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Biomedical subjects

I Bonilla

Publications and source records attributed to I Bonilla.

At least 19 recordsLinked to original sources
PubMed2004

Myeloperoxidase release after allergen-specific conjunctival challenge.

BACKGROUND: Allergen-specific conjunctival challenge is a fruitful and complete tool in evaluating pathophysiological phenomena of allergic inflammation. After challenge, a significant neutrophil infiltrate occurred in allergic subjects. The primary (azurophilic) granules of neutrophils contain a variety of enzymes that might potentiate inflammation, such as myeloperoxidase (MPO). It is not known whether allergen-specific conjunctival challenge (ASCC) is able to elicit MPO release. We also investigated the possible role of immunotherapy (IT) in the release of MPO. METHOD: The groups studied included Dactylis glomerata-sensitive adult atopic patients suffering from seasonal allergic rhinoconjunctivitis, and healthy adult nonatopic volunteer controls. One group of allergic patients received no specific hyposensitization (not-IT allergic group). A second group of allergic patients had been immunotherapy-treated with Dactylis glomerata extract for the preceding three years and continued to receive a maintenance dose within the highest potency of the extract (IT-allergic group). ASCC with Dactylis glomerata was performed outside the pollen season in all subjects. Myeloperoxidase was assayed by MPO-enzyme immunoassay method. RESULTS: Thirty minutes after challenge, myeloperoxidase levels in the non-immunotherapy allergic patients were significantly higher compared than in the healthy group (p<0.001). The levels of myeloperoxidase released in the immunotherapy allergic group were significantly lower than those in the nonimmunotherapy allergic group (p<0.001) and higher than those in nonallergic subjects (p<0.001). CONCLUSION: These results indicate that after ASCC there is a release of MPO. Our study suggests that immunotherapy actively modifies the release of MPO after ASCC.

Allergens↗
PubMed2004

A calcium signal is involved in heterocyst differentiation in the cyanobacterium Anabaena sp. PCC7120.

The impact of calcium signals in virtually all cells has led to the study of their role in prokaryotic organisms as stress response modulators. Cell differentiation in adverse conditions is a common Ca(2+)-requiring response. Nitrogen starvation induces the differentiation of N(2)-fixing heterocysts in the filamentous cyanobacterium Anabaena sp. PCC7120. This paper reports the use of a recombinant strain of this organism expressing the photoprotein aequorin to monitor the intracellular free-calcium concentration during the course of heterocyst differentiation. A specific calcium signature that is triggered exclusively when cells are deprived of combined nitrogen and generated by intracellular calcium stores was identified. The intracellular calcium signal was manipulated by treatment with specific calcium drugs, and the effect of such manipulation on the process of heterocyst differentiation was subsequently assessed. Suppression, magnification or poor regulation of this signal prevented the process of heterocyst differentiation, thereby suggesting that a calcium signal with a defined set of kinetic parameters may be required for differentiation. A hetR mutant of Anabaena sp. PCC7120 that cannot differentiate into heterocysts retains, however, the capacity to generate the calcium transient in response to nitrogen deprivation, strongly suggesting that Ca(2+) may be involved in a very early step of the differentiation process.

Adaptation, Physiological↗
PubMed2003

Allergen-dependent CD14 modulation and apoptosis in monocytes from allergic patients.

BACKGROUND: CD14 is a most important monocyte surface molecule. Recently, it has been reported that there is an important relationship between CD14 and immunoglobulin E, and that regulation of CD14 expression is an effector mechanism mediating apoptosis of monocytes. OBJECTIVE: The present study was designed to determine whether specific allergens were able to modulate CD14 expression and apoptosis by monocytes from allergic patients or whether specific immunotherapy (IT) might affect these processes. METHODS: One group of adult allergic asthmatic patients had received IT for the previous 3 years. Another similar group was not treated with IT. We challenged peripheral blood monocytes from both groups of asthmatic patients in vitro with the specific allergen that produced clinical symptoms in asthmatic patients. The cells were also challenged with allergen to which the patients were not sensitive. Monocytes from normal subjects were also challenged with allergens. Expression of CD14 on the monocyte surface was analyzed by flow cytometry, and soluble CD14 (sCD14) in culture supernatant by enzyme-linked immunosorbent assay. The three groups of subjects were challenged with allergens, and apoptosis was analyzed by flow cytometry. RESULTS: When monocytes from non-IT-treated asthmatic patients were cultivated with the allergens to which the patients were sensitive, a significant up-regulation on the monocyte surface was observed compared with results from the healthy group (P < 0.003) and from the IT asthmatic group (P < 0.003). A significantly higher sCD14 level was observed in the culture supernatant of the monocytes from the IT asthmatic group were observed compared with those from the healthy group (P < 0.001) and those from the non-IT asthmatic group (P < 0.001). A significantly higher apoptosis level was observed in monocytes from the IT asthmatic group compared with those from the healthy group (P < 0.001) and those from the non-IT asthmatic group (<0.001). CONCLUSIONS: We present evidence that the expression of CD14 on the surface of monocytes and the apoptosis of the same cells can be modulated by an allergen-dependent mechanism. These processes can be affected by IT.

