Peripheral blood-derived stem cells can be successfully cryopreserved without using controlled-rate freezing.
Explore the source record for details and available documents.
Blood Cells↗
Biomedical subjects
M Damon
Publications and source records attributed to M Damon.
Effects of a single dose of methotrexate on 5- and 12-lipoxygenase products in patients with rheumatoid arthritis.
The inhibition of 5-lipoxygenase could be involved in the mechanism of action of methotrexate (MTX). We studied 8 patients with active rheumatoid arthritis (RA) immediately before and the day after the first dose of MTX (10 mg intramuscularly). Leukotriene B4 (LTB4) formation by polymorphonuclear leukocytes was significantly decreased (32%, p less than 0.01). This essentially involved released LTB4. A slight decrease was also obtained in omega-oxidation products. Similar results were obtained for plasma LTB4 (30%, p less than 0.02). A non-significant decrease in 5-HETE was noted. Conversely, 12-HETE was not modified. Our results suggest MTX has an effect on the 5-lipoxygenase pathway, particularly at the LTA4 epoxide hydrolase step, since 5-HETE and 6-trans-LTB4 isomers are not involved.
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
Formation of lipoxins and leukotrienes by human alveolar macrophages incubated with 15(S)-HETE: a model for cellular cooperation between macrophages and airway epithelial cells.
Human alveolar macrophages (AM) from bronchoalveolar lavage of asthmatic patients (AP) and healthy volunteers (HS) were compared for their respective capacities to produce lipoxins and leukotrienes when stimulated by calcium ionophore A23187 with or without 15(S)-HETE. The metabolites were analyzed using an isocratic RP-HPLC system and their formation profiles evaluated on the basis of chromatographic behaviour, UV spectral characteristics and co-elution with synthetic standards. Without 15-HETE, AM from AP produced more LTB4 and 5-HETE than those from HS. In the presence of 15-HETE, human AM were able to produce 5,15-diHETE and lipoxins. Moreover, the total amount of lipoxins synthesized by AM from AP was 2 fold higher than that synthesized by AM from HS, thus showing an enhanced cell activation via the 5-lipoxygenase (5-LO) pathway. These results presented AM as in vitro 15-HETE metabolizing cells and suggested some hypothesis about human AM 5-LO regulation mechanism. The enhanced 5-LO activity in AM from AP suggested that in vivo they could participate in cell to cell interaction mechanisms involved in inflammatory lung diseases and might also take up and transform 15-HETE predominantly released by airway epithelial cells.
Adult↗
Airway macrophages releasability in bronchial asthma.
Asthma is a multifactorial disease on genetic basis. Its development is influenced by maternal and environmental factors, i.e. allergens and adjuvants. Early identification of candidates at high risk for development of asthma will enable giving recommendations on preventive measures focussing on exposure to tobacco smoke and other pollutants, indoor and outdoor allergens and possibly viral infections during infancy.
Asthma↗
[Effects of methotrexate on leukotriene and derivated lipoxygenase synthesis in polynuclear neutrophils in rheumatoid polyarthritis].
Methotrexate (MTX) has been proved to be effective in rheumatoid arthritis (RA). The mechanism of action of MTX in this disease remains unelucidated but may involve inhibition of the enzyme 5-lipoxygenase. Arachidonic acid metabolites were studied in eight patients with active RA immediately prior to and 24 hours after the first intramuscular injection of 10 mg MTX. None of the patients were taking corticosteroids. Nonsteroidal antiinflammatory drugs were withdrawn four days before the study. Reverse phase high-performance liquid chromatography was used to quantitate metabolites produced by 5-lipoxygenase (5-LO) and 12-LO in plasma (full spectrum of blood cells) and purified neutrophils (PN) after stimulation with calcium ionophore A 21387 (50 microM for 30 minutes and 5 microM for 5 minutes, respectively). LTB4 production by PNs was significantly decreased (-32%, p < 0.01) 24 hours after MTX administration. A moderate (-17%), nonsignificant (NS) fall in LTB4 omega-oxidation products (wP) was seen. Production of 5-HETE was also slightly decreased (-15%, NS). Findings in plasma were comparable, with a significant decrease in total LTB4 (-29.8%, p < 0.01) and moderate falls in wP (-18.8%, NS) and in 5-HETE production (-17%, NS). Production of 12-HETE was unchanged. These findings suggest that MTX in a single dose is responsible for a decrease in the synthesis of LTB4 and 5-LO products in neutrophils and other blood cells in RA patients but does not affect 12-LO activity.
