Search PubMed⌕ Search

Biomedical subjects

E Brambilla

Publications and source records attributed to E Brambilla.

At least 163 records · Page 9Linked to original sources
PubMed1994

Gamma interferon activates a previously undescribed Ca2+ influx in T lymphocytes from patients with multiple sclerosis.

Multiple sclerosis (MS) is an immune-mediated demyelinating disease of the central nervous system. The etiology of the disease is still unknown. Activated T lymphocytes are considered essential in mediating the inflammatory process leading to demyelination in MS. They operate through a complex network of cytokines among which gamma interferon (gamma-IFN) plays a key role. Here we report that exposure to gamma-IFN of T lymphocytes from patients with MS activates, by a protein kinase C-mediated pathway, a previously undescribed gamma-IFN-activated Ca2+ influx, functionally coupled to the gamma-IFN receptor. The influx mainly expressed by CD4+ T lymphocytes, was found in 12 of 15 (80%) patients with clinically active MS and in 14 of 30 (46%) patients with stable MS. The influx was found in only 3 of 24 (12%) control patients and in none of the 15 healthy subjects studied. Our results document the appearance in MS lymphocytes of a gamma-IFN-activated, protein kinase C-dependent, Ca2+ influx that might be due to the expression of a new cation-specific plasmalemma channel. This finding suggests that at least part of gamma-IFN's contribution to the pathogenesis of MS is exerted through a Ca(2+)-dependent regulation of T lymphocyte activity.

Adult↗
PubMed1994

Intracellular Ca2+ stores of T lymphocytes: changes induced by in vitro and in vivo activation.

Intracellular Ca2+ stores were investigated in resting and activated splenic T lymphocytes from Lewis rats. Activation was obtained either in vitro (spleen cells isolated from "naive" rats exposed to concanavalin A for 24 h) or in vivo (spleen cells from rats with fully developed symptoms of experimental allergic encephalomyelitis). In both experimental conditions several changes of Ca2+ homeostasis were observed with respect to resting lymphocytes: (1) a threefold increase of the total intracellular calcium (from 1.15 to 3.5 mmol/l); (2) a moderate increase of the pool sensitive to inositol 1,4,5-trisphosphate (IP3), investigated both in intact T lymphocytes (fura-2 and 45Ca(2+)-release techniques in cells challenged with phytohemagglutinin) and in T lymphocytes permeabilized with beta-escin (45Ca2+ release induced by saturating concentrations of IP3); and (3) the appearance of a pool released by the endoplasmic reticulum (ER) Ca2+ ATPase inhibitor thapsigargin (Tg), but insensitive to IP3, which, therefore, appears to be localized in areas of the ER devoid of the cognate receptor. The latter two findings were paralleled in activated lymphocytes by an increase of expression of ER markers, involved (calreticulin; Ca2+ ATPase) or not (protein disulfide isomerase) in the regulation of Ca2+ homeostasis. In contrast, calnexin (another ER marker) and the receptor for IP3 were increased to only a moderate extent. Finally, an enlargement of non-ER Ca2+ pools was observed in the cells pretreated with Tg in which 45Ca2+ release was induced by the Ca2+ ionophore ionomycin. Our results document structural and functional changes of intracellular Ca2+ stores which might play an important regulatory role in activated T lymphocytes.

Animals↗
PubMed1994

Oral administration of fluoride in pregnant women, and the relation between concentration in maternal plasma and in amniotic fluid.

The aim was to measure the ionic fluoride concentration in maternal plasma and in amniotic fluid after oral administration of different doses of sodium fluoride (NaF) to 121 pregnant women. They were divided into six groups, according to the dose administered; 0 for the control group and 0.25, 0.50, 0.75, 1.00 or 1.25 mg of F- for the others. The subjects were instructed to take the corresponding NaF dose both 24 and 3 h before amniocentesis. Amniotic fluid (5 ml) and venous blood (5 ml) were obtained from each subject. Ionic fluoride concentration was measured with an expanded-scale potentiometer and a selective fluoride electrode. The results showed that F- concentration in amniotic fluid and, presumably, in fetal circulation, was not significantly different in groups taking 0.25, 0.50, 0.75 or 1.00 mg/day of F-. The F- concentration in amniotic fluid of the 1.25 mg/day group was, however, significantly higher than in all the other groups.

