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A Vercellone

Publications and source records attributed to A Vercellone.

At least 19 recordsLinked to original sources
PubMed2000

New structural insights into the molecular deciphering of mycobacterial lipoglycan binding to C-type lectins: lipoarabinomannan glycoform characterization and quantification by capillary electrophoresis at the subnanomole level.

Lipoarabinomannans are key molecules of the mycobacterial envelopes involved in many steps of tuberculosis immunopathogenesis. Several of the biological activities of lipoarabinomannans are mediated by their ability to bind human C-type lectins, such as the macrophage mannose receptor, the mannose-binding protein and the surfactant proteins A and D. The lipoarabinomannan mannooligosaccharide caps have been demonstrated to be involved in the binding to the lectin carbohydrate recognition domains. We report an original analytical approach, based on capillary electrophoresis monitored by laser-induced fluorescence, allowing the absolute quantification, in nanomole quantities of lipoarabinomannan, of the number of mannooligosaccharide units per lipoarabinomannan molecule. Moreover, this analytical approach was successful for the glycosidic linkage determination of the mannooligosaccharide motifs and has been applied to the comparative analysis of parietal and cellular lipoarabinomannans of Mycobacterium bovis BCG and Mycobacterium tuberculosis H37Rv, H37Ra and Erdman strains. Significant differences were observed in the amounts of the various mannooligosaccharide units between lipoarabinomannans of different strains and between parietal and cellular lipoarabinomannans of the same strain. Nevertheless, no relationship was found between the number of mannooligosaccharide caps and the virulence of the corresponding strain. The results of the present study should help us to gain more understanding of the molecular basis of lipoarabinomannan discrimination in the process of binding to C-type lectins.

Acetylation↗
PubMed2000

Mycobacterium tuberculosis H37Rv parietal and cellular lipoarabinomannans. Characterization of the acyl- and glyco-forms.

Mannosylated lipoarabinomannans are multifaceted molecules. They have been shown to exert an immunosuppressive role in the immunopathogenesis of tuberculosis. They are also described as antigens of host double negative alphabeta T-cells. Delimitation of ManLAMs epitopes require knowledge of the precise structure of these molecules. The two major functional domains (the cap motifs and the phosphatidylinositol anchor) of the parietal and cellular ManLAMs of Mycobacterium tuberculosis H37Rv were investigated here. Using capillary electrophoresis, we established that parietal and cellular ManLAMs share the same capping motifs, mono-, di-, and trimannosyl units with the same relative abundance. By (31)P NMR analysis of the native LAMs in Me(2)SO-d(6), the major acyl-form of both parietal and cellular H37Rv ManLAM anchors, typified by the P3 phosphorus resonance, comprised a diacylglycerol unit. Three other acyl-forms were characterized in the cellular ManLAMs. Comparative analysis of the cellular Mycobacterium bovis BCG and M. tuberculosis ManLAM acyl-forms revealed the presence of the same populations, but with different relative abundance. The biological importance of the H37Rv ManLAM acyl-form characterization is discussed, particularly concerning the molecular mechanisms of binding of ManLAMs to the CD1 proteins involved in the presentation of ManLAMs to T-cell receptors.

Acylation↗
PubMed1999

Structural definition of arabinomannans from Mycobacterium bovis BCG.

The structures of the hydrophilic parietal and cellular arabinomannans isolated from Mycobacterium bovis BCG cell wall [Nigou et al. (1997) J Biol Chem 272: 23094-103] were investigated. Their molecular mass as determined by MALDI-TOF mass spectrometry was around 16 kDa. Concerning cap structure, capillary electrophoresis analysis demonstrated that dimannoside (Manpalpha1-->2Manp) was the most abundant motif (65-75%). Using two-dimensional 1H-13C NMR spectroscopy, the mannan core was unambiguously demonstrated to be composed of -->6Manpalpha1--> backbone substituted at some O-2 by a single Manp unit. The branching degree was determined as 84%. Finally, arabinomannans were found to be devoid of the phosphatidyl-myo-inositol anchor and, by aminonaphthalene disulfonate tagging, the mannan core was shown to contain a reducing end. This constitutes the main difference between arabinomannans and lipoarabinomannans from Mycobacterium bovis BCG.

