophil detachment from the endothelium, promoting their reentry into the circulation. Taken together, our results demonstrate that PARS inhibition reduces local and systemic inflammation. Part of the antiinflammatory effects of PARS inhibition is due to reduced neutrophil recruitment, which may be related to maintained endothelial integrity.

Animals↗

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

Glucocorticoids modulate the cellular disposition of lipocortin 1 in the rat brain in vivo and in vitro.

The role of glucocorticoids in the regulation of lipocortin 1 (LC1) mRNA/protein expression in the brain is uncertain. This study has examined the influence of dexamethasone on the disposition of LC1 protein in various central and peripheral tissues in the rat. LC1 was readily detectable in all tissues studied by Western blot analysis. Exposure to dexamethasone in vitro (0.1 microM, 3 h) or in vivo (20 micrograms/100 g i.p., 24 h before autopsy) had no discernible effects on intracellular LC1 levels but increased the amount of LC1 attached to the outer surface of cells (pericellular LC1) in cortex, hippocampus, hypothalamus, pituitary gland and peritoneal macrophages (in vitro only). The results suggest that in central tissues, as in the periphery, glucocorticoids promote the translocation of LC1 from intracellular to pericellular sites.

Animals↗