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Biomedical subjects

G Orlandini

Publications and source records attributed to G Orlandini.

At least 19 recordsLinked to original sources
PubMed2000

Secretin increases the paracellular permeability of CAPAN-1 pancreatic duct cells.

The effects of secretin, the physiological secretagogue for pancreatic ducts, were studied in CAPAN-1 pancreatic duct carcinoma cells. When grown to confluence on plastic dishes, CAPAN-1 cells form domes and exhibit marked increases in culture content of Na+ and urea distribution space (UDS). This parameter is measured as an index of both intracellular and dome compartments under the conditions adopted. Both Na increase and dome formation are inhibited by long term incubation with phorbols, DIDS, DPC, EIPA, H2DIDS, and brefeldin. Short term treatment with secretin or 8-Br-cAMP/teophylline causes dome collapse and a marked decrease in UDS and culture content of Na. Secretin-induced sodium decrease is not abolished by ion channel inhibitors, suggesting that diffusion routes other than ion channels are involved in hormone effects. This hypothesis is also in agreement with data obtained on CAPAN-1 cells cultured on permeable inserts, where no change in Na content or UDS is detected upon secretin treatment. Confluent monolayers exhibit a high transepithelial resistance (Rms) which is markedly and reversibly decreased by secretin. The hormone also decreases the transepithelial voltage (Vms) and raises the monolayer permeability to mannitol. It is concluded that secretin enhances the paracellular permeability of pancreatic duct cells. This effect of secretin, unknown thus far, may be involved in the mechanism of pancreatic secretion in vivo.

Cell Membrane Permeability↗
PubMed2000

Fluorescent in situ hybridisation on tissue sections: a quantitative approach with confocal laser scanning microscopy.

The use of fluorescent detection methods in association with digital microscopy technologies is an innovative approach for tissue localisation of messenger RNA. The success of such methods relies on the tissue preservation, local availability of the probe and on the existence of high resolution tridimensional analysis systems. Cryostatic sections, mild denaturation, short oligonucleotide probes (20mer) and confocal laser scanning microscopy allow the fulfillment of all these conditions avoiding photobleaching and tissue autofluorescence. In this paper, we describe in detail a method for in situ hybridisation set up with digoxigenin-coupled oligonucleotide complementary to beta-actin mRNA as a probe and an anti-hapten fluorescent antibody as second step for detecting specific hybridisation. Fluorescence was analysed by means of a confocal laser scanning microscope (CLSM) that provides images with low out-of-focus blurring also with relatively low numerical aperture (NA) objectives. We propose also an easy method to perform semi-quantitative thresholding analysis which allows to discriminate between background and specific signal.

Humans↗
PubMed1999

Natural anti-endothelial cell antibodies (AECA).

Natural AECA constitute a pool of autoantibodies circulating in healthy subjects, which react with a restricted and conserved set of endothelial antigens and establish idiotypic interactions within the immunoglobulin networks. Normal IgG interacts with living endothelial cells and is internalized with a mechanism involving microtubules and resembling that of ligand-receptor internalization. IgG-endothelial cell interaction appears to be dependent on the variable region of antibodies and is followed by modifications of endothelial cell function. Natural AECA increase anti-inflammatory properties of endothelial cells through the selective inhibition of thromboxane A2, endothelin and metalloproteinase-9 secretion, and also through the inhibition of endothelial cell proinflammatory response to TNF-alpha. We have gathered evidence demonstrating that natural AECA constitute a strictly controlled autoantibody pool, interact with living endothelial cells and take part in the regulation of endothelial function, through direct anti-inflammatory effects.

Antibody Specificity↗
PubMed1999

Histological adaptation of orthotopic ileal neobladder mucosa: 4-year follow-up of 30 patients.