Arthritis, Rheumatoid↗
Sulfidopeptide leukotrienes contribute to human alveolar macrophage activation in asthma.
The mechanism involved in amplification of the local inflammatory process, characteristic of asthma, was investigated through the role of human alveolar macrophages. During asthma attacks, mast cells and eosinophils are known to be activated in order to release arachidonic acid derived inflammation mediators such as sulfidopeptide leukotrienes. It is now known that these metabolites, particularly leukotriene C4, are present in bronchoalveolar lavage from asthmatic patients. Alveolar macrophages, recovered by bronchoalveolar lavage and purified by adherence, are able to transform LTC4 into LTE4. In four asthmatic patients with severe local inflammation as determined by fibrobronchoscopy, these phagocytes, incubated in the presence of LTC4, also generated LTB4 and 5-HETE, which remained within the cells. These preliminary results are discussed relative to amplification of the local process, involving cooperation between the different cells involved in airway responsiveness.
Adult↗
Releasability of airway macrophages in bronchial asthma.
Explore the source record for details and available documents.
Airway Obstruction↗
Neutrophil chemotactic activity of PAF, histamine and neuromediators in bronchial asthma.
Human blood polymorphonuclear neutrophils (PMN) are thought to be involved in the pathogenesis of asthma through their recruitment into the bronchoalveolar lumen and the lung by local release of chemotactic factors. Therefore chemotactic activities of several mediators (PAF, histamine and three neuropeptides substance P, VIP and a somatostatin analog) were compared on blood PMN from both healthy subjects (HS) and asthmatic patients (AP). The maximal response to PAF was significantly different (P less than 0.05) with cells from both groups. Moreover activity for the HS peaked at 10(-6) M, whereas the AP showed peak chemotactic activity at 10(-8) M. Histamine had no chemoattractant effect on PMN. Substance P did not induce PMN locomotion, whereas VIP induced a chemotactic response in a dose-dependent manner, particularly with cells from HS as compared to those from AP. BIM 23014 (a somatostatin analog) exhibited chemotactic activity which was also more pronounced with PMN from HS as compared to those from AP. Our findings showed that blood PMN could be involved in asthma through their heightened locomotor reactions to mediators which are known to be released locally by activated cells in bronchoalveolar lumen.
Adolescent↗
Reactive oxygen species released by human blood neutrophils piroxicam-treated healthy subjects.
Explore the source record for details and available documents.
Adult↗
Enhanced arachidonic acid metabolism and human neutrophil migration in asthma.
In stable state asthmatic patients (AP) without any airway obstruction, the capacity of peripheral blood polymorphonuclear neutrophils (PMN) to produce 5-lipoxygenase metabolites and to migrate, was investigated and compared with the response in healthy subjects (HS). After calcium-ionophore A23187 stimulation, PMN from AP and HS produced LTB4, its hydroxylated derivatives: omega-OH-and omega-CO2H-LTB4) (omega-LTB4, i.e 6-trans-LTB4 and 5,6-diHETE isomers, and 5-HETE. We found an increase in LTB4 (+59%), omega-LTB4 (+39%), 6-trans-LTB4 (+128%), and free 5-HETE (+63%) generation of AP as compared with HS. Unstimulated migration was enhanced in AP (122 +/- 27 PMN/10 high power fields (hpf) in AP versus 74 +/- 25 PMN/10 hpf in HS, p less than 0.025) and suggested a greater capacity of PMN from AP to migrate. This was confirmed by the PAF-induced chemotaxis studies which showed, in AP, a greater PAF-sensitivity of PMN (10(-6) M versus 10(-5) M in HS) and a greater chemotaxis response (600 +/- 50 PMN versus 200 +/- 35 PMN in HS). In AP, we compared the capacity of PMN to generate LTB4 and 5-HETE with their capacity to migrate. We found an inverse correlation (r = 0.86, p less than 0.007) of intracellular free 5-HETE with chemotaxis to PAF.
