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M Felaco

Publications and source records attributed to M Felaco.

At least 19 recordsLinked to original sources
PubMed2003

Oxygen supply modulates MCP-1 release in monocytes from young and aged rats: decrease of MCP-1 transcription and translation is age-related.

Hyper or hypoxia may affect the immune system's chemokine production. Monocyte chemotactic protein-1 (MCP-1), an important chemotactic cytokine can be activated by active oxygen species. Groups of rats were exposed to hypoxic and hyperoxic environmental conditions for 60 h and MCP-1 was determined in their peripheral blood mononuclear cells by Elisa and Reverse Transcriptase Polymerase Chain Reaction (RT-PCR). In this study we evaluated if the ability of monocytes to produce MCP-1 under basal conditions or after stimulation with lipopolysaccharide (LPS) or phytohaemagglutinin (PHA) was differently affected by exposure to hyper or hypoxic conditions in young and aged rats. MCP-1 expression and production in monocyte/macrophages from rats at normoxic conditions was reduced in aged subjects. However, spontaneous, LPS or PHA-induced MCP-1 production was up-regulated by exposure to hyperoxic conditions in both young (62 +/- 8, 99 +/- 7, 102 +/- 8 pg/ml, respectively) and aged rats (79 +/- 4, 112 +/- 9, 117 +/- 10 pg/ ml, respectively). We conclude that hyperoxia is an important regulator of MCP-1 release and support the hypothesis that increased % of O2 may serve to initiate MCP-1 production which then serves to recruit and regulate the distribution of mononuclear cells to the sites of inflammation.

Aging↗
PubMed2002

Inhibition of MCP-1 and MIP-2 transcription and translation by mimosine in muscle tissue infected with the parasite Trichinella spiralis.

Mimosine is a non-toxic plant aminoacid which is an effective inhibitor of DNA replication by acting at the S-phase. In this study we infected mice with T. spiralis, a nematode parasite, and studied the inflammatory response through the determination of MIP-2, a C-X-C chemokine and MCP-1, a C-C chemokine in the inflamed area around the parasitic cyst. The animals were infected and their diaphragms were tested for inflammatory response. MCP-1 and MIP-2 was tested after 1, 10, 20, 30, and 40 days post inoculation, before and after mimosine treatment. The inflammatory index was calculated by counting the white blood cells around the nematode cysts, while expression of MIP-2 and MCP-1 was calculated by ELISA method and transcription by Northern blot and RT-PCR. Here we found that mimosine strongly inhibited the inflammatory index in the diaphragmatic tissue at 10, 20, 30 and 40 days post-treatment. In these experiments, mimosine had no effect on the number of cysts produced. In addition, we found that MCP-1 transcription and translation was completely inhibited by mimosine, while MIP-2 transcription and translation was partially inhibited at 30 and 40 days; yet it was totally inhibited after 10 and 20 days in encysted diaphragm tissue infected by T. spiralis. Our studies suggest that mimosine has an inhibitory effect through the inhibition of cytoplasmatic serine hydroxymethyltransferase altering the cell cycle of white blood cells. This study suggests for the first time the premise that mimosine acts as an anti-inflammatory compound.

Animals↗
PubMed2001

Simvastatin reduces reperfusion injury by modulating nitric oxide synthase expression: an ex vivo study in isolated working rat hearts.

OBJECTIVE: We tested the hypothesis of beneficial effects of the 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA)-reductase inhibitor simvastatin in a model of ischemia-reperfusion, and investigated potential mechanisms. METHODS: Isolated working rat hearts were subjected to 15 min global ischemia and 22-180 min reperfusion in the presence or absence of simvastatin (10-100 microM). We evaluated creatinephosphokinase and nitrite levels in coronary effluent, heart weight changes, microvascular permeability (extravasation of fluoresceine-labeled albumin), ultrastructural alterations, and the expression of endothelial (e) and inducible (i) nitric oxide synthase (NOS) (by reverse-transcribed polymerase chain reaction and Western blotting) in the presence or absence of the transcriptional inhibitor actinomycin-D. RESULTS: Simvastatin (25 microM) significantly reduced myocardial damage and vascular hyperpermeability, concomitant with a reduction in endothelial and cardiomyocyte lesions. Protection became less evident at 50 microM and reverted to increased damage at 100 microM. At 25 microM, simvastatin significantly increased eNOS mRNA and protein compared with untreated hearts, probably due to a post-transcriptional regulation since unaltered by animal pretreatment with actinomycin D. Simvastatin also significantly decreased iNOS mRNA and protein, as well as nitrite production after ischemia-reperfusion. The addition of the NOS inhibitor N(pi)-nitro-L-arginine methylester (L-NAME, 30 microM) to 25 microM simvastatin-treated hearts significantly reduced cardioprotection against ischemia-reperfusion. CONCLUSIONS: In this model, in the absence of perfusing granulocytes, the acute administration of a pharmacologically relevant simvastatin concentration reduces ischemia-reperfusion injury and prevents coronary endothelial cell and cardiomyocyte damage by cholesterol-independent, NO-dependent mechanisms.