Carbohydrate Sequence↗
PubMed1998

Relationships between the structure and the roles of lipoarabinomannans and related glycoconjugates in tuberculosis pathogenesis.

The mechanisms and the molecular basis of the mycobacteria virulence remain obscure. However, recent findings provide evidences that, among the glycoconjugates which compose the mycobacterial envelope, lipoglycans including lipoarabinomannan, lipomannan and phosphatidyl-myo-inositol mannosides, are involved in the major steps of the tuberculosis immunopathogenesis. These steps are the mycobacterial phagocytosis process and the macrophage activation via the regulation of the production and secretion of cytokines. In this article, we examine recent observations about comparative structural models of the lipoglycans from the pathogenic Mycobacterium tuberculosis strain, and the vaccinal M. bovis BCG strain, and finally avirulent mycobacteria strains. We also consider the role of the lipoglycans in the mycobacterial phagocytosis process and in the regulation of the macrophage microbicidal activity.

Antigens, Bacterial↗
PubMed1997

Comparative structural study of the mannosylated-lipoarabinomannans from Mycobacterium bovis BCG vaccine strains: characterization and localization of succinates.

In order to investigate the molecular basis of the differences observed in the immunogenicity of various BCG vaccines, which may play a major role in BCG vaccination efficiency, the presence and the partial structure of the lipoarabinomannans (LAMs) were investigated in four different BCG strains (Pasteur, Glaxo, Copenhagen, and Japanese strains). According to their cell-wall extraction mode, the LAMs were subdivided into two classes, namely, parietal LAMs and cellular LAMs. From a structural point of view, the cellular LAMs and parietal LAMs both belong to the ManLAM class since the relevant structural features, described for ManLAMs, that is, the phosphatidyl-myo-Ins anchor and the manno-oligosaccharide caps were characterized. In addition, thanks to NMR experiments realized on the native ManLAMs, we prove the existence of succinyl groups and we determine their location on the C-2 of the 3,5-di-O-alpha-D-Araf units. The average number of succinyl groups was estimated at one and four per ManLAMs molecule for Glaxo and Japanese vaccines, respectively, and two for Pasteur and Copenhagen strains.

BCG Vaccine↗
PubMed1996

Complete NMR structural elucidation of the capsular polysaccharide adjuvant from Klebsiella I-714.

The capsular polysaccharide (CPS) produced by the non-pathogenic Klebsiella strain I-714, selected for its immunomodulating activity, has been purified and almost completely detoxified in a previous study [Adam, O., Vercellone, A., Paul, F., Monsan, P. F. & Puzo, G. (1995) Anal. Biochem. 225, 321-327]. The present report concerns the structural elucidation of this CPS by several one-dimensional and two-dimensional 1H-NMR and 13C-NMR experiments performed on the native molecule. It was found to be a high molecular-mass branched polymer constituted by a hexasaccharide repeating unit of following structure: 4)-alpha-L-Rhap-(1-->3)-beta-D-Galp-(1-->2)-alpha-L-Rhap-(1- ->4)-beta-D- GlcpA-(1-->3)-¿alpha-L-Rhap-(1-->2)-¿-alpha-D-Galp-(1. The presence of two glycosidic substitutions on the Galp residue resulted in a strong overlapping of its proton signals, preventing direct assignment of both proton and carbon resonances. However, assignment could be achieved using the two-bond and three-bond 1H-13C heteronuclear coupling observed in the 1H-13C heteronuclear multiple-bond correlation (HMBC) spectrum.

Adjuvants, Immunologic↗
PubMed1996

Incidence of biopsy-proven primary glomerulonephritis in an Italian province.