OBJECTIVE: For 4 years we have monitored the histological evolution of ileal neobladders in a single cohort of 30 patients in order to systematically describe the histological changes occurring after surgery. The aim of the study was to evaluate the long-term evolution of many histological parameters with functional relevance as to the metabolic outcome of the reservoirs. METHODS: Ileal samples were collected during surgery and by random biopsies during cystoscopy 6, 12, 18, 24, 36 and 48 months later. At each step qualitative and quantitative assessment of the histological and cytological conditions of the samples was carried out. RESULTS: Morphological changes develop relatively early but the situation tends to level out in about 1 year. The morphological changes are topographically uneven and, although mucosal flattening becomes progressively prevalent, areas with shortened villi persist indefinitely. Goblet cells prevail over enterocytes and the secretive pattern shifts towards sialomucins. The overall replication rate decreases initially but tends to restore in 1 year. Dysplasia or atrophy were never recorded. CONCLUSIONS: The 4-year systematic follow-up revealed a typical histological adaptation pattern in the ileal neobladder without signs of dysplasia. The changes seem to be induced by the aggressive environment and develop in the time lag required for functional adaptation of the epithelium.

Adult↗
PubMed1999

Food allergy and Helicobacter pylori infection.

BACKGROUND: Most antigens reach the immune system through mucosae. Gastrointestinal mucosa is a barrier for alimentary antigens. Inflammatory processes, such as Helicobacter pylori-associated gastritis, could alter the integrity of the gastric barrier, increase the mucosal permeability, and enhance crossing of food antigens which may stimulate allergic reactions. PURPOSE: The aim of this study was to establish whether patients with symptomatic food allergy and detectable immunoglobulin E (IgE) to alimentary antigens were infected by Helicobacter pylori more often than controls, and to determine the phenotype of the infecting Helicobacter pylori. PATIENTS AND METHODS: Thirty-eight consecutive patients with symptomatic food allergy and serum IgE to alimentary antigens, and 53 consecutive age-matched controls (subjects without food allergy and detectable levels of IgE anti-alimentary antigens) living in the same area and attending the same institution were investigated serologically to determine the prevalence of Helicobacter pylori infection, and an immune response to CagA, a marker of the most pathogenic strains. IgE to alimentary allergens were measured by a commercial kit. RESULTS: The prevalence of Helicobacter pylori infection in patients with food allergy and controls was similar (42.1% and 47.1%, respectively). Anti-CagA antibodies in Helicobacter pylori-infected persons were detected in 62.5% of patients with food allergy, and 28.0% of controls (p = 0.030, odds ratio = 4.29, RR = 2.23). The mean IgE level to the most common alimentary antigens was increased in CagA-positive, with respect to the CagA-negative, patients. CONCLUSIONS: The enhanced mucosal and inflammatory lesions commonly found in individuals infected by CagA-positive Helicobacter pylori strains could increase the epithelial permeability and render non-selective the passage of allergens which, in atopic persons, could directly stimulate an IgE response. Infection by CagA-positive Helicobacter pylori may increase the risk of food allergy.

Adolescent↗
PubMed1999

The infection by Helicobacter pylori strains expressing CagA is highly prevalent in women with autoimmune thyroid disorders.

UNLABELLED: H. pylori infection is putatively associated with extra-digestive disorders and may also play a role in the development of autoimmune thyroid diseases (ATD). It was recently found that monoclonal antibodies to an H. pylori strain with cagA-positivity reacted with follicular cells of the thyroid gland, and that an H. pylori organism possessing the cag pathogenicity island carried a gene encoding for an endogenous peroxidase. The aims of this study was (1); To ascertain whether the infection by strains endowed with an increased inflammatory potential (those expressing CagA) could further enhance the risk of developing ATD (2); To verify the possible existence of an immune cross-reactivity between autoantibodies to peroxidase and thyroglobulin and H. pylori antigens (3). To establish whether thyroid colloid antigens could cross-react with an anti-H. pylori serum. The study was partly designed retrospectively. We examined 41 consecutive women with ATD, and, as a control, 33 consecutive age- and socio-economic class-matched women without autoimmune thyroid disorders, living in the same area as patients, occurred at the same institution in the same period (six months). Both patients and controls were examined serologically for H. pylori infection and CagA status by Western blotting. Some serum samples were absorbed with H. pylori to determine whether the antibody levels decreased. Colloid proteins were resolved electrophoretically and matched with a hyperimmune serum raised in rabbits against a CagA-positive H. pylori. Thirty-two patients (78.0%) tested seropositive for H. pylori infection, vs. 16 controls (48.4%) (P = 0.008, OR = 3.78, RR = 1.61). The prevalence of anti-CagA antibodies was 71.8% in infected patients, and 50% in infected controls (P = 0.161, n.s.). The overall prevalence of infection by CagA-positive H. pylori was significantly higher in patients with ATD (23/41, or 56.0%) than that in controls (8/33, or 24.2%) (P = 0.006, OR = 3.99, RR = 2.31). The other tests gave negative or inexplicable results. IN CONCLUSION: CagA-positive H. pylori infection increases the risk of ATD development.

