Search PubMed⌕ Search

Biomedical subjects

S Finotto

Publications and source records attributed to S Finotto.

17 recordsLinked to original sources
PubMed1999

Cutting edge: chronic intestinal inflammation in STAT-4 transgenic mice: characterization of disease and adoptive transfer by TNF- plus IFN-gamma-producing CD4+ T cells that respond to bacterial antigens.

We generated transgenic mice for STAT-4, a regulatory protein specifically associated with IL-12 signaling, under the control of a CMV promoter. These mice expressed strikingly increased nuclear STAT-4 levels in lamina propria CD4+ T lymphocytes upon systemic administration of dinitrophenyl-keyhole limpet hemocyanin and developed chronic transmural colitis characterized by infiltrates of mainly CD4+ T lymphocytes. The latter cells produced predominantly TNF and IFN-gamma but not IL-4 upon activation with alphaCD3/CD28 or autologous bacterial Ags, consistent with a Th1-type cell response. Furthermore, chronic colitis in STAT-4 transgenic mice could be adoptively transferred to SCID mice by colonic and splenic CD4+ T cells that were activated with Ags from autologous bacterial flora. These data establish a critical molecular signaling pathway involving STAT-4 for the pathogenesis of chronic intestinal inflammation, and targeting of this pathway may be relevant for the treatment of colitis in humans.

Adoptive Transfer↗
PubMed1999

Tumour necrosis factor (TNF) production by T cell receptor-primed T lymphocytes is a target for low dose methotrexate in rheumatoid arthritis.

Methotrexate (MTX) is an effective immunosuppressive agent in various chronic inflammatory diseases such as rheumatoid arthritis (RA). However, its mechanisms of action are only partially understood. In this study, we assessed the effects of MTX on the differentiation of peripheral blood (PB) CD4+CD45RA 'naive' and CD4+CD45RO 'memory' T cells from healthy controls and patients with RA. Accordingly, purified T cells were primed and restimulated in vitro via the T cell receptor (TCR) in the presence of IL-2 to generate effector T cells secreting large amounts of Th1 and Th2 cytokines. We observed that low doses of MTX strongly suppress TNF and to a lesser extent interferon-gamma (IFN-gamma) production by T cells from both healthy donors and RA patients when present during T cell priming via the TCR. Similar data were obtained for TCR-primed synovial fluid mononuclear cells in RA. In contrast, production of IL-4 by TCR-primed CD45RA T cells was significantly increased upon MTX treatment. Interestingly, MTX did not enhance IL-4 production when present during restimulation of effector CD45RO T cells, although it still suppressed TNF production. The results indicate that MTX effects depend on the stage of T cell activation and identify TNF production by TCR-primed T lymphocytes as a target for low-dose MTX treatment in RA. These findings could explain the delayed clinical effects of MTX and may contribute to its potent anti-inflammatory and immunoregulatory properties.

Adult↗
PubMed1999

Analysis of mice carrying targeted mutations of the glucocorticoid receptor gene argues against an essential role of glucocorticoid signalling for generating adrenal chromaffin cells.

Molecular mechanisms underlying the generation of distinct cell phenotypes is a key issue in developmental biology. A major paradigm of determination of neural cell fate concerns the development of sympathetic neurones and neuroendocrine chromaffin cells from a common sympathoadrenal (SA) progenitor cell. Two decades of in vitro experiments have suggested an essential role of glucocorticoid receptor (GR)-mediated signalling in generating chromaffin cells. Targeted mutation of the GR should consequently abolish chromaffin cells. The present analysis of mice lacking GR gene product demonstrates that animals have normal numbers of adrenal chromaffin cells. Moreover, there are no differences in terms of apoptosis and proliferation or in expression of several markers (e.g. GAP43, acetylcholinesterase, adhesion molecule L1) of chromaffin cells in GR-deficient and wild-type mice. However, GR mutant mice lack the adrenaline-synthesizing enzyme PNMT and secretogranin II. Chromaffin cells of GR-deficient mice exhibit the typical ultrastructural features of this cell phenotype, including the large chromaffin granules that distinguish them from sympathetic neurones. Peripherin, an intermediate filament of sympathetic neurones, is undetectable in chromaffin cells of GR mutants. Finally, when stimulated with nerve growth factor in vitro, identical proportions of chromaffin cells from GR-deficient and wild-type mice extend neuritic processes. We conclude that important phenotypic features of chromaffin cells that distinguish them from sympathetic neurones develop normally in the absence of GR-mediated signalling. Most importantly, chromaffin cells in GR-deficient mice do not convert to a neuronal phenotype. These data strongly suggest that the dogma of an essential role of glucocorticoid signalling for the development of chromaffin cells must be abandoned.