Adult↗
PubMed2002

Respiratory burst in neutrophils from asthmatic patients.

BACKGROUND: The role of oxygen radicals has been implicated in disease processes of asthma. We have previously shown that specific allergens were able to activate respiratory burst by neutrophils from allergic patients sensitized to allergens of the same type as those which produce clinical allergy. OBJECTIVES: In this study, we attempted to evaluate the production of respiratory burst by an anti-IgE Ab in neutrophils from asthmatic allergic patients (with and without immunotherapy treatment) and in neutrophils from healthy subjects. METHOD: Neutrophils were stimulated by 10 microg/mL of anti-IgE Ab for 15 min at 37 degrees C. The production of respiratory burst from neutrophils was assayed by luminol-amplified chemiluminescence method. RESULTS: The respiratory burst was significantly higher in neutrophils from non-IT-asthmatic patients than in neutrophils from both healthy (p < 0.001) and IT-asthmatic (p < 0.001) groups. The IT-asthmatic group presented levels of respiratory burst approximately equal to those from non-allergic subjects (p=0.426). CONCLUSIONS: We conclude that neutrophils obtained from allergic asthmatic patients have an increased propensity to generate respiratory bursts, in comparison with neutrophils from healthy subjects. Immunotherapy actively modifies the respiratory burst by neutrophils from allergic asthmatic patients.

Antibodies↗
PubMed2001

IgE-mediated downregulation of L-selectin (CD62L) on lymphocytes from asthmatic patients.

BACKGROUND: L-selectin (CD62L) mediates the binding of lymphocytes to high endothelial venules of peripheral lymph nodes and is also involved in leukocyte attachment to the endothelium at sites of inflammation. Although it has been demonstrated that L-selectin is shed after lymphocyte activation, it is unknown whether the expression of L-selectin on the surface of lymphocytes can be modulated by an IgE-dependent mechanism or whether immunotherapy (IT) might affect this mechanism. METHODS: One group of adult allergic asthmatic patients had received IT for the previous 3 years. Another similar group was not treated with IT. We challenged peripheral blood lymphocytes from both groups of asthmatic patients in vitro with an anti-IgE antibody (Ab). Expression of L-selectin on the lymphocyte surface was analyzed by flow cytometry, and the levels of soluble L-selectin (sL-selectin) on culture supernatant by ELISA. RESULTS: L-selectin was downregulated from the surface of lymphocytes in a time- and anti-IgE antibody dose-dependent manner (with a concomitant upregulation of shed L-selectin in the supernatant). When lymphocytes from non-IT asthmatic patients were cultivated with anti-IgE Ab, a statistically significantly greater CD62L downmodulation on the lymphocyte surface was observed compared with lymphocytes from the healthy group (P<0.002) and from the IT-asthmatic group (P<0.001). When lymphocytes from non-IT asthmatic patients were cultivated with anti-IgE Ab, a significantly greater sL-selectin level in the culture supernatant was observed compared with lymphocytes from the healthy group (P<0.001) and with lymphocytes from IT-asthmatic group (P<0.001). CONCLUSIONS: We present evidence that the expression of L-selectin on the surface of lymphocytes can be modulated by an IgE-dependent mechanism. This mechanism can be affected by IT.

Antibodies↗
PubMed2001

IgE-dependent release of myeloperoxidase by neutrophils from allergic patients.