Adolescent↗
PubMed1999

CagA-positive Helicobacter pylori infection may increase the risk of food allergy development.

The aim of this study was to test whether patients with symptomatic food allergy and significant levels of immunoglobulin E (IgE) to alimentary antigens were more likely infected by H. pylori, especially by strains expressing the CagA protein, with respect to controls. A group of 38 patients with symptomatic food allergy and 53 age-matched controls were examined serologically for H. pylori infectious status, and for CagA seropositivity. IgE to alimentary allergens were measured by a commercial kit. The prevalence of H. pylori infection in patients with food allergy and controls was similar (42.1%, and 48.3%, respectively). However, anti-CagA antibodies in H. pylori-infected persons were detected in 62.5% of patients with food allergy, and 28% of controls (P = 0.030, odds ratio = 4.29). The mean level of IgE to the most common alimentary antigens in serum samples from infected patients with anti-CagA antibodies was significantly higher than in CagA-negative infected patients: 3.28 kU/L (SD 3.93), vs. 1.99 kU/L (SD 1.53), P = 0.002, 95% confidence interval = 0.61 to 2.53). Infection by CagA-positive H. pylori increases the risk of developing food allergy.

Adolescent↗
PubMed1998

Enhancement of convective transport by internal filtration in a modified experimental hemodialyzer: technical note.

BACKGROUND: Hemodialysis using high flux membranes today is a commonly used therapy. The primary advantage is the larger spectrum of molecules removed with these membranes, and the mechanism of removal is in part due to a phenomenon of filtration and backfiltration along the length of the hollow fibers. We hypothesized that increasing the filtration and backfiltration fluxes by modifying the structure of the dialyzer could enhance the convective transport of various solutes. METHODS: A modified high flux dialyzer was compared to the standard model in terms of pressure profiles, filtration-backfiltration rates and solute clearances. The modification consisted on the placement of a O-ring around the fiber bundle to create a resistance for the flow of the dialysis solution external to the fibers. The study on filtration fluxes was carried out using a scintigraphic method previously described, and solute clearances were studied during ultrafiltration-controlled hemodialysis sessions. RESULTS: Utilizing a net filtration condition proximal to zero, the rates of proximal filtration and distal backfiltration in the experimental dialyzer were significantly enhanced in comparison with the standard dialyzer. The pressure drop in the dialysate compartment could be increased significantly, thus permitting an increase in the positive transmembrane pressure in the first half of the dialyzer and a parallel increase in the negative transmembrane pressure in the second half of the dialyzer. This resulted in a significant enhancement of the convective transport of middle-large solutes as demonstrated by the increase in vitamin B12 and inulin clearances. CONCLUSIONS: This approach suggests that changes in design of the dialyzer may affect its performance. The use of internal filtration is suggested to improve convection and dialyzer efficiency for larger solutes without the requirement for high volumes of replacement fluid, as is the case for current hemodiafiltration techniques.

Hemodiafiltration↗
PubMed1998

Influence of age on enzyme activities of pyrimidine metabolism in the chicken heart.