Between January 1, 1970, and December 31, 1994, 1,926 cases of biopsy-proven primary glomerulonephritis (PGN) were diagnosed in an adult population (> 15 years of age) in a northwestern region of Italy with approximately 3.5 million inhabitants. The principal long-term changes were an increase in the absolute number of biopsies per year, an increase in the mean age of patients undergoing biopsy (from 29.3 +/- 12.2 years to 47.0 +/- 17.8 years), an increase in the percentage of patients older than 65 years (from 1.7% to 20.4%), and an increase in the percentage of isolated urinary abnormalities as an indication for biopsy (from 3.5% to 29.6%). In the total biopsy material, immunoglobulin A glomerulonephritis (IgA-GN) is the most frequent type (26%), followed by membranous glomerulonephritis (MGN; 20%). An incidence study was begun in 1990; this survey was restricted to the population of the province of Torino (approximately 2 million inhabitants) as only this area completely refers to the nephrologic centers that entered patients into this study. The overall incidence of PGN is 4.68 new cases/yr/10(5) population with a predominance of males (> 2:1); IgA-GN is the most common type (1.47/yr/10(5) population [34.5%]) in the overall population. In the elderly, cases of PGN are twice as high as in adults (8.19/yr/10(5) population v 4.02/yr/10(5) population in the 65 to 74 year and 45 to 54 year age groups, respectively); MGN mainly accounts for this high incidence (3.4/yr/10(5) population), while the nephrotic syndrome is the most common indication for biopsy (53.8%). A comparison with the incidence in the same area in the early 1970s is evaluable only for PGN, which was mainly registered in the age groups for which an unrestricted biopsy policy was already in place (15 to 35 years). In contrast with a misleading increase of all types of PGN, which is in reality due to the extension of the biopsy policy to older and asymptomatic patients, membranoproliferative glomerulonephritis type I shows a countercurrent decrease from 0.43 to 0.13/yr/10(5) population. Evidence of a simultaneous decrease in severe cardiac valvulopathy, due to rheumatic fever, is also provided. We feel that before epidemiologic conclusions can be reached, a clear understanding of one's own biopsy policy is essential. An apparent change in the PGN rate in our region over the last 25 years mainly depends on modifications in our biopsy policy, most probably coupled with a change in the threshold of detection of symptoms in the general population. At present, according to our experience, IgA-GN is the most common type of PGN in the total bioptic material, as demonstrated in other European countries, while the elderly show a peculiar pattern with a higher PGN incidence, mainly represented by MGN and heralded by the nephrotic syndrome. We also confirm that membranoproliferative glomerulonephritis type I is indeed decreasing in parallel with changes in the microbiologic environment.

Adolescent↗
PubMed1996

Cytometric detection of mycobacterial surface antigens: exposure of mannosyl epitopes and of the arabinan segment of arabinomannans.

The physical arrangement of cell envelope components leads to the exposure of selected structural motifs which in turn may influence host-parasite interactions. To gain insight into the exposed epitopes, the present study describes a flow cytometric method designed to probe defined molecules on dispersed mycobacteria. The hydrophobic fluorophore N-hexadecanoyl aminofluorescein inserted in the mycobacterial cell envelope permitted focusing of fluorescence-activated cell sorter analysis on cells that were further labeled with defined monoclonal antibodies and fluorochrome-coupled streptavidin. The use of antibodies directed against the lipooligosaccharide of Mycobacterium tuberculosis demonstrated the specific detection of the antigen on the cell surface of a Canetti-like strain of M. tuberculosis, and not on those of mycobacterial strains that were devoid of the glycolipid. Thus, the method was applied to investigate the relative amounts of surface-exposed mannosylated compounds and D-arabinan-containing substances of different strains of the tubercle bacillus and a strain of the rapidly growing nonpathogenic species Mycobacterium smegmatis. Both M. tuberculosis and M. smegmatis are endowed with mannosyl and arabinan epitopes on their surfaces, although there are many differences in terms of exposed mannosyl epitopes between the various strains of the tubercle bacillus examined. These differences are correlated with the amounts of terminal mannosyl residues that cap the surface-exposed arabinomannans (A. Ortalo-Magné, A. B. Andersen, and M. Daffé, Microbiology 142:927-935, 1996) but not with the degrees of virulence of the strains. This novel approach could provide new insights into the distribution of defined surface-exposed antigens and thereby into the architecture of the cell envelopes.