BACKGROUND: The three forms of IgE receptor: the heterotrimeric high-affinity receptor for IgE (FcepsilonRI), the low-affinity receptor for IgE (FcepsilonRII/CD23) and the Mac-2/IgE-binding protein (epsilonBP), have previously been found on human neutrophils. We have previously shown that specific allergens are able to activate functional responses by neutrophils from allergic patients sensitized to those allergens. Neutrophils are present in the sites of allergic inflammation. The primary (azurophilic) granules of neutrophils contain a variety of enzymes that might potentiate inflammation, such as myeloperoxidase (MPO). It is not known whether specific allergens are able to elicit MPO release by neutrophils from allergic patients. METHODS: Neutrophils were challenged in vitro with the specific allergen that produced clinical symptoms in asthmatic patients. Also, the cells were challenged with allergens that the patients were not sensitive to. Neutrophils from normal subjects were also challenged with allergens. RESULTS: The in vitro challenge of neutrophils with allergens that the patients were sensitive to elicited a release of MPO by these cells. The in vitro activation of neutrophils was highly allergen-specific, in such a way that allergens other than those accounting for clinical symptoms did not evoke MPO release, and allergens were ineffective on neutrophils from healthy donors. CONCLUSION: An IgE-dependent mechanism might promote MPO release by neutrophils at allergic sites.

Adult↗
PubMed2001

Elevated secretion of myeloperoxidase by neutrophils from asthmatic patients: the effect of immunotherapy.

BACKGROUND: There is increasing evidence of neutrophil participation in asthma and the allergic process. After activation, neutrophils release myeloperoxidase (MPO) together with other granule enzymes. OBJECTIVES: In this study we attempted to evaluate the release of MPO in vitro by neutrophils from asthmatic patients and the relationship between neutrophil degranulation and lung function, measured as FEV(1), of the patients. We also investigated the possible role of immunotherapy in the release of MPO by neutrophils. METHODS: Neutrophils were stimulated with formyl-methionyl-leucyl-phenylalanine for 45 minutes at 37 degrees C. MPO released from neutrophils was assayed by using an MPO enzyme immunoassay. RESULTS: Neutrophils released statistically significantly higher MPO levels in the asthmatic patients not receiving immunotherapy than in the healthy group. A significant inverse correlation was observed in the asthmatic group not receiving immunotherapy between MPO secretion and lung function, measured as FEV(1), of the patients. Neutrophils of the asthmatic group receiving immunotherapy released significantly less MPO than did those of the asthmatic group not receiving immunotherapy, with MPO levels equal to those from nonallergic subjects. CONCLUSIONS: We conclude that neutrophils obtained from allergic asthmatic patients have an increased propensity to release MPO. The experiments described here provide evidence that there is a significant inverse relationship between levels of MPO released by neutrophils from allergic patients and lung function, as assessed by FEV(1). Our study suggests that immunotherapy actively modifies the release of MPO in vitro by neutrophils from allergic asthmatic patients.

Adult↗
PubMed2001

Lectin-like glycoprotein PsNLEC-1 is not correctly glycosylated and targeted in boron-deficient pea nodules.

Symbiosome development was studied in pea root nodules from plants growing in the absence of boron (B). Rhizobia released into the host cells of nodules from B-deficient plants developed to abnormal endophytic forms with an altered electrophoretic lipopolysaccharide pattern. Immunostaining after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electroblotting of nodule homogenates with antibodies that recognize glycoprotein components showed that two previously described lectin-like glycoproteins (PsNLEC-1A and PsNLEC-1B) did not harbor the carbohydrate epitope normally recognized by specific monoclonal antibodies. Material derived from B-deficient nodules, however, still contained three antigenic isoforms with similar electrophoretic mobilities to PsNLEC-1 isoforms A, B, and C. These could be detected following immunoblotting and immunostaining with a specific antiserum originating from the purified PsNLEC protein that had been heterologously expressed in Escherichia coli. Immunogold localization of PsNLEC-1 sugar epitopes in B-deficient nodules showed that they were associated mostly with cytoplasmic vesicles rather than normal localization in the symbiosome compartment of mature infected cells. These results suggest that a modification of the glycosyl-moieties of PsNLEC-1 and an alteration of vesicle targeting occur during the development of pea nodules in the absence of B, and that these changes are associated with the development of aberrant nonfunctional symbiosomes.