Administration, Oral↗
PubMed1994

Absence of central nervous system pathology in severe combined immunodeficiency mice intraperitoneally injected with peripheral blood lymphocytes from multiple sclerosis patients.

In order to reproduce some of the pathological features of multiple sclerosis (MS) we transplanted peripheral blood lymphocytes (PBLs) from seven patients with MS into the peritoneal cavity of 28 severe combined immunodeficiency (MS-SCID) mice. Seven SCID mice were also transplanted with PBLs from two healthy subjects (hu-SCID). Animals were sacrificed between 2 and 8 weeks after transplantation (a.t.). Polymerase chain reaction (PCR) using primers able to amplify the HLA-DQ alpha region showed presence of human cells in neural tissues of MS-SCID mice. Immunocytochemical analysis revealed the scattered appearance of human lymphocytes (mostly CD45RO+ T cells) in the meningeal space and choroid plexuses of MS-SCID brains. However, human lymphocytes were similarly found in brains of hu-SCID mice. Both groups of mice never showed signs or symptoms of neurological impairment. Our results indicate that the simple transplantation of lymphocytes from MS patients into SCID mice is not likely to produce an MS-like pathology.

Animals↗
PubMed1994

Steroidogenic adrenocortical cells synthesize alpha 2-macroglobulin in vitro, not in vivo.

We previously identified alpha 2-macroglobulin as the major protein secreted by primary cultures of adrenocortical cells. We report here that in the adrenal gland, the distribution of alpha 2-macroglobulin in the adrenocortical tissue is restricted to the endothelium of blood vessels and that no immunoreactivity is found in steroidogenic cells. A time course study revealed that freshly dissociated bovine adrenocortical cells were void of alpha 2-macroglobulin immunoreactivity whereas the proportion of alpha 2-macroglobulin-positive cells reached more than two-thirds of the population between day 4 and day 7 of culture. Double immunoenzymatic labeling of 6-day-old cultures revealed a co-localization of alpha 2-macroglobulin and the steroidogenic enzyme P-450SCC. Treatment of 5-day-old cultures (expressing alpha 2-macroglobulin) for 24 h by either ACTH (10(-9)-10(-6) M) or alpha 2-macroglobulin (2.5 mg/ml) resulted in a marked decrease of the expression of alpha 2-macroglobulin. These data indicate that ACTH and plasmatic alpha 2-macroglobulin could physiologically repress alpha 2-macroglobulin expression in the adrenal cortex in vivo.

Adrenal Cortex↗
PubMed1994

Immunoscintigraphy using 111in-labelled F(ab')2 fragments of anti-carcinoembryonic antigen (CEA) monoclonal antibody for staging of non-small cell lung carcinoma.

28 patients with primitive lung cancers were imaged by immunoscintigraphy (IS) with 111indium-labelled F(ab')2 anti-carcinoembryonic antigen (CEA), to assess this technique for mediastinal staging. IS revealed primitive tumours in 21 cases in whom mediastinal extension was assessed. There was concordance between clinical staging and IS confirmed by surgery in 17 cases, and discordance in 4. After surgery, discordance was in favour of IS in 2 cases (1 true positive and 1 true negative) and in favour of clinical staging in 2 (false positive of immunoscintigraphy). Anti-CEA IS could be useful for improving mediastinal staging of lung cancer.

Antibodies, Monoclonal↗
PubMed1994

Detection of anti-acetylcholine receptor antibody by an ELISA using human receptor from a rhabdomyosarcoma cell line.