Adolescent↗
[Aberrant expression of HLA-DR antigens of the MHC class II in bronchial epithelial cells in asthmatic patients].
The study of cellular events in the bronchi of asthmatic patients shows significant epithelial destruction. The ciliated cells are more often destroyed than others in the respiratory epithelium. Using highly specific monoclonal antibodies, we detected that the epithelial cells were positive for HLA-DR antigen expression, whereas those from healthy subjects were negative. The aberrant expression could be explained by the presence of T lymphocytes in the asthmatic bronchial mucosa. The activated lymphocytes (CD4+) are known to release gamma interferon, which is a lymphokine that induces or enhances the expression of HLA-DR antigens. These cells, which are known to mediate cytotoxicity in an Ag-specific and Ia-restricted way, could take part in the shedding process of epithelial cells in asthma.
Asthma↗
Phagocytosis induction of chemiluminescence and chemoattractant increased superoxide anion release from activated human alveolar macrophages in asthma.
Human alveolar macrophages (AM) were demonstrated to generate reactive toxic derivatives of oxygen in many pulmonary disorders. These cells are involved in local inflammation which characterizes bronchial asthma. In the present work, we studied the ability of stimulated macrophages from healthy volunteers, and asthmatic patients to generate oxygen species in vitro. AM obtained by bronchoalveolar lavage were purified by adherence. The production of oxygen species was measured by luminol-enhanced chemiluminescence (CL) after challenge with opsonized zymosan. The maximal values were significantly (p less than 0.03 and p less than 0.01) higher in AM from asthmatics than in AM from healthy subjects. A significant correlation (p less than 0.01) was observed between maximal value of CL and the severity of asthma as assessed by the clinical score. But, no difference was observed between AM from asthmatics in a stable state and healthy subjects. On the other hand, assays for superoxide anion generation emphasized the activation state of these macrophages stimulated by formyl-peptides.
Adult↗
Inhibition by histamine of platelet-activating-factor-induced neutrophil chemotaxis in bronchial asthma.
Circulating human polymorphonuclear neutrophils are involved in asthma after their migration into the lung by local chemotactic factors. Investigation of the locomotion of neutrophils in Boyden chambers, showed that the chemotactic intensity of the platelet-activating factor (PAF) was similar in cells from healthy subjects and allergic asthmatics, although the optimal effect of the mediator was observed at 10(-6) M and 10(-8) M, respectively. Histamine had no direct chemoattractant effect on neutrophils but inhibited PAF-induced chemotaxis of neutrophils from healthy subjects and allergic asthmatics. This study provides additional evidence that neutrophils are involved in asthma, and points out the interaction between PAF and histamine in the migration of neutrophils to the lung.
Asthma↗
Increased generation of the arachidonic metabolites LTB4 and 5-HETE by human alveolar macrophages in patients with asthma: effect in vitro of nedocromil sodium.
Alveolar macrophages (AM) are the principal resident phagocytes in the human lung, and play a major role in local defence against environmental agents. It is now known that during asthma these cells take part in the amplification of the inflammatory mechanism. It has been demonstrated in vitro that they can be activated to generate leukotriene B4 (LTB4) and 5-hydroxyeicosatetraenoic acid (5-HETE), mediators with potent pharmacological properties. These two arachidonic metabolites were identified and quantified by reversed phase high performance liquid chromatography (HPLC) performed in cell suspensions, and in cell free supernatants. AM from asthmatics, after stimulation by the calcium ionophore A23187 or opsonized zymosan, released significantly (p less than 0.05) more LTB4 than those from healthy subjects. The increase in LTB4 release could be evidence for in vivo activation. On the other hand, the levels of 5-HETE in the AM from asthmatics were significantly (p less than 0.03) higher than those in cells from healthy subjects. This intracellular increase could be correlated with a greater migratory ability of these inflammatory macrophages, as observed for eosinophils. The clinical efficacy of nedocromil sodium may be partly related to the decreases in LTB4 releasability and intracellular 5-HETE levels observed only in AM from asthmatic patients.
Adolescent↗
Phosphoinositide breakdown and superoxide anion release in formyl-peptide-stimulated human alveolar macrophages. Comparison between quiescent and activated cells.