Analysis of Variance↗
PubMed2001

Histochemical and biochemical analysis of phospholipase C isoforms in normal human gastric mucosa cells.

The expression and activity of PIP2-specific phospholipase C (PLC) in healthy human gastric mucosa cells were investigated by means of Western blotting, immunohistochemistry and in vitro activity assays. The results provide direct evidence for an almost exclusive expression of the PLC beta family and at the same time supply a cellular cartography of each represented isoform of this family. In this context, the putative roles of each isoform in the signaling events regulating the gastric mucosa metabolic machinery are discussed. These data provide a unique map of the specific expression and cellular distribution of the most represented PLC isoforms in healthy human gastric mucosa cells, which may constitute a reference point in future studies aimed at highlighting possible cytochemical and biochemical hallmarks of metaplastic or malignant transformation.

Blotting, Western↗
PubMed2001

Differential expression and secretion of RANTES and MCP-1 in activated peripheral blood mononuclear cell cultures of atopic subjects.

RANTES and MCP-1 represent a link between the activation of monocytes, lymphocytes, basophils, mast cells and eosinophils in inflammatory disorders, such as the late phase allergic reaction. These C-C chemokines also play a role in regulating Th cell cytokine production and leukocyte trafficking. In this study, we determined the expression and secretion of RANTES and MCP-1 from PHA-activated PBMC of healthy and atopic subjects with no symptoms. Levels of RANTES from PHA-activated PBMC of atopic patients were higher, at 18 and 24 h incubations (42+/-5.5 and 48+/-4), compared to controls (20+/-4 and 35+/-4), respectively; while MCP-1 was not (12+/-3 and 17+/-3) compared to controls (10.5+/-3 and 15+/-2), respectively. This effect was also revealed on RANTES mRNA expression, as determined by reverse transcriptase polymerase chain reaction (RT-PCR) analysis. In addition, PHA-activated PBMC of atopic subjects produce more IL-4 (five times more) than healthy subjects, while IFN-gamma did not vary. RANTES, compared to MCP-1, may have more influence on signal transduction pathways, either in physiologic or inflammatory states and may induce profound effects on the regulation of cell activity. The differential production of RANTES and MCP-1 may lead to diverse regulation of the function and development of cells involved in the allergic response. These studies emphasize the importance of chemokine selectivity during inflammation.

Cells, Cultured↗
PubMed2001

Ultrastructural modifications and phosphatidylinositol-3-kinase expression and activity in myocardial tissue deriving from rats in different experimental conditions.

Oxygen supply is essential in the maintenance of the physiological cell metabolism. In fact, both lower and higher O2 concentrations induce modifications of the enzymatic activity of the cell which determine, in turn, morphological changes at nuclear and cytoplasmic level. Among the molecules involved in the maintenance of the cellular homeostasis, the signal transduction pathway PI-3-kinase/AKT-1 should be included. Here we suggest a relationship between the modulation of this pathway and the morphological modifications occurring "in vivo" in myocardial tissue upon hypoxic and hyperoxic stress. In particular, down regulation of this pathway, which when activated is known to deliver an anti-apoptotic signal, is concomitant to the maintenance of the apoptotic events occurring in these cells in response to oxidative stresses.

Animals↗
PubMed2001

Endothelial nitric oxide synthase (eNOS) expression and localization in healthy and diabetic rat hearts.