A simple and reliable enzyme-linked immunosorbent assay (ELISA) has been developed for the measurement of anti-acetylcholine receptor (AChR) antibodies. The test utilizes a membrane-bound AChR obtained from a human rhabdomyosarcoma cell line (TE671) as antigen and employs an affinity-purified rabbit anti-human immunoglobulin G alkaline phosphatase-conjugated antibody as labelled antibody. To assess the sensitivity and the specificity of our assay we tested serum samples from 13 anti-AChR antibody-positive myasthenia gravis (MG) patients known to contain between 2 and 120 nmol/l of anti-AChR antibody, three anti-AChR antibody-negative MG patients, and 70 control subjects including patients with other neurological and autoimmune diseases. A panel of six different anti-AChR monoclonal antibodies and membranes from a AChR-negative rat adrenal pheochromocytoma cell line (PC 12) were also used in competitive studies. The test showed to be specific and able to detect as low as 2.0 nmol/l of anti-AChR antibodies. Moreover, we found a good correspondence between anti-AChR antibody levels measured in the serum samples tested by our assay and levels measured by the routinely adopted radioimmuno assay (RIA) using human-AChR (r = 0.96). Cross-reaction phenomena were observed only using serum samples containing high-titer anti-DNA antibodies. The proposed ELISA, circumventing the limitation of the commonly used RIA (radioactivity and amputated legs as source of human antigen), can be considered as an useful screening test for the diagnosis of myasthenia gravis.

Antibodies, Monoclonal↗
PubMed1994

F(ab) secondary antibodies: a general method for double immunolabeling with primary antisera from the same species. Efficiency control by chemiluminescence.

We propose a general solution to the problem of using antibodies originating in the same species for double immunohistochemical labeling. It relies on the use of a two-step protocol in which a secondary polyclonal monovalent F(ab) antibody present in the first step blocks access in the second of the secondary antibody to the primary antibody, which is continuously present from the first step. The monovalence of the F(ab) fragment eliminates the possibility of its linking the primary antibody from the second step. We designed two efficiency tests to explore the limits of the method by the very sensitive chemiluminescent system applied to sections of human pituitary tissue. They confirmed both the validity of the method and the necessity of adapting working conditions to obtain a complete lack of interference.

Animals↗
PubMed1994

p53 immunolabeling in archival paraffin-embedded tissues: optimal protocol based on microwave heating for eight antibodies on lung carcinomas.

The prognostic value of p53 gene mutations is dealt with by several recent reports. However, retrospective assessment of p53 tumor status on archived samples has been prevented by p53 epitope alteration during routine fixation and embedding procedures. This study aimed at establishing a reproducible low-cost protocol to retrieve not only N-terminal, but also midregion and C-terminal, epitopes, with special attention to possible artifacts induced by epitope retrieval procedures. Using microwave heating, we compared the epitope retrieval efficiency of five solutions with eight commercial antibodies on 21 lung carcinomas for which frozen tissue and samples fixed with formalin and Bouin's liquid were available. All eight epitopes were retrieved, citrate buffer proving efficient for seven. PAb 240 epitope was restored by target unmasking fluid only. No false positivity was observed. Fixation-induced loss of p53 immunoreactivity was minimal for formalin (two of 10 tumors for one antibody each), more significant for Bouin (six of 10 tumors for one to five antibodies). On the other hand, staining intensity was maintained or even improved, and nonspecific staining reduced, through fixation. We conclude that p53 stabilization can be detected on routinely processed archival tumor samples with a reliability similar to that of frozen tissue by means of a microwave-based procedure and a panel of at least three antibodies, with epitopes on the N-terminal, C-terminal, and midpart of the molecule.

Antibodies, Monoclonal↗
PubMed1994

[Respective roles of the products of the mdr1 and p53 genes in drug resistance of bronchial cancers].

Expression of mdr1 and p53 was assessed on 119 cases of bronchial carcinomas, and compared with clinical chemoresistance. mdr1 expression was evaluated by immunohistochemistry (IHC), using the monoclonal antibody JSB1. The study of p53 expression was performed by both IHC, using six different antibodies, and Northern blotting. We observed a correlation between the expression of mdr1 and the presence of a mutation of p53 in neuro endocrine (NE) carcinomas (P = 0.02). Correlation was not observed when non NE carcinomas were evaluated, nor was found any correlation between mdr1 expression and clinical chemoresistance in patients with small cell lung carcinoma (SCLC). The frequency of complete response however was significantly higher in patients whose tumor did no express mdr1 (P = 0.02). Chemoresistance correlated well with the phenotype of p53 mutant in SCC (P = 0.015). We conclude that p53 mutation is a better predictive factor of clinical chemoresistance in SCLC than mdr1 IHC detection.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1994

[Small cell bronchial cancer in a 17-year-old young man].