Adrenal Glands↗
PubMed1997

Glucocorticoids decrease tissue mast cell number by reducing the production of the c-kit ligand, stem cell factor, by resident cells: in vitro and in vivo evidence in murine systems.

The local delivery of glucocorticoids to tissues significantly decreases mast cell number. This pharmacologic effect of glucocorticoids is believed to be one of the mechanisms by which glucocorticoids regulate allergic inflammation. To determine the mechanism by which glucocorticoids are able to exert this effect, we first applied the glucocorticoid fluocinonide to mouse dermis and observed that the decrease in mast cell number was associated with an increase in mast cell apoptosis. This did not appear to be due to a direct effect of the glucocorticoid on mast cells, as the addition of 0.01-1.0 microM of the glucocorticoid dexamethasone into stem cell factor (SCF)-dependent mast cell cultures did not enhance mast cell death. However, addition of dexamethasone to cultured fibroblasts did result in a downregulation of SCF mRNA and a significant decrease in SCF protein production. Similarly, immunohistochemistry performed on fluocinonide-treated mouse dermis revealed a decrease in immunoreactive SCF. Administration of SCF at sites of fluocinonide administration to the dermis abolished the mast cell-depleting effect of this glucocorticoid. Thus, glucocorticoids decrease tissue mast cell number by downregulating tissue SCF production required for the survival of local mast cells. This observation may be applicable to the design of improved strategies to treat mast cell-mediated disorders.

3T3 Cells↗
PubMed1997

Generation of cell diversity in the peripheral autonomic nervous system: the sympathoadrenal cell lineage revisited.

Based on recent evidence from in vitro and gene knock-out/knock-in studies this short review summarizes the molecular scenario underlying the development of autonomic neurons from the neural crest. The focus is on the sympathoadrenal (SA) cell lineage. While migrating ventrally precursors of this cell lineage are exposed to signals from notochord/ventral neural tube probably including the protein sonic hedgehog. These and signals in the region of the dorsal aorta (members of the family of bone morphogenetic proteins), where SA progenitor cells subsequently assemble, are essential for the induction of the adrenergic phenotype. SA progenitor cells subsequently differentiate into paravertebral and prevertebral sympathetic neurons, intra- and extra-adrenal chromaffin cells and intermediate SIF (small intensely fluorescent) cells. Based on in vitro studies with isolated SA and chromaffin progenitor cells glucocorticoids have been claimed as essential for suppressing a neuronal commitment and channeling SA cells towards the chromaffin phenotype. Unexpectedly, mice deficient for a functional glucocorticoid receptor possess the full complement of adrenal chromaffin cells at birth. We present a hypothetical scenario consistent with these data, in which chromaffin cell development would be the default pathway in the SA cell lineage, while development into a neuronal direction requires specific growth factor signaling, which is probably distinct for paravertebral and prevertebral sympathetic neurons.

Animals↗
PubMed1996

Specific inhibition of beta-tryptase expression in a human mast cell line by granulocyte-macrophage colony-stimulating factor produced by airways structural cells.

The growth and differentiation of mast cells are regulated by cytokines produced in tissue microenvironments. We previously reported that mast cells isolated from the epithelial compartment of nasal polyp tissue contain significantly less tryptase when compared with mast cells isolated from the stroma of the same tissue. In an attempt to explore this finding, we analyzed the ability of supernatants obtained from cultured nasal polyp epithelial cells (NP-EpCM) or nasal polyp fibroblasts (NP-FbCM) to regulate the tryptase content of the immature human mast cell line HMC-1. HMC-1 cells were cultured for 7 days in Iscove's modified Dulbecco's medium (IMDM) with 30% of either NP-FbCM or NP-EpCM or 20% MoCM (supernatant of a leukemic T cell line). As assessed by radioimmunoassay and test for enzymatic activity, all three conditioned media were shown to significantly decrease tryptase protein expression in HMC-1, when compared with cultures performed with IMDM alone (NP-EpCM P < 0.001; NP-FbCM P < 0.04; MoCM P < 0.004). In addition, Northern blot analysis demonstrated lower tryptase mRNA levels upon exposure to all three conditioned media tested, suggesting that tryptase downregulation occurs at the transcriptional level. In further studies we found that preincubation of MoCM with anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) completely blocked the observed downregulation of tryptase expression mediated by this conditioned medium. The findings suggest that GM-CSF has a suppressive effect on expression of protease in mast cells, and may thus play a modulatory role in determining the extent of tissue inflammation in allergic airways disease.