Several studies suggest that nitric oxide (NO) production is reduced in diabetes and that the decrease of NO may be related to the pathogenesis of diabetic endothelial damage. NO synthase (NOS) catalyses the conversion of L-arginine to L-citrulline in the presence of oxygen and NADPH-diaphorase (NADPH-d). In this study, we evaluated the expression of endothelial NOS (eNOS) enzyme and its co-enzyme in diabetic rat hearts. Male Wistar rats (n = 20, 4 mo old) and 20 male Bio Breeding Wistar (BB/W) rats of the same age were used; the Wistar rats represent the control non-diabetic rats while the BB/W rats represent the diabetic group. After the hearts were excised, the NADPH-d co-enzyme was visualized by a histochemical method and the endothelial isoform of NOS was localized by immunohistochemistry. In addition, eNOS gene expression was estimated by rt-PCR, and eNOS protein level was detected by Western blot analysis. The eNOS visualization, which involved immunoprecipitation, and the NADPH-d visualization, which involved histochemical staining, were both diminished in endothelial cells of the vascular wall of diabetic hearts, compared to non-diabetic hearts. The eNOS protein level, evaluated by Western blotting, was evident as an intense band in cardiac homogenates of non-diabetic and diabetic rats. The expression of mRNA for eNOS did not differ significantly between the two groups. These findings indicate that, in this rat heart model, diabetes does not influence the overall eNOS protein level or its mRNA level. However, there a diminution in the deposition of eNOS in cardiac endothelial cells of diabetic rats, versus non-diabetic controls, suggesting a relation between eNOS and the loss of vasodilatory response that is observed in diabetes.

Animals↗
PubMed2000

Catcholamine and nitric oxide systems as targets of chronic lead exposure in inducing selective functional impairment.

Rats were exposed for ten months to 60 ppm of lead (Pb, as acetate) in drinking water to further assess cardiovascular effects of chronic Pb exposure. At the end of the treatment, mean blood Pb was 3.1+/-0.3 microg/dL in the control rats and 22.8+/-1.2 microg/dL in the Pb-exposed rats (means+/-SE, n=12 in each group); these values were not comparable to those of humans. Pb greatly increased plasma levels of noradrenaline (NA) and adrenaline (A), but not those of L-DOPA and dopamine; monoaminoxidase activity was augmented by Pb, mostly in the aorta and in the liver; the aorta, liver, heart and kidney showed discrete histopathological alterations in the Pb-exposed rats, in which plasma levels of nitric oxide (NO, determined as L-citrulline) were reduced. Pb was able to induce blood hypertension, resulting from increase of cardiac inotropism and, mostly, total peripheral resistance. These data were discussed also in relation to those obtained in our previous studies carried out in rats exposed to Pb in drinking water (15-60 ppm) for periods ranging from five to eighteen months. Pb appeared to increase both sympathetic nerve activity by central mechanisms (thus increasing plasma NA and A) and cyclic adenosine monophosphate (cAMP)-dependent availability of calcium ions (Ca++) for contractile mechanisms in the vascular and cardiac myocells (also through an increased vascular alpha2- and myocardial beta1-adrenoreceptor reactivity). The reduction of plasma NO, contributing to increase vascular resistance and cardiac inotropism, was explained as a result of actions of Pb on enzyme activities concerned with the kallikrein-kinin (KK) and renin-angiotensin-aldosterone (RAA) systems. It was concluded that chronic Pb exposure is able to affect selective neuroendocrine (i.e., catecholamine), au- tacoidal (i.e., KK and RAA) and transductional pathways (i.e., cAMP, NO, Ca++) involved in the cardiovascular function.

Animals↗
PubMed2000

Endothelial NOS expression and ischemia-reperfusion in isolated working rat heart from hypoxic and hyperoxic conditions.

Induction of endothelial nitric oxide synthase (eNOS) contributes to the mechanism of heart protection against ischemia-reperfusion damage. We analyzed the effects of hypoxia and hyperoxia on eNOS expression in isolated working rat hearts after ischemia-reperfusion damage. Adult male Wistar rats were submitted to chronic hypoxia (2 weeks) and hyperoxia (72 h). The hearts were submitted to 15 min of ischemia and reperfused for 60 min, then we evaluated hemodynamic parameters and creatine phosphokinase (CPK) release. eNOS expression was estimated by RT-PCR; enzyme localization was evaluated by immunohistochemistry and the eNOS protein levels were detected by Western blot. All hemodynamic parameters in hypoxic conditions were better with respect to other groups. The CPK release was lower in hypoxic (P<0.01) than in normoxic and hyperoxic conditions. The eNOS deposition was significantly higher in the hypoxic group versus the normoxic or hyperoxic groups. The eNOS protein and mRNA levels were increased by hypoxia versus both other groups. Chronic hypoxic exposure may decrease injury and increase eNOS protein and mRNA levels in heart subjected to ischemia-reperfusion.

Animals↗
PubMed2000

Localization of the e-NOS enzyme in endothelial cells and odontoblasts of healthy human dental pulp.