The effect of the aging on the activities of enzymes involved in UMP-CMP metabolism were evaluated in the heart of newborn (1-day-old), young (20-day-old), adult (12-month-old), and aged (30-month-old) chickens. In newborn animals, UMP metabolism proceeds preferentially towards cytidine compounds rather than to breakdown. In addition, two pathways different from those involved in de novo synthesis may contribute to the synthesis of UMP: one, through cytosine deaminase that shows its maximal activity; the other, by uridine kinase, the main "salvage" enzyme of pyrimidine nucleotides. In young chickens, pyrimidine metabolism regards especially UMP. In fact, the lower activities of cytidylate phosphatase and cytosine deaminase, together with the remarkable increase of uridine kinase indicate that the metabolic flux converges on the main salvage pathway. In adult chickens, pyrimidine catabolism is enhanced, as supported by the maximal activity of the enzymes involved in UMP-CMP breakdown. On the contrary, the remarkable reduction of the anabolic enzymes suggests a limited resort to the salvage pathways. Finally, in aged chickens a reduced pyrimidine catabolism and a greater utilization of the salvage pathways appear to take place, thus contributing to the maintenance of pyrimidine nucleotide pool.

5'-Nucleotidase↗
PubMed1998

Histomorphology of an ileal orthotopic neobladder: 24 Year's follow-up.

No morphological data are so far available about long-term (over 15 years) adaptation and outcome of ileal neobladders. In this single case report we have studied a 72-year-old man who underwent reconstruction of an ileal neobladder 24 years ago. The ileal epithelium shows changes toward a colonic aspect with villous atrophy and increased goblet cell number although, as in normal ileum, sialomucins are the most abundant secretory products. Neuroendocrine cells are reduced to a few elements of a single morphological class. Even in the presence of a low apoptotic rate reduced cell proliferation causes mucosal thinning. These adaptive mechanisms grant an excellent clinical and functional result even beyond 20 years after surgery. It is suggested that ileal neobladder can constitute an important model for the study of epithelial differentiation control.

Aged↗
PubMed1998

Comparison of annexin V and calcein-AM as early vital markers of apoptosis in adherent cells by confocal laser microscopy.

Although morphological criteria for apoptosis are in general reliable, no systematic comparison of the techniques employed thus far has yet been performed. In this study, using confocal laser microscopy, we compared the performance of annexin V-FITC and calcein-AM for early detection of apoptosis in living adherent cells. Experiments were carried out on two distinct cell lines, PC 12 and NIH3T3, endowed with different shape and adhesion properties. The apoptotic process was followed for a prolonged period in the same cells of a predetermined field by means of a special flow chamber. Our results show that both probes allowed the detection of apoptotic cells in either cell line. However, some cells that clearly exhibited apoptotic changes on calcein visualization were annexin-negative. In NIH3T3 cells, annexin negativity of apoptotic cells was correlated with the preservation of cell shape and adhesion properties. These findings show that, at least in PC12 and NIH3T3 cells, annexin might be less sensitive than calcein-AM for early apoptosis detection and, for NIH3T3 cells, suggest that phosphatidilserine exposure is in some way linked to changes in cell shape and/or adhesion to culture substrate. (J Histochem Cytochem 46:895-900, 1998)

3T3 Cells↗
PubMed1998

Reduction of mononuclear cytokine production in hemodialysis patients treated with steam-sterilized low-flux polysulphone membranes.

An increased cytokine production, correlated with long term complications of uremic disease, has been described during hemodialysis. To identify possible differences in the cytokine release of differently sterilized membranes, we enrolled six uremic patients on chronic hemodialysis. The patients underwent dialysis with ETO-sterilized low-flux polysulphone membranes (F6, Fresenius AG) for at least three months (A1), they were then switched to steam-sterilized polysulphone membranes (F6-HPS Fresenius AG) and further evaluations after one (B1) and two months (B2) were carried out. A final evaluation (A2) was made one month after switching back to F6 dialyzers. At each time period, samples were drawn to measure IL-1beta released by cultured mononuclear cells (MN). Moreover, dialysate samples were collected to test endotoxin levels. C3a and C5a levels were assessed at 0, 5, 15 and 60 min from starting hemodialysis. Anti-ETO IgE levels were also assayed at A1, B1 and A2. The LAL test revealed a good quality dialysate. The mean pre-dialysis IL-1beta levels were 215 pg/million cells at A1; falling to 49 at B1, and 54 at B2 (p<0.01); there was then a sharp rebound at A2: 284, p<0.01. Post-dialysis levels followed the same pattern. No correlation between the dialysate endotoxin level and cytokine release was found. Complement activation did not change and in all the phases of the study no anti-ETO IgE was detected in any of the subjects. Our data suggest that the steam sterilized polysulphone membrane induces a lower cytokine release than the ETO sterilized membrane, although the mechanism by which it does so remains to be clarified.