Blotting, Northern↗
PubMed1995

A subset of patients with recurrent spontaneous abortion is deficient in transforming growth factor beta-2-producing "suppressor cells" in uterine tissue near the placental attachment site.

PROBLEM: To determine if patients with unexplained recurrent miscarriage have a deficiency of decidual immunosuppressor cells that produce transforming growth factor beta type 2, as has been found in mice with abortion due to rejection and/or trophoblast failure. METHODS: Decidual biopsy specimens were taken as near to the placental attachment site as possible under ultrasound guidance from first trimester legal termination (control) patients with recurrent miscarriage and non-viable pregnancy, and from patients with sporadic missed abortion. The tissue was tested for TGF beta-2+ suppressor cells by in situ hybridization, immunohistochemistry, and analysis of supernatants. RESULTS: TGF beta-2-related suppressor molecules similar but not identical to those identified in pregnant mice were released by decidual lymphoid cells. Fifty percent of 14 recurrent miscarriage patients showed a lack of suppressor cells and 59% were subnormal in comparison to 20 controls and 5 sporadic miscarriage patients, where 80-85% of the patients had detectable suppressor cells. CONCLUSIONS: Suppressor cell deficiency is compatible with a role for rejection and/or trophoblast failure in some patients with recurrent miscarriage. Presence of suppressor cells in most patients with missed abortion (4/5) is compatible with an alternative cause of fetal death, similar to findings reported in genetic fetal death mice.

Abortion, Habitual↗
PubMed1994

TNF-alpha production by eosinophils in upper airways inflammation (nasal polyposis).

TNF-alpha is a cytokine with a wide spectrum of proinflammatory activities. Nasal polyps (NP), which occur in association with allergic rhinitis and asthma, are characterized by a marked infiltration of activated eosinophils, epithelial damage, and varying degrees of stromal fibrosis. By using Southern blot analysis after a reverse transcription-PCR, we detected a signal specific for TNF-alpha mRNA in five of seven NP samples, but not in control nasal mucosal samples. With in situ hybridization, we detected cells expressing mRNA specific for TNF-alpha in eight of thirteen NP samples, but not in four control samples. Counterstaining with chromotrope 2R demonstrated that virtually all cells expressing TNF-alpha message were eosinophils. The ratio of eosinophils expressing TNF-alpha to the total number of eosinophils varied greatly among tissues; as an average, we observed 24% TNF-alpha-positive eosinophils. Using a mAb against human TNF-alpha, we demonstrated TNF-alpha localization in 12 NP tissues (48.8 +/- 16.5 positive cells/mm2) but not in three control samples. Morphologically, cells localizing TNF-alpha were both mononucleated and polynucleated with only a small number of eosinophils, as determined by aniline blue counterstaining. Studies of purified blood eosinophils from a patient with hypereosinophilic syndrome and from four healthy donors indicated that TNF-alpha is produced, but rapidly secreted, so that TNF-alpha mRNA-positive cells contain little detectable protein. Furthermore, cells that retain detectable TNF-alpha may not contain sufficient TNF-alpha mRNA to be detected by using the probe developed for our studies. Together, these findings identify a novel mechanism by which eosinophils may contribute to mucosal inflammation and provide an approach to future investigation.

Eosinophilia↗
PubMed1994

Functional heterogeneity of mast cells isolated from different microenvironments within nasal polyp tissue.

Nasal polyposis is a chronic inflammatory condition of the upper airways characterized by infiltration of activated inflammatory cells, including mast cells, both in the epithelium and in the stroma. The aim of this work was to study human mast cells derived from two different anatomical sites within the same nasal polyp tissue. To this end, we isolated two distinct mast cell populations, one from the epithelial and the other from the stromal layers of individual human nasal polyp tissues. We examined the mediator content of the two mast cell populations and found that stromal mast cells had a significantly higher content of tryptase compared with the epithelial mast cells from the same tissue. In addition, mast cells from the stromal compartment, but not those from the epithelium, released a significant amount of histamine after anti-IgE stimulation. By contrast, both populations released over 50% of the total histamine after non-specific stimuli (A23187 10(-6) M). The content of mediators and the response to immunological activation were not significantly altered in patients receiving topical steroid therapy. It remains to be determined if the observed differences are the result of an intrinsic characteristic of the mast cell populations localized to separate tissue compartments, or reflect a different in vivo exposure to stimuli such as antigens, or different surrounding structural or infiltrating cells. In conclusion, these data provide evidence of functional heterogeneity and differences in mediator content between mast cell subpopulations from a single human tissue. The failure of release of epithelial mast cell mediators from an immunologic stimulus may have implications concerning acute effects of antigen exposure in nasal polyposis.

Adult↗
PubMed1993

Identification of epithelial cells in bronchoalveolar lavage.

1. Damage to the bronchial epithelium occurs after the inhalation of toxic substances and allergens, and through virus infections and it may lead to increased desquamation of epithelial cells in bronchoalveolar lavage (BAL). 2. In this study we compared two methods of staining the epithelial cells of BAL, the conventional cytochemical May Grunwald-Giemsa stain (MGG) and an immunocytochemical technique using a monoclonal antibody anti-human cytokeratin (CK) detected with APAAP immuno-alkaline phosphatase. BAL was obtained from 13 subjects and the epithelial cells were cytocentrifuged either immediately after collection (fraction A) or after washing (fraction B). 3. Higher percentages of epithelial cells were identified in fraction A with CK (20.0 +/- 5.1%) than in fraction A with MGG (11.2 +/- 2.3%), which recognized only ciliated epithelial cells. In fact a proportion of CK-positive cells (34%) in fraction A were not ciliated. Underestimation of epithelial cells by MGG compared to CK was more pronounced in fraction B (8.0 +/- 2.9% and 22.9 +/- 3.0%, respectively) as there was a relative loss of ciliated CK+ cells after washings. 4. These results suggest that immunocytochemical staining with an anti-cytokeratin monoclonal antibody is more sensitive than using the MGG stain in detecting epithelial cells in BAL.

Adolescent↗
PubMed1991

Increase in numbers of CD8 positive lymphocytes and eosinophils in peripheral blood of subjects with late asthmatic reactions induced by toluene diisocyanate.

Occupational asthma induced by toluene diisocyanate (TDI) shares several features with allergic asthma, but the mechanism of action of TDI is poorly understood. Ten sensitised subjects, previously shown to develop a dual or late asthmatic reaction after inhaling TDI were examined. In each subject, forced expiratory volume in one second (FEV1) was measured and venous blood was taken before, and 30 minutes and eight, 24, 48, and 72 hours after exposure to TDI (0.005-0.015 ppm for 10-30 minutes). Filtered air was used as a control. Differential leucocyte counts were determined and phenotypic analysis was performed by immunofluorescence on mononuclear cells using monoclonal antibodies (anti-CD3, anti-CD4, anti-CD8, and anti-HLA-DR). Five subjects developed a dual asthmatic reaction and five had a late reaction. Percentage of CD8 positive lymphocytes increased significantly eight hours after exposure to TDI (from 27 +/- 3 (SEM) % to 42.1 +/- 5%) in the subjects with an isolated late reaction. A delayed significant further increase in suppressor/cytotoxic T lymphocytes was seen in seven of the 10 subjects 48 hours after active exposure (from 27 +/- 2% to 42 +/- 4.8%), irrespective of the type of asthmatic reaction developed after exposure to TDI. Eosinophil percentage increased from 2.5% +/- 1.0 to 6.4% +/- 1.2 24 hours after exposure to TDI and the increase was sustained for up to 48 hours (4.7 +/- 1.1%). No significant variations of FEV1 or cell percentages were seen in the controls. In conclusion, the events triggered by exposure to TDI in sensitised subjects included changes in lung function and systemic effects which lasted longer than bonchoconstriction and concerned suppressor/cytotoxic lymphocytes and eosinophils. These results suggest that TDI induced late asthmatic reactions may be associated with an immunological response to TDI or to its products.

Adult↗
PubMed1991

Granulocyte/macrophage colony-stimulating factor (GM-CSF) gene expression by eosinophils in nasal polyposis.

Nasal polyposis is a chronic inflammatory condition of the upper airways characterized by infiltration of activated inflammatory cells, particularly eosinophils. Granulocyte/macrophage colony-stimulating factor (GM-CSF) is a cytokine with powerful biologic effects including the regulation of survival, proliferation, and activation of granulocytes as well as differentiation of hemopoietic cells. To examine the potential role of GM-CSF in the pathogenesis of this condition, we investigated gene expression and production of GM-CSF in nasal polyp tissues as well as in the normal nasal mucosa. Immunoreactive GM-CSF was detected by enzyme-linked immunosorbent assay in the 24-h supernatant of nasal polyp tissues placed in culture. By Northern blot analysis and Southern blot analysis following a reverse-transcription polymerase chain reaction using a human GM-CSF cDNA probe, we detected GM-CSF mRNA in nasal polyp tissues, as well as in the tissue from a patient with allergic rhinitis, but not in the normal nasal mucosa. By in situ hybridization using the same probe, cells expressing mRNA specific for GM-CSF were observed in nasal polyp tissues and in the allergic nasal mucosa. In addition, by the combination of in situ hybridization and counterstaining with chromotrope 2R, we demonstrated that approximately 30% of eosinophils infiltrating the polyp tissue express the GM-CSF gene. These results suggest a novel mechanism by which eosinophils may contribute to the pathogenesis of chronic inflammatory diseases such as nasal polyposis, allergic rhinitis, and, by implication, asthma.

Base Sequence↗
PubMed1989

RBE-LET relationship for the survival of V79 cells irradiated with low energy protons.

The survival of V79 Chinese hamster cells irradiated with proton beams with energies of 0.73, 0.84, 1.16, 1.70 and 3.36 MeV, corresponding to LET values, evaluated at the cell midplane, of 34.5, 30.4, 23.9, 17.8 and 10.6 keV/micron respectively, have been studied in the dose range 0.5-6.0 Gy. As a reference, the survival curve obtained with 200 kV X-rays was used. The initial shoulder, typical of survival curves obtained with sparsely ionizing radiation, decreases as the LET increases and completely disappears at 23.9 keV/micron. This value corresponds to the maximum of the RBE, expressed as the initial slope ratio. In the energy range we have considered, the RBEs for protons are higher than those reported for other ions of comparable LET and the RBE-LET relationship results shifted to lower LET values. Our data seem to indicate that the RBE-LET curve depends on the type of radiation and this could imply that LET is not a good reference for the dose-effectiveness relationship.

Animals↗
PubMed1989

Ambroxol inhibits airway hyperresponsiveness induced by ozone in dogs.

To follow up previous observations that airway hyperresponsiveness induced by ozone is linked to airway inflammation and particularly to the release of arachidonic acid metabolites, we investigated the effect of ambroxol (a mucoactive and surfactant-stimulating drug that has recently been discovered to inhibit the release of arachidonic acid from cell membrane phospholipids) on airway hyperresponsiveness and bronchoalveolar neutrophilia induced by ozone in dogs. One group of 5 dogs was studied before treatment with nebulized saline and then after exposure to ozone (3 ppm, 1 h); another group of 6 dogs was studied before treatment with ambroxol (100 breaths of a 1% solution) and after exposure to ozone. On each occasion, we measured airway responsiveness to acetylcholine and counted the number of cells in bronchoalveolar lavage fluid. When the dogs were given the saline placebo, ozone induced a marked increase in airway responsiveness to acetylcholine and a marked influx of neutrophils in the airways. When the dogs were given ambroxol, ozone induced the same increase in the number of neutrophils in bronchoalveolar lavage, but did not increase the degree of airway responsiveness to acetylcholine. We conclude that ambroxol inhibits ozone-induced airway hyperresponsiveness in dogs, probably by inhibiting the formation and release of oxygenation products of arachidonic acid from neutrophils.

Acetylcholine↗
PubMed1987

The micronucleus assay in exfoliated cells of the human buccal mucosa.

The frequency of micronuclei resulting from chromosome breaks, and that of micronuclei deriving from spindle disturbances was determined in exfoliated cells of the human buccal mucosa in 50 normal individuals. Several confounding factors, such as age, smoking habits, alcohol consumption, etc., were taken into account. While the frequencies of micronuclei resulting from chromosome breaks and of cells with this kind of micronuclei were about double in smokers as compared with non-smokers, the difference being highly statistically significant, the frequencies of cells with spindle disturbances were almost the same in the two groups. No statistically significant correlation was found for any of the other variables examined. In two patients suffering from cancer of the oral cavity the variation of the frequency of the micronuclei during the progress of radiotherapy and following its interruption was determined. It was found that gamma rays induced only micronuclei resulting from chromosome breaks, whose frequency increased linearly with the applied dose, and was lowered to the initial background level 7-12 days after the end of radiotherapy.

Adult↗