Nitric oxide synthases (NOS) are important enzymes present in different cells such as endothelial cells, macrophages, etc. Recently, it has been found that nitric oxide (NO) is responsible for vasodilation, blood pressure regulation, platelet aggregation, cardiac contractility, and the mediation of immunity during bacterial infections and inflammation. However, the production and role of NO in various structures of the oral cavity have not been investigated extensively. The aim of this study was to evaluate the presence of e-NOS in healthy human odontoblasts and endothelial cells of the dental pulp. Twenty healthy human dental pulps were collected and frozen and pulp slices were obtained using a cryostat. The e-NOS enzyme was revealed by immunohistochemical analysis and the enzyme level was detected by Western blotting and mRNA expression by RT-PCR. The immunohistochemical results demonstrated, for the first time, the presence of e-NOS in odontoblasts and in endothelial cells. The presence of e-NOS m-RNA was confirmed by RT-PCR and the expression of the protein by Western blotting. These results clearly show that the e-NOS enzyme is present in both odontoblasts and endothelial cells of healthy human pulp. The presence of e-NOS in the odontoblast and endothelial cells of the dental pulp may mediate local vasodilation and cell proliferation.

Blotting, Western↗
PubMed1999

Effect of electromagnetic fields on several CD markers and transcription and expression of CD4.

We carried out flow cytometric analysis for multiparametric evaluation of cell surface markers related to cellular functions. Specifically, we studied the expression of CD4, CD8, CD3, CD16, CD19, HLA-DR, and CD14 macrophage receptors expression and cell cycle progression on cells exposed to ELF-EMF. In addition, we tested the effects of ELF-EMF on CD4 mRNA protein transcription and translation and the cell-cycle progression using an immunofluorescence method. Our data show that same CD surface marker expression are weakly influenced by electromagnetic fields, with no differences between cells exposed or not exposed to ELF-EMFs. However, when the CD4 protein generation was studied, an indication of protein production was found in lymphocytes exposed to ELF-EMF, as evidenced by immunofluorescence, Western blotting and RT-PCR analysis. CD16 and CD14 expression were affected by EMF exposure at all times studied (24, 48, 72 h). The results obtained with cell cycle analysis show that after 48 h of exposure to ELF-EMF, PHA-activated and not activated cells in S phase increase with respect to non-exposed cells. The findings from this study demonstrate that under our defined experimental conditions there is evidence that ELF-EMF has a slight effect on CD4, CD14 and CD16 receptor expression, while the other CD receptors are not affected.

Antigens, CD↗
PubMed1999

Impact of extremely low frequency electromagnetic fields on CD4 expression in peripheral blood mononuclear cells.

There is increasing evidence suggesting that extremely low frequency electromagnetic fields (ELF-EMF) may influence several cell functions. Here the effects of ELF-EMF were studied on the expression of CD4+ cell surface receptors of human peripheral blood mononuclear cells (PBMC) using fluorescence-activated cell sorter (FACScan). The expression of CD4+ in ELF-EMF exposed (24, 48 and 72 h) and not exposed PBMC were not statistically significant. In addition, a flow cytometric analysis was determined by using a fluorescent labeled antibody, at 24 and 72 h incubations. The amount of bound antibody was distributed with a slight difference in the ELF-EMF-exposed PBMC compared to the not exposed cells. Moreover, DNA CD4+ expression in PBMC strongly increased in exposed cells, resting and activated with Phytohaemaglutinin (PHA). When polymerase chain reaction was performed on CD4+ mRNA of PBMC an increase of CD4+ mRNA expression was found after the resting cells were exposed to ELF-EMF at 24 h compared to not exposed cells, while at 48 and 72 h no difference was found. In the cell cycle progression analysis, the PBMC exposed to ELF-EMF presented a significant increase of percentage expression of cell cycle progression in the S phase compared to not exposed cells; while in G1 and G2 phases, there were no differences. Our results provide new evidence that ELF-EMF can affect CD4+ expression in PBMC and describe an additional biological activity for ELF-EMF affecting CD4+ transcription and translation protein and the increase of the percentage expression of the cell cycle progression of the S phase.

Blotting, Western↗
PubMed1998

Does chronic hypoxia increase rat carotid body nitric oxide?

The carotid body plays an important role in ventilatory adaptation during chronic hypoxia. Nitric oxide (NO) may act as a regulator in neurotransmission, influencing the carotid body chemosensory discharge. The aim of the study was to understand if NO could contribute to the adaptation process during chronic hypoxia. The rats were kept in chronic hypoxia (10-11% inspired oxygen) for 12 days, while the controls were kept in room air (21% O2). The distribution for diaphorase activity and immunohistochemistry for nitric oxide synthase (NOS) showed that chronic hypoxia induces an increase in NOS activity in the carotid body. It was concluded that NO release increased during chronic hypoxia and causes an inhibitory effect on carotid chemosensory discharge of the rat carotid body.