Antibodies, Bacterial↗
PubMed1996

[Diabetes mellitus is currently one of the most frequent causes or associated causes of uremia. Data from the Piedmont registry of dialysis and transplantation (update, December 1993)].

A regular dialytic treatment of diabetic patients is until accepted from about twenty years in many areas. Aim of this work was a retrospective analysis of main clinical and survival data of diabetic patients (diabetic nephropathy or diabetes as comorbidous factor = 659 cases) admitted for dialysis in Piedmont (Northern Italy Region about 4,400,000 inhabitants) in the period 1981-1993 (functional recovery and follow-up < 1 month excluded). A progressive increment in incidence of diabetic patients was seen mostly in the aged. At 12/31/1993, 263 of 2404 patients admitted for dialysis were diabetics (10.9%); the majority of them was treated in Hospital Centers with bicarbonate haemodialysis (54.4%), while a small group was treated with CAPD (12.9%). During the years ¿80 was seen a progressive leaving of CAPD as first choice method in this population and in the last period the orienteering is the utilization of mixed methods (diffusive-convective as first choice). As regards the survival are not prominent significant differences between this cohort and the cohort affected by vasculopathy as comorbidous factor (86.2 and 54.2% in diabetics vs 78.6 and 55.2% in patients affected by vasculopathy at 1 and 3 years--p = 0.3481; patients aged 45-64 years). In conclusion the cohort of diabetic patients represent a good marker of the clinical problems of the elder population with high clinic risk, in progressive increasing in our Region.

Adult↗
PubMed1995

Platelet-activating factor directly stimulates in vitro migration of endothelial cells and promotes in vivo angiogenesis by a heparin-dependent mechanism.

The aim of the present study was to investigate the angiogenic properties of platelet-activating factor (PAF). In vitro PAF was shown to induce a dose-dependent migration of human endothelial cells (EC) across the polycarbonate filters in Boyden's chambers. In contrast, D-PAF, the biologically inactive enantiomer, and Lyso-PAF did not stimulate a significant migration of EC. This effect of PAF was not associated with a proliferative response of EC to this mediator. Moreover, the ability of PAF to stimulate the migration of EC was independent of the presence of heparin in the medium. WEB 2170, a specific PAF receptor antagonist, prevented the migration of EC induced by PAF, thus suggesting a receptor-dependent stimulation. The expression of PAF receptor gene by EC was confirmed by reverse transcriptase-PCR and Southern blot analysis. The in vivo angiogenic effect of PAF was studied in mice using a model in which Matrigel was used for the delivery of mediators. PAF induced a dose-dependent angiogenic response, which at pharmacologic concentrations (1-5 microM) did not require heparin, but at physiologic concentrations (5-50 nM) required the presence of heparin at doses that were not angiogenic per se. The angiogenesis induced by 50 nM PAF was, indeed, inhibited both by protamine and by the PAF receptor antagonist WEB 2170. The angiogenic effect of D-PAF and Lyso-PAF was not significant. Neutralizing Abs to basic fibroblast growth factor induced a slight but not statistically significant reduction of the angiogenesis induced by PAF.

Animals↗
PubMed1995

A nondegradative route for the removal of endotoxin from exopolysaccharides.