Human alveolar macrophages (AMs) from allergic asthmatics (AAs) showed continuous Li+-sensitive production of IP1, indicating that the cells were continuously activated. Furthermore, whereas the accumulation of IP1, IP2 and IP3 rapidly increased by as much as 125-175% in chemotactic-factor-stimulated AMs from healthy subjects, stimulation of cells from AAs increased these inositol phosphates only slightly. This moderate production could be due to a permanent state of activation leading to a depleted pool of polyphosphoinositides, corroborating the greater capacity of these cells to generate superoxide anion after stimulation by a chemoattractant. The activation state could be due to the action of priming agents, which are known to be released into the inflammatory sites.
Cells, Cultured↗
Evidence for specific binding of 15 hydroxyeicosatetraenoic acid to pituitary rat cells.
Specific receptors for [3H]-15 HETE have been identified on GH3 cells, a cloned strain of rat pituitary cells. With incremental inputs of radioligand and a constant cell number, specific [3H]-15 HETE binding reached a plateau indicative of saturable binding sites. Ligand analysis of the Scatchard plot demonstrated a single class of high affinity binding sites with a dissociation constant (Kd) of 0.75 nM. 12 HETE competed with radiolabeled 15 HETE (IC50 = 1 x 10(-6) +/- 0.8 M). In contrast, arachidonic acid, leukotriene B4, prostaglandins E2 and F2 alpha did not compete with [3H]-15 HETE.
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
15-Lipoxygenase products stimulate prolactin secretion from a cloned strain of rat pituitary cells.
Arachidonic acid and its lipoxygenase products may contribute to the process of prolactin (PRL) release. In the present study we investigate the role of 15-hydroperoxyeicosatetraenoic acid (15-HPETE) and 15-hydroxyeicosatetraenoic acid (15-HETE) on PRL secretion from GH3 cells. The incubation of GH3 cells with the lipoxygenase product 15-HETE significantly increased PRL release in a concentration-dependent manner. Nordihydroguaiaretic acid (NDGA), which reduces the production of arachidonate metabolites via the lipoxygenase pathway, also reduced basal and TRH or arachidonic-acid-stimulated-PRL release. The inhibitory effect of NDGA on PRL release could be overcome by the addition of 15-HETE. The time course curve of PRL release from cells challenged by 15-HETE had the same profile as that of cells stimulated by TRH. The stimulating effect of 15-HPETE (ED50 = 0.7 x 10(-9) M), which is the direct precursor of 15-HETE, on PRL release was greater than TRH or 15-HETE (ED50 = 6.5 x 10(-9) M). Furthermore 15-HPETE and 15-HETE seemed to affect the release of newly synthesized PRL. These data indicate that 15-HETE and 15-HPETE could be important intracellular components in the control of PRL secretion and may account for at least a part of arachidonate-induced PRL release from GH3.
Animals↗
Correlation in inflammatory fibroblasts between the number of glucocorticoid receptors and PGE2 release.
The anti-inflammatory effects of glucocorticoids are mediated through steroid receptor occupancy and there is a significant correlation between the extent of receptor saturation and the extent of the biological effects. In a previously published study, we found that the number of these receptors was higher in inflammatory fibroblasts than in quiescent ones. PGE2 release, measured at the same time as the number of steroid receptors, was higher when the cells were from inflammatory tissue. Our aim, in the present study, was to determine whether the PGE2 released by cells during inflammatory processes could participate in increasing the number of steroid receptors. Fibroblasts obtained from rat quiescent subcutaneous connective tissue and granulomas were subcultured in monolayers. The specific binding of [3H]dexamethasone was assessed and analyzed by a method described by Kalimi et al. After a freeze-thaw cycle, we observed a decrease in the number of receptors in inflammatory fibroblasts. When the frozen and thawed fibroblasts were subcultured in the presence of PGE2 (10(-8) M), the number of receptors was enhanced in fibroblasts from inflammatory tissue. Cycloheximide (3 X 10(-7) M) prevented this increase. The release of PGE2 decreased after freezing and then increased simultaneously with the number of receptors in inflammatory cells. These findings suggest that PGE2 may play a role in regulating steroid effects on fibroblast function.
Animals↗