Adaptation, Physiological↗
PubMed1998

Sodium nitroprusside, a NO donor, modifies Ca2+ transport and mechanical properties in frog skeletal muscle.

Recently it has been hypothesized that, in skeletal muscle, NO produced directly by high-frequency stimulation could produce contraction through reactions with thiol groups on the sarcoplasmic reticulum (SR). However, a possible cGMP-mediated relaxing effect, similar to that seen in smooth muscle, has also been demonstrated. We used purified SR preparations and single fibres from frog fast muscles incubated with different concentrations of sodium nitroprusside (SNP) in this study. The results obtained from a long low-frequency stimulation, together with those from a study on Ca2+ transport regulation, showed that the presence of NO precursor induced: an acceleration of the onset of fatigue in single fibres; a decreased vesicular Ca2+ content due to increased Ca2+ release; a shift to open status in SR Ca2+ channels; an increase in SR Ca2+ pump activity. The data presented in this paper seem to indicate that the increased NO in the muscle fibres can influence muscle activity in different ways, perhaps depending on the metabolic status of the muscle and target (filaments, sarcolemma, SR) with which the NO (or its derivatives) acts.

Animals↗
PubMed1998

Induction of monocyte chemotactic protein-1 (MCP-1) and TNF alpha by Trichinella spiralis in serum of mice in vivo.

MCP-1 is a small (8-10 KDa) protein and a prototype member of the CC chemokine beta subfamily, which plays a critical role in acute and chronic inflammation. Recent evidence suggests an important role for MCP- 1, MCP-2 and MCP-3 in a number of pathological states, including delayed type hypersensitivity conditions, parasitic infections and rheumatoid arthritis. Forty BALB-c mice were treated with the parasite Trichinella spiralis. After the infection the animals were sacrificed at different periods from the initial infection and MCP-1 and TNFalpha were quantified in the mouse serum. The level of MCP-1 in the serum of mice infected with 100 larvae increases from 27.5+/-7.0 pg/ml at day 23, to a maximum level of 31.5+/-5.0 pg/ml at day 33, then decreased to 14.6+/-2.0 pg/ml at day 47. When the mice were infected with 200 larvae of T. spiralis the maximum increase was 34.4+/-2.5 pg/ml found on day 23. From day 33 to day 47 MCP-1 levels were decreased. In addition, in infected mice levels of TNFalpha were detectable in the serum as early as day 1. The level of TNFalpha was maximum at day 35 (3812+/-224 pg/ml). Serum from non-infected mice contained no detectable levels of either MCP-1 or TNFalpha. However, even if MCP-1 seems to be implicated in Trichinellosis, its exact role and function in inflammatory parasitic diseases remains to be determined.

Animals↗
PubMed1998

Mast cell recruitment after subcutaneous injection of RANTES in the sole of the rat paw.

The effect of hrRANTES was studied after the injection in the sole of the rat paw, an area particularly rich in mast cells. Subcutaneous injections of RANTES 50 ng/10 microl produced an erythematous reaction which was inhibited by anti-RANTES antibody 50 microg/rat injected in the tail vein 30 min before hrRANTES 50 ng/10 microl was injected. In another set of experiments the animals were injected subcutaneously in the sole of the paw with PBS 10 microl (control), LPS (100 ng/10 microl) hrRANTES 50 ng/10 microl or anti-RANTES 50 microl/rat injected in the tail vein 30 min before hrRANTES 50 ng/10 microl was injected. The biopsies were analysed after 4 h and counted in an optic field. hrRANTES produced a strong recruitment of mast cells selectively coloured with 0.1% toluidine blue and inhibited by anti-RANTES antibody. In addition to the optical and electron microscope study, in some of the excised tissue Northern blot analysis for histidine decarboxylase (HDC) mRNA was performed to estimate the amount of histamine generation in the tissue of the injection sites. We found that subcutaneous injections of hrRANTES 50 ng/10 microl in the sole of the rat paw produced an accumulation of a great number of mast cells compared to PBS 10 microl (negative control) or LPS 100 ng/10 microl (positive control) after 4 h. The hrRANTES effect was inhibited by anti-RANTES antibody injected in the tail vein 30 min before hrRANTES exposure. Moreover, hrRANTES increased HDC mRNA and histamine generation.

Animals↗