Animals↗
PubMed2000

MucR is necessary for galactoglucan production in Sinorhizobium meliloti EFB1.

Sinorhizobium meliloti can produce two types of acidic exopolysaccharides, succinoglycan and galactoglucan, that are interchangeable for infection of alfalfa nodules. Strain SU47 and derivatives produce only succinoglycan, unless it grows under phosphate limitation or carries a mutation in either of two regulatory loci, mucR or expR. It has been proposed that MucR acts as a transcriptional repressor that blocks the expression of the exp genes responsible for galactoglucan production. Strain EFB1 simultaneously produces both exopolysaccharides. Heterologous expression of lacZ transcriptional fusions of the expE promoters has shown that genetic background is more important that promoter sequence for exp gene expression, since expE promoters from both strains are expressed at high level in EFB1 and not in SU47. We have found that mucR is present in mucoid and nonmucoid strains, and in EFB1 differs from SU47 in only one conservative amino acid change. MucR proteins from both strains are interchangeable. An mucR mutant of EFB1 cannot produce galactoglucan and does not express mucS.

Bacterial Proteins↗
PubMed2000

Use of recombinant aequorin to study calcium homeostasis and monitor calcium transients in response to heat and cold shock in cyanobacteria.

We investigated the possibility of Ca(2+) signaling in cyanobacteria (blue-green algae) by measuring intracellular free Ca(2+) levels ([Ca(2+)](i)) in a recombinant strain of the nitrogen fixing cyanobacterium Anabaena strain sp. PCC7120, which constitutively expresses the Ca(2+)-binding photoprotein apoaequorin. The homeostasis of intracellular Ca(2+) in response to increasing external Ca(2+) has been studied in this strain. The resting level of free Ca(2+) in Anabaena was found to be between 100 and 200 nM. Additions of increasing concentrations of external Ca(2+) gave a transient burst of [Ca(2+)](i) followed by a very quick decline, reaching a plateau within seconds that brought the level of [Ca(2+)](i) back to the resting value. These results indicate that Anabaena strain sp. PCC7120 is able to regulate its internal Ca(2+) levels. We also monitored Ca(2+) transients in our recombinant strain in response to heat and cold shock. The cell's response to both stresses was dependent on the way they were induced. The use of inhibitors suggests that heat shock mobilizes cytosolic Ca(2+) from both intracellular and extracellular sources, while the Ca(2+) source for cold shock signaling is mostly extracellular.

Aequorin↗
PubMed2000

PCR use of highly conserved DNA regions for identification of Sinorhizobium meliloti.

A PCR identification method in which four primers that recognize homologous conserved regions in the Sinorhizobium meliloti genome are used was developed and tested. The regions used for identification were the nodbox 4 locus, which is located in one of the symbiotic megaplasmids, and the mucR gene, which is located in the chromosome. The new method was used to establish a collection of S. meliloti strains from polluted soils.

Bacterial Proteins↗
PubMed1998

Exopolysaccharide II production is regulated by salt in the halotolerant strain Rhizobium meliloti EFB1.

The halotolerant strain Rhizobium meliloti EFB1 modifies the production of extracellular polysaccharides in response to salt. EFB1 colonies grown in the presence of 0.3 M NaCl show a decrease in mucoidy, and in salt-supplemented liquid medium this organism produces 40% less exopolysaccharides. We isolated transposon-induced mutant that, when grown in the absence of salt, had a colony morphology (nonmucoid) similar to the colony morphology of the wild type grown in the presence of salt. Calcofluor fluorescence, proton nuclear magnetic resonance spectroscopy, and genetic analysis of the mutant indicated that galactoglucan, which is not produced under normal conditions by other R. meliloti strains, is produced by strain EFB1 and that production of this compound decreases when the organism is grown in the presence of salt. The mutant was found to be affected in a genetic region highly homologous to genes for galactoglucan production in R. meliloti Rm2011 (expE genes). However, sequence divergence occurs in a putative expE promoter region. A transcriptional fusion of the promoter with lacZ demonstrated that, unlike R. meliloti Rm2011, galactoglucan is produced constitutively by EFB1 and that its expression is reduced 10-fold during exponential growth in the presence of salt.

Base Sequence↗