The potentiality of the Triton X-114 phase separation technique for the removal of lipopolysaccharide (LPS, endotoxin) from Klebsiella sp. I-714 exopolysaccharide (EPS) has been investigated. Classical purification and chemical detoxification methods were evaluated for their effectiveness in removing residual LPS, while preserving structural and functional integrity of EPS. Ultracentrifugation, Detoxi-Gel, and ion-exchange chromatography did not remove endotoxin, except gel filtration chromatography performed at 60 degrees C in sodium deoxycholate buffer. In this case, the bioactivity of the purified EPS fraction was significantly lowered, as was seen after alkaline hydrolysis treatment. Moreover, the acetic acid detoxification procedure hydrolyzed EPS. As an alternative, phase partitioning of EPS in Triton X-114 at low temperature provided a fast, mild, and efficient method for the removal of LPS as shown by a 100-fold reduction in Limulus amebocyte lysate (LAL) activity and only a 2-fold reduction in bioactivity. Gel filtration chromatography performed at 4 degrees C with Triton X-114 buffer and phase partitioning with the more hydrophilic Triton X-100 nonionic detergent at 75 degrees C led to a similar decrease in LAL activity. This novel application of Triton X-114 partitioning is a nondegradative alternative to the chemical detoxification of gram-negative bacterial EPS for vaccine production. Purification of endotoxin-contaminated polysaccharides prior to screening for biological activity should also benefit from this technique. The extraction scheme using Triton X-114 can be easily used in large-scale purification processes.

Cell Extracts↗
PubMed1995

Are serum albumin and cholesterol reliable outcome markers in elderly dialysis patients?

Albumin and cholesterol are considered reliable outcome markers in dialysis patients; their influence, however, may also be related to non-independent factors, such as age and presence of co-morbid conditions. The aim of the study was an analysis of four outcome markers, assessed at start of dialysis: age, high risk conditions, cholesterol and albumin levels. Data were obtained from the Piedmont Dialysis and Transplantation Registry (northern Italy, about 4,400,000 inhabitants, 21 dialysis centres, open acceptance since mid-1970s, 5661 patients on file at 31 December 1992). Prevalence of albumin and cholesterol in the normal range increases with age; in each age group prevalence in the range is higher in patients at high risk. However, influence of these biochemical parameters is evident also in no-risk cohorts, thus identifying a subgroup with poorer prognosis also in the population without any identified classic risk factor. The influence of albumin, more evident in the population studied compared with cholesterol, is reflected by impaired survival of low-albumin patients (age > or = 65 high risk at 1 year: 60.7% vs 76.6%, P = 0.0052; age > or = 65 non-high risk, at 1 year: 76.5% vs 90.7%, P = 0.0001). In conclusion, albumin and cholesterol, assessed at start of dialysis, are reliable outcome markers even in elderly patients, identifying, in this high mortality cohort, a subgroup with poorer prognosis. If and how their effect may be reversed by dialysis therapy remains to be assessed.

Adolescent↗
PubMed1993

Hemolytic-uremic syndrome during recombinant alpha-interferon treatment for hairy cell leukemia.

We report a case of hemolytic uremic syndrome in a patient suffering from hairy cell leukemia during recombinant alpha-interferon treatment. We believe that this is the first report of the occurrence of this peculiar kind of acute renal failure following alpha-interferon therapy. This association may suggest possible speculations regarding side effects of interferon treatment and pathogenesis of hemolytic uremic syndrome.

Biopsy↗
PubMed1993

Effect of cytokines on the cytoskeleton of resident glomerular cells.

Resident glomerular cells have complex cytoskeletal organizations relevant to maintaining functional and structural integrity. The ability of cells to change shape, develop coordinate directed movements, replicate and interact with contiguous cells or extracellular matrix depends on cytoskeletal functions. Several cytokines directly or indirectly change cytoskeletal organization. Cytokines may alter cytoskeletal organization in cultured glomerular epithelial and mesangial cells and vascular endothelial cells mainly by mechanisms involving production of secondary mediators, such as PAF and leukotrienes that stimulate cell contraction, or PGE2 and PGI2 that stimulate cell relaxation. Consequent assembly and disassembly of microfilaments leads to functional responses of cells that may account for the physiopathological changes induced by cytokines in vivo.

Animals↗