Adult↗
PubMed1997

Membrane potential changes visualized in complete growth media through confocal laser scanning microscopy of bis-oxonol-loaded cells.

Confocal laser scanning microscopy (CLSM) was employed to visualize and measure membrane potential changes in several types of cultured adherent cells, such as human fibroblasts, mouse mammary tumor C127 cells, and human saphenous vein endothelial cells, preloaded with the anionic dye bis-1, 3,-diethylthiobarbituratetrimethineoxonol (bis-oxonol). The fluorescence of cell-associated bis-oxonol was detected in a single confocal plane. An original flow-chamber apparatus was employed to replace the extracellular medium, avoiding alterations of the plane selected for observation. In all the cell types and the experimental situations tested the intracellular distribution of the dye was typical; perinuclear zones accumulated the dye which, conversely, was excluded by the nucleus. Fluorescence was calibrated versus the membrane potential by varying the extracellular concentration of sodium in the presence of gramicidin. With this approach membrane potential was measured (i) in cultured human fibroblasts incubated under anisotonic conditions, (ii) in heterogeneous cell populations which respond unevenly to potential perturbing conditions, and (iii) in human macrovascular endothelial cells maintained in high-serum, complete growth medium. The results obtained indicate that CLSM can be successfully employed to measure changes of membrane potential in single, bis-oxonol-loaded adherent cells under experimental conditions which severely hinder conventional spectrofluorimetric approaches.

Animals↗
PubMed1997

Effects of reduced preload on diastolic filling in essential hypertensive patients with increased left ventricular mass.

Doppler-derived indices of diastolic filling are widely used in the routine evaluation of essential hypertensives. However, these indices are affected by loading conditions and systolic performance. This study aimed at monitoring the transmitral flow pattern and indices of left ventricular systolic function during acute nonpharmacological isolated reduction of preload in essential hypertensives with left ventricular hypertrophy. Nine essential hypertensive patients with left ventricular hypertrophy and nine age- and sex-matched normotensive controls underwent echocardiographic and Doppler evaluation of both systolic function and diastolic filling indices at baseline and during lower body suction at -40 mm Hg. Lower body suction caused a similar decrease in end-diastolic volume index, stroke volume index, and midwall fractional shortening in the normotensives and hypertensives. Circumferential end-systolic stress was unaffected in both groups. Acceleration time of early diastolic filling and isovolumic relaxation time increased in the normotensives but not in the hypertensives. Deceleration time of early diastolic filling increased in both groups. The ratio of peak velocities during early filling and at atrial contraction decreased in the normotensives, whereas it was unchanged in the hypertensives; this was due to the fact that early filling velocity decreased in both groups, whereas peak velocity at atrial contraction decreased only in the hypertensives. We conclude that Doppler-derived diastolic filling indices are not affected by a reduction of preload in essential hypertensives with left ventricular hypertrophy.

Diastole↗
PubMed1997

Binding and internalization of human IgG by living cultured endothelial cells.

Interactions between circulating IgG and endothelial cells (EC) in humans have been described only in conditions associated with pathologic immunoglobulins and/or activated or damaged EC. In this study we provide evidence that normal human IgG includes one/some antibody species that bind to and are internalized by living EC in culture. This novel function of EC and natural autoantibodies is of potential importance for the understanding of physiologic interactions between vessels and the immune system and for the clarification of pathogenesis of vasculitis and mechanisms of action of pooled IgG used in the therapy of such conditions.

Autoantibodies↗
PubMed1997

Use of Fourier shape descriptors to improve the reproducibility of echographic measurements of arterial intima-media thickness.