We report a case of small cell lung cancer in a 17 year-old man. He was admitted to our unit suffering from a two month history of pain left shoulder. Chest X-rays showed a large round mass in the left upper lobe. The chest CT scan revealed a tumor with evidence of first left rib involvement. Histological examination, after surgical biopsy revealed small cell carcinoma, confirmed by two independent pathological physicians. Small cell lung carcinoma is very rare in the under 20 year age group and we compare this case with other types of lung cancer in young patients described in the literature.

Adolescent↗
PubMed1993

[Mechanisms of lung oncogenesis].

Precancerous lesions of the bronchial epithelium are dysplasias and in situ carcinomas. Squamous metaplasia has not yet been considered as a true malignant state. Epithelial cells, which are able to proliferate (non terminally differentiated) in bronchial tree and alveoli, are the candidates for malignant proliferation (basal cells, mucus cells, Clara cells and type II pneumonocytes). Their initial growth is probably promoted by deregulated autocrine growth factors (EGF, GRP, IGF1), or their receptors (EGF-R). Under continuous carcinogens exposition these proliferating cells accumulate multiple genetic abnormalities affecting dominant oncogenes such as myc and ras, and recessive tumor suppressor genes such as Rb and p53. Neither the order of intervention of these genetic factors nor their correlation with premalignant states have been demonstrated.

Humans↗
PubMed1993

Nonspecific increased serum levels of secretory component in lung tumors: relationship to the gene expression of the transmembrane receptor form.

Polymeric immunoglobulin receptor (pIg-R) is synthesized by epithelial cells lining the bronchial mucosa. It is released in secretions as free secretory component (SC) or bound to Ig as secretory Ig (S-IgA and S-IgM). To evaluate the usefulness of SC and pIg-R expression as tumour markers, we measured SC and secretory Ig, using enzyme-linked immunosorbent assay, in the serum of 45 patients with lung carcinomas, in the serum of 10 patients with non-neoplastic diseases, and in the serum of 45 control subjects. We also studied the immunohistochemical expression of pIg-R and its mRNA in tumors from 20 out of the 45 patients. Serum levels of SC and S-IgA were similarly and significantly elevated in patients with lung cancer (squamous cell carcinoma [25 cases], small cell carcinoma [7 cases], adenocarcinoma [13 cases]) and with non-neoplastic diseases, as compared with control subject levels (P < 0.001). The highest SC levels were found in patients with adenocarcinoma although the mean SC level was not different from other pathologic conditions. pIg-R was usually not detected in the cells of small cell carcinoma or of squamous cell carcinoma, whereas it was found in the cells of five adenocarcinomas and in the two in situ carcinomas under study. The specific mRNA analysis usually agreed with the immunolocalization of pIg-R. A single band at 3.8 kb was detected in the positive tumor tissues and in normal lung tissues. However, the signal was weak in one case of squamous carcinoma and stronger in two out of three adenocarcinomas, than in normal tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

[Expression of oncogenes C-myc, C-raf and N-ras in advanced cancers of the upper respiratory and digestive tracts. Correlation with tumor clinical response to chemotherapy].

A correlation between oncogenes expression (N-ras, C-myc, C-raf) and response to neoadjuvant chemotherapy has not been established in carcinology of the head and neck yet. We have studied the expression of these three oncogenes in 24 tumors patients with squamous cell carcinoma previously untreated. Tumors samples were biopsed before initiation of treatment. After extraction of total RNAs, oncogene expression was measured by northern and slot blot analysis using 32P radiolabeled probes. The endoscopic, histological and clinical response was evaluated after chemotherapy (5-FU: 1 g/m2/d, D1 to D5; CDDP: 100 mg/m2 at D1, 3 courses every 3 weeks). There was no significant difference of N-ras expression between responding or resistant tumors to chemotherapy. However, there was a significant difference of C-myc (p < 0.05) and C-raf (p < 0.01) expression between the two patients population: C-raf and C-myc expression were higher in the responding tumors than in the resistant tumors to chemotherapy. In conclusion, C-myc and C-raf expression could be a marker for chemosensitivity.

Adult↗
PubMed1993

Lipoprotein structure in male subjects during in vivo lipolysis: effect of an anti-lipolytic treatment with acipimox.

Plasma free fatty acid (