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Biomedical subjects

S Gasperini

Publications and source records attributed to S Gasperini.

At least 19 recordsLinked to original sources
PubMed2006

Hyperkalemia after acute metabolic decompensation in two children with vitamin B12-unresponsive methylmalonic acidemia and normal renal function.

The patients affected by vitamin B12-unresponsive methylmalonic acidemia (MMA) on the long run develop chronic renal disease with interstitial nephropathy and progressive renal insufficiency. The mechanism of nephrotoxicity in vitamin B12-unresponsive MMA is not yet known. Chronic hyporeninemic hypoaldosteronism has been found in many cases of methylmalonic acidemia, hyperkalemia and renal tubular acidosis type 4. We report 2 patients affected by B12-unresponsive methylmalonic acidemia diagnosed at the age of 23 months and 5 years, respectively, with normal glomerular filtration and function. They showed hyporeninemic hypoaldosteronism and significant hyperkalemia requiring sodium potassium exchange resin (Kayexalate) therapy after an episode of metabolic decompensation leading to diagnosis of MMA. In both children, hyporeninemic hypoaldosteronism and hyperkalemia disappeared after 6 months of good metabolic control.

Child, Preschool↗
PubMed2002

Two novel genetic lesions and a common BH4-responsive mutation of the PAH gene in Italian patients with hyperphenylalaninemia.

Hyperphenylalaninemia (HPA), due to a deficiency of phenylalanine hydroxylase (PAH) enzyme, is caused by mutations in the PAH gene. Molecular analysis in 23 Italian patients with PAH deficiency identified two novel (P281R, L287V) and 20 previously described genetic lesions in the PAH gene. The detection of the A403V amino acid substitution in combination with null mutations in patients with BH4-responsive PAH deficiency leads us to correlate it with BH4 responsiveness.

DNA Mutational Analysis↗
PubMed2002

Two new mutations in children affected by partial biotinidase deficiency ascertained by newborn screening.

Mutation analysis performed on DNA from 6 Italian patients with partial biotinidase deficiency ascertained by newborn screening allowed the identification of two new mutations, c1211C > T (T404I) and a single base deletion c594delC. All patients were compound heterozygous for the D444H amino acid substitution showing that this mutation is also common in Italian patients affected by partial biotinidase deficiency.

Amidohydrolases↗
PubMed2001

Up-regulation of IL-10R1 expression is required to render human neutrophils fully responsive to IL-10.

We have recently shown that IL-10 fails to trigger Stat3 and Stat1 tyrosine phosphorylation in freshly isolated human neutrophils. In this study, we report that IL-10 can nonetheless induce Stat3 tyrosine phosphorylation and the binding of Stat1 and Stat3 to the IFN-gamma response region or the high-affinity synthetic derivative of the c-sis-inducible element in neutrophils that have been cultured for at least 3 h with LPS. Similarly, the ability of IL-10 to up-regulate suppressor of cytokine signaling (SOCS)-3 mRNA was dramatically enhanced in cultured neutrophils and, as a result, translated into the SOCS-3 protein. Since neutrophils' acquisition of responsiveness to IL-10 required de novo protein synthesis, we assessed whether expression of IL-10R1 or IL-10R2 was modulated in cultured neutrophils. We detected constitutive IL-10R1 mRNA and protein expression in circulating neutrophils, at levels which were much lower than those observed in autologous monocytes or lymphocytes. In contrast, IL-10R2 expression was comparable in both cell types. However, IL-10R1 (but not IL-10R2) mRNA and protein expression was substantially increased in neutrophils stimulated by LPS. The ability of IL-10 to activate Stat3 tyrosine phosphorylation and SOCS-3 synthesis and to regulate IL-1 receptor antagonist and macrophage-inflammatory protein 1beta release in LPS-treated neutrophils correlated with this increased IL-10R1 expression, and was abolished by neutralizing anti-IL-10R1 and anti-IL-10R2 Abs. Our results demonstrate that the capacity of neutrophils to respond to IL-10, as assessed by Stat3 tyrosine phosphorylation, SOCS-3 expression, and modulation of cytokine production, is very dependent on the level of expression of IL-10R1.

Cell Separation↗
PubMed2000

The neutrophil as a cellular source of chemokines.

Neutrophils are known to play an important role in inflammatory responses by virtue of their ability to perform a series of effector functions that collectively represent a major mechanism of innate immunity against injury and infection. In recent years, however, it has become obvious that the contribution of neutrophils to host defence and natural immunity extends well beyond their traditional role as professional phagocytes. Indeed, neutrophils can be induced to express a number of genes whose products lie at the core of inflammatory and immune responses. These include not only Fc receptors, complement components, cationic antimicrobial and NADPH oxidase proteins, but also a variety of cytokines (including tumour necrosis factor-alpha, interleukin (IL)-1beta, IL-1R alpha, IL-12 and vascular endothelial growth factor), and chemokines such as IL-8, growth-related gene product, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, interferon-gamma-inducible protein of 10 kDa and monokine induced by interferon-gamma. Because these chemokines are primarily chemotactic for neutrophils, monocytes, immature dendritic cells and T-lymphocyte subsets, a potential role for neutrophils in orchestrating the sequential recruitment of distinct leukocyte types to the inflamed tissue is likely to occur. The purpose of this review is to summarize the essential features of the production of chemokines by polymorphonuclear neutrophil leukocytes and the contribution that we have made to characterize some aspects of this newly discovered crucial function of neutrophils.

Animals↗
PubMed1999

Interleukin-10 (IL-10) selectively enhances CIS3/SOCS3 mRNA expression in human neutrophils: evidence for an IL-10-induced pathway that is independent of STAT protein activation.

We have recently shown that, in human neutrophils, interleukin-10 (IL-10) fails to induce specific DNA-binding activities to the gamma-interferon response region (GRR), a regulatory element located in the FcgammaRI gene promoter, which is required for transcriptional activation by IL-10 and interferon gamma (IFNgamma) in monocytic cells. In this study, we report that IL-10 is also unable to induce the binding of STAT1 or STAT3 to the serum-inducible element (hSIE/m67), despite the fact that both proteins are expressed in neutrophils. Whereas IFNgamma and granulocyte colony-stimulating factor (G-CSF) are efficient inducers of STAT1 and STAT3 tyrosine phosphorylation in polymorphonuclear neutrophils (PMN), IL-10 fails to trigger STAT1 and STAT3 tyrosine and serine phosphorylation, therefore explaining its inability to induce the FcgammaRI expression in these cells. By contrast, we demonstrate that IL-10 alone represents an efficient stimulus of CIS3/SOCS3 mRNA expression in neutrophils. CIS3/SOCS3 belongs to the recently cloned cytokine-inducible SH2-containing protein (CIS) gene family (which also includes CIS1, CIS2, CIS4, CIS5, and JAB) that is believed to be, at least in part, under the control of STAT transcription factors and whose products are potential modulators of cytokine signaling. Moreover, IL-10 synergizes with lipopolysaccharide (LPS) in upregulating CIS3/SOCS3 mRNA expression in PMN through a mechanism that involves mRNA stabilization. In contrast to CIS3/SOCS3, mRNA transcripts encoding other family members are unaffected by IL-10 in neutrophils. Finally, transfection of CIS3/SOCS3 in murine M1 myeloid cells suppresses LPS-induced growth arrest, macrophage-like differentiation, and nitric oxide synthesis, but not IL-6 mRNA expression. Collectively, our data suggest that, in neutrophils, the activation of STAT1 and STAT3 phosphorylation is neither required for CIS3/SOCS3 induction by IL-10 nor involved in the regulatory effects of IL-10 on cytokine production.

Animals↗
PubMed1999

Gene expression and production of the monokine induced by IFN-gamma (MIG), IFN-inducible T cell alpha chemoattractant (I-TAC), and IFN-gamma-inducible protein-10 (IP-10) chemokines by human neutrophils.

Monokine induced by IFN-gamma (MIG), IFN-inducible T cell alpha chemoattractant (I-TAC), and IFN-gamma-inducible protein of 10 kDa (IP-10) are related members of the CXC chemokine subfamily that bind to a common receptor, CXCR3, and that are produced by different cell types in response to IFN-gamma. We have recently reported that human polymorphonuclear neutrophils (PMN) have the capacity to release IP-10. Herein, we show that PMN also have the ability to produce MIG and to express I-TAC mRNA in response to IFN-gamma in combination with either TNF-alpha or LPS. While IFN-gamma, alone or in association with agonists such as fMLP, IL-8, granulocyte (G)-CSF and granulocyte-macrophage (GM)-CSF, failed to influence MIG, IP-10, and I-TAC gene expression, IFN-alpha, in combination with TNF-alpha, LPS, or IL-1beta, resulted in a considerable induction of IP-10 release by neutrophils. Furthermore, IL-10 and IL-4 significantly suppressed the expression of MIG, IP-10, and I-TAC mRNA and the extracellular production of MIG and IP-10 in neutrophils stimulated with IFN-gamma plus either LPS or TNF-alpha. Finally, supernatants harvested from stimulated PMN induced migration and rapid integrin-dependent adhesion of CXCR3-expressing lymphocytes; these activities were significantly reduced by neutralizing anti-MIG and anti-IP-10 Abs, suggesting that they were mediated by MIG and IP-10 present in the supernatants. Since MIG, IP-10, and I-TAC are potent chemoattractants for NK cells and Th1 lymphocytes, the ability of neutrophils to produce these chemokines might contribute not only to the progression and evolution of the inflammatory response, but also to the regulation of the immune response.

Apoptosis↗
PubMed1998

Involvement of the IP-10 chemokine in sarcoid granulomatous reactions.

The accumulation of T cells and monocytes at sites of ongoing inflammation represents the earliest step in the series of events that lead to granuloma formation in sarcoidosis. In this study, we evaluated the pulmonary production of IFN-inducible protein 10 (IP-10), a CXC chemokine that stimulates the directional migration of activated T cells. Striking levels of IP-10 were demonstrated in the bronchoalveolar lavage (BAL) fluid of 24 patients with pulmonary sarcoidosis and lymphocytic alveolitis, as compared with patients with inactive disease or control subjects. A positive correlation was demonstrated between IP-10 levels and the number of sarcoid CD45R0+/CD4+ cells in the BAL. Immunochemistry, performed with an anti-human IP-10 polyclonal Ab in lymph nodes displaying prominent sarcoid granulomas, showed that cells bearing IP-10 were mainly epithelioid cells and CD68+ macrophages located inside granulomatous areas. Macrophages recovered from the BAL of sarcoid patients stained positive for IP-10 protein. Furthermore, alveolar macrophages isolated from sarcoid patients with T cell alveolitis and cultured for 24 h in presence of IFN-gamma secreted definite levels of IP-10 capable of inducing T cell chemiotaxis. Interestingly, alveolar lymphocytes recovered from patients with active sarcoidosis were CD4+ T cells expressing Th1 cytokines (IL-2 and IFN-gamma) and high levels of CXCR3. Taken together, these data suggest the potential role of IP-10 in regulating the migration and activation of T cells toward sites of sarcoid inflammatory process and the consequent granuloma formation.

Adult↗
PubMed1998

Impaired cytokine production by neutrophils isolated from patients with AIDS.

OBJECTIVES: To determine the ability of neutrophils isolated from HIV-seropositive patients to produce proinflammatory cytokines. DESIGN: The in vitro responsiveness of polymorphonuclear neutrophils (PMN) and peripheral blood mononuclear cells (PBMC) to lipopolysaccharide (LPS), used in the presence or absence of interferon (IFN)gamma, was determined in 47 HIV-positive patients with advanced stages of virus infection. METHODS: Cytokines in cell-free supernatants were measured by enzyme-linked immunosorbent assay or radioimmunoassay. RESULTS: Cell-free supernatants from PMN isolated from the peripheral blood of HIV-positive patients and stimulated with LPS contained increased amounts of tumour necrosis factor (TNF)-alpha and interleukin (IL)-8 with respect to supernatants obtained from PMN of normal donors. In contrast, release of IL-1beta and IL-1ra (IL-1 receptor antagonist) in response to LPS, or LPS plus IFNgamma, was found to be lower in PMN from HIV-positive patients than in PMN from controls, but was significant only in the case of IL-1ra. Furthermore, the release of IL-12 induced by LPS or LPS plus IFNgamma did not significantly differ between PMN from HIV-positive patients and healthy donors. Concerning PBMC, the production of TNF-alpha and IL-12 in response to LPS, or LPS plus IFNgamma, was found to be significantly higher in cells isolated from HIV-positive patients, whereas the release of IL-1beta was significantly lower. In the case of IL-8, no statistically significant difference was found between PBMC isolated from HIV-positive patients and healthy donors. CONCLUSIONS: Collectively, the data suggest that in HIV-positive patients with advanced stages of disease, the ability of PMN to produce specific cytokines in response to LPS is significantly altered. Alterations in this ability might contribute to the evolution of HIV disease.

Adolescent↗
PubMed1998

Modulation by interferon-gamma of the production and gene expression of IL-1 receptor antagonist in human neutrophils.

In this report, we show that interferon-gamma (IFN-gamma) modulates the production of IL-1ra in activated human neutrophils. In lipopolysaccharide-stimulated cells, IFN-gamma increased the release of IL-1ra without modulating IL-1ra mRNA accumulation; under these conditions, IFN-gamma only marginally enhanced IL-1ra de novo synthesis, while IL-1ra was more efficiently secreted. In response to the formylated peptide, fMLP, neutrophils released small but significant amounts of IL-1ra, yet without an increase in IL-1ra mRNA over constitutive levels. Following IFN-gamma treatment, however, the fMLP-elicited IL-1ra production was greatly potentiated, and this was accompanied by a transient increased accumulation of IL-1ra mRNA. Finally, opsonized yeast particles were found to induce IL-1ra formation at late incubation times, and prior treatment of neutrophils with IFN-gamma moderately enhanced this response. Collectively, our results demonstrate that in neutrophils activated by different classes of agonists, IFN-gamma modulates the release of IL-1ra by acting through distinct mechanisms.

Cells, Cultured↗
PubMed1998

High affinity receptor for IgG (Fc gamma RI/CD64) gene and STAT protein binding to the IFN-gamma response region (GRR) are regulated differentially in human neutrophils and monocytes by IL-10.

Since IL-10 has been shown to up-regulate the expression of the high affinity receptor for IgG (FcgammaRI/CD64) in human monocytes, we examined whether the cytokine exerts a similar action toward polymorphonuclear neutrophils (PMN). Unexpectedly, we found that in neutrophils, IL-10 failed to induce either the mRNA accumulation or the surface expression of FcgammaRI. Consistent with these findings, stimulation of PMN with IFN-gamma, but not with IL-10, resulted in the induction of specific DNA-binding activities to the IFN-gamma response region (GRR), a regulatory element located in the FcgammaRI gene promoter, required for transcriptional activation. In electrophoretic mobility shift assays (EMSAs), we confirmed that in PBMC, IL-10 induces the binding to the GRR of both STAT1 and STAT3, two members of the STAT family. In neutrophils, however, these activators did not bind to the GRR in response to IL-10, despite the fact that both STAT1 and STAT3 are expressed in these cells. On the other hand, IFN-gamma was an efficient inducer of STAT1 binding to the GRR in both PMN and PBMC. The lack of inducible GRR-binding activity in IL-10-treated PMN could not be ascribed to a lack of IL-10R, and did not appear to reflect an inhibitory effect of the cytokine. Taken together, our data suggest that IL-10 is unable to induce FcgammaRI gene expression in neutrophils because the intracellular signaling pathway triggered by the cytokine is impaired at the level of, or upstream of, STAT1 and/or STAT3 activation.

DNA-Binding Proteins↗
PubMed1997

Regulated production of the interferon-gamma-inducible protein-10 (IP-10) chemokine by human neutrophils.

Interferon-gamma (IFN-gamma)-inducible protein-10 (IP-10), a member of the C-X-C sub-family of chemokines, is known to be produced by monocytes, lymphocytes, keratinocytes and endothelial cells in response to IFN-gamma. Here, we show that human polymorphonuclear neutrophils (PMN) also have the ability to produce IP-10. IFN-gamma alone had a modest effect on IP-10 mRNA accumulation, whereas tumor necrosis factor-alpha (TNF-alpha), yeast particles opsonized with IgG (Y-IgG), lipopolysaccharide (LPS), and formyl-methionyl-leucyl-phenylalanine (fMLP) all failed to up-regulate IP-10 gene expression. However, stimulation of neutrophils with IFN-gamma in combination with either TNF-alpha or LPS (but not with Y-IgG or fMLP) resulted in a considerable induction of IP-10 mRNA transcripts, as well as in the extracellular release of the protein. In contrast, the best inducer of IP-10 release from peripheral blood mononuclear cells was IFN-gamma alone. Furthermore, mRNA stabilization analyses demonstrated that IP-10 mRNA isolated from PMN stimulated with IFN-gamma only, or with IFN-gamma plus either TNF-alpha or LPS, had similar half-lives. Finally, we found that interleukin-10, a known inhibitor of chemokine production in PMN, moderately suppressed the extracellular production of IP-10 in neutrophils stimulated with IFN-gamma plus either LPS or TNF-alpha. Since IP-10 is a potent chemoattractant for activated T lymphocytes, the ability of neutrophils to produce IP-10 might contribute to the evolution and progression of the inflammatory response.

Cells, Cultured↗
PubMed1996

Granulocyte colony-stimulating factor induces the binding of STAT1 and STAT3 to the IFNgamma response region within the promoter of the Fc(gamma)RI/CD64 gene in human neutrophils.

Granulocyte colony-stimulating factor (G-CSF) has been recently shown to induce the high-affinity Fc receptor for IgG (Fc(gammaRI)/CD64) in human polymorphonuclear neutrophils (PMN). To elucidate the molecular mechanisms whereby G-CSF exerts this effect, we examined whether the cytokine induces the binding of transcription factors to the IFNgamma response region (GRR), a well characterized regulatory element in the Fc(gammaRI) promoter that is responsible for the transcriptional induction of this gene. Using electrophoretic mobility shift assays, we show that in human PMN, G-CSF activates a GRR-binding complex which contains members of the signal transducer and activator of transcription (STAT) family of proteins, namely STAT1 and STAT3. In keeping with this result, treatment of neutrophils with G-CSF led to tyrosine phosphorylation of STAT3, as determined by immunoprecipitation followed by immunoblotting with antiphosphotyrosine antibodies. This is the first demonstration that in human neutrophils, the induction by G-CSF of Fc(gammaRI) gene expression may be mediated by the binding of STAT1 and STAT3 to the GRR sequence.

Binding Sites↗
PubMed1995

Lipopolysaccharide-induced interleukin-8 gene expression in human granulocytes: transcriptional inhibition by interferon-gamma.

We recently showed that lipopolysaccharide (LPS) is a potent inducer of interleukin-8 (IL-8) expression in human polymorphonuclear leucocytes (PMN), at the level of both mRNA and protein, and that interferon-gamma (IFN gamma) inhibits IL-8 mRNA accumulation in stimulated PMN. To further define the molecular basis of the regulation of IL-8 gene expression in PMN, we investigated the effects of LPS and IFN gamma at both the transcriptional and post-transcriptional levels. As determined by Northern blot analysis, new protein synthesis was not required for the induction of IL-8 mRNA expression by LPS. Neither did the half-life of IL-8 mRNA in LPS-treated PMN differ from that observed in untreated cells. However, nuclear run-on analysis revealed that LPS increased the transcription of the IL-8 and IL-1 beta genes and that, in LPS-activated cells, IFN gamma markedly inhibited the rate of IL-8 gene transcription, but not that of IL-1 beta. IFN gamma did not affect IL-8 mRNA stability in LPS-treated PMN, indicating that the cytokine does not regulate LPS-induced IL-8 gene expression through post-transcriptional events. These results provide the first evidence that human granulocytes can actively transcribe the IL-8 gene, and that transcriptional inhibition is the mechanism by which IFN gamma inhibits IL-8 gene expression in PMN.

Cell Nucleus↗
PubMed1995

Interleukin-10 decreases tyrosine phosphorylation of discrete lipopolysaccharide-induced phosphoproteins in human granulocytes.

Although Interleukin-10 (IL-10) has been recently shown to modulate lipopolysaccharide (LPS)-induced release of cytokines in human granulocytes, the intracellular signalling pathways of LPS have been only partially defined, while those of IL-10 remain unknown. The present study shows that LPS induces an increase in tyrosine phosphorylation of a discrete number of proteins, in a time- and concentration-dependent manner. In addition, IL-10 negatively influenced protein tyrosine phosphorylation in LPS-treated human polymorphonuclear leukocytes (PMN). The effect of IL-10 was evident only after 60 min LPS-stimulation and was detected by analysing either cell lysates or lysates which were previously immunoprecipitated with anti-phosphotyrosine antibodies. Amongst the tyrosine phosphoproteins mostly affected by IL-10 in LPS-stimulated cells were the species with molecular weights ranging from 46 to 49 kDa. The identity and possible function of these proteins remain unknown. Taken together, our results suggest that tyrosine phosphorylation may constitute one of the intracellular events that mediate LPS and IL-10 responses in granulocytes.

Granulocytes↗
PubMed1995

Interleukin-12 production by human polymorphonuclear leukocytes.

Human polymorphonuclear leukocytes (PMN) stimulated by lipopolysaccharide (LPS) produce interleukin-12 (IL-12). Both the free IL-12 p40 chain and minute amounts of the biologically active IL-12 p70 heterodimers are produced by PMN. Interferon-gamma (IFN-gamma) enhanced the LPS-induced secretion of both the free IL-12 p40 chain and the p70 heterodimer by approximately fivefold. As observed for other IL-12-producing cell types, the ratio of free p40 chain to p70 heterodimer secreted by LPS-stimulated PMN was approximately 20:1. LPS induced a 100-fold increase of IL-12 p40 mRNA, but had minimal effect on p35 mRNA accumulation. IFN-gamma enhanced the LPS-induced accumulation of p40 mRNA and directly induced a several-fold increase in the accumulation of p35 mRNA. Therefore, the combined effect of LPS and IFN-gamma induced sufficient expression of both p40 and p35 to attain production of the biologically active p70 heterodimer at physiologically relevant concentrations. The ratio between p40 and p35 mRNA abundance in PMN stimulated with both LPS and IFN-gamma was approximately 200:1, explaining the secretion of the free p40 chain in much higher concentrations than the p70 heterodimer. IL-10, an inhibitor of the production of various cytokines in PMN, also suppressed IL-12 mRNA accumulation and secretion by PMN. Because of the important immunoregulatory function of IL-12, in particular induction of IFN-gamma production and facilitation of T helper cell type 1 response, the ability of PMN to produce IL-12 suggests that neutrophils may play an active role in the regulatory interaction between innate resistance and adaptive immunity.

Blotting, Northern↗