BACKGROUND: A major source of error in the longitudinal assessment of the intima-media thickness (IMT) is the difficulty in retrieving the same echographic view of the vessel. OBJECTIVE: To present a method for increasing the reproducibility of IMT measurements by ultrasound in large arteries. METHOD: The Fourier descriptor is a well-known means of describing an object's shape. By means of the discrete Fourier transform (DFT), the shape was represented in a frequency domain; the computational advantages of the DFT then permitted a measure of unlikeness between different shapes (the 'distance' measure; DM) to be defined and used as a criterion for reproducing the contour. When the sonographer compared successive images of a complex vascular segment, like the carotid bifurcation, the identity of the echographic cut was deduced from the identity of the vessel's contour. The best match of the baseline image was the view that minimized the contour DM. RESULTS: Preliminary studies in the carotid artery bifurcations of eight subjects showed that the DM responds to systematic variations in the ultrasound interrogation angle and reveals minimal changes in transducer position. Duplicate scans of 12 subjects were performed by three sonographers with different strategies for acquisition of the same images: a low DM was associated with a low difference in pairs of IMT measurements. Data were classified into two groups (normal or borderline vessels with a pooled mean IMT of 0.62 mm and overtly thickened segments with a pooled mean IMT of 1.31 mm). When minimization of the DM was the criterion for the acquisition of replicate scans, the mean absolute difference of paired data for the mean IMT of the distal common carotid artery was 0.03 +/- 0.02 mm for the first group and 0.06 +/- 0.03 mm for the second group. This is a significant reduction in comparison with non-quantitative alternative criteria for image reproduction. For the maximum IMT of the same segments the mean absolute differences were 0.07 +/- 0.03 and 0.13 +/- 0.06 mm in the first and second groups, respectively. CONCLUSION: This method can be applied to the serial assessment of single atherosclerotic segments. The computational time is negligible. By reducing the scatter in sequential IMT data, longitudinal investigations (e.g. of the results of antihypertensive therapy) with shorter durations and smaller sample groups may be rendered feasible.

Algorithms↗
PubMed1996

Effect of a new model of hemodialysis potassium removal on the control of ventricular arrhythmias.

The primary aim of this multicenter, prospective, randomized cross-over study was to clarify whether a new model of hemodialysis (HD) potassium (K) removal using a decreasing intra-HD dialysate K concentration and a constant plasma-dialysate K gradient (treatment B) is capable of reducing the arrhythmogenic effect of standard HD, which has a constant dialysate K concentration and decreasing plasma-dialysate K gradient (treatment A). The secondary aim was to verify whether this new model is clinically safe. In treatment B, the initial dialysate K concentration had to be 1.5 mEq/liter less than the plasma K concentration, and exponentially decrease to 2.5 mEq/liter at the end of HD. Forty-two chronic HD patients with an increase in premature ventricular complexes (PVC) during dialysis were enrolled from 18 participating centers, and randomly assigned to either sequence 1 (ABA) or sequence 2 (BAB). A pool of 333 of 378 expected ECG Holter recordings were checked for signal quality; 269 (71%) from 36 patients (86%) had a satisfactory signal quality and 108 were selected for analysis (1 per patient per period). There was a difference in the natural logarithm of the increase in PVC/hr and PVC couplets/hr during HD between treatments A and B (1.70 +/- 1.59 vs. 1.09 +/- 1.76 and 0.94 +/- 0.86 vs. 0.64 +/- 1.01, a reduction of 36% and 32%, P = 0.011 and 0.047, respectively) without any carry over effect (P = 0.61 and 0.24, respectively). The fact that this decrease of one third is due to a lower plasma-dialysate K gradient is supported by the observation that it was more evident during the first than the last two hours of HD (a reduction in the natural logarithm of the increase in PVC/hr and PVC couplets/hr of 60% and 60%, P 0.002 and 0.009, vs. 26% and 17%, P = 0.098 and 0.332, respectively): the initial plasma-dialysate K gradient was 2.3 times lower during treatment B than during treatment A, without adversely affecting pre-HD plasma K levels. These results could have a considerably clinical impact not only because of the possibility of physiologically decreasing the arrhythmogenic effect of HD, but also because this effect can be considered a "marker" of the electrophysiological derangement induced by the administration of standard HD three times a week for years ("electric disequilibrium syndrome").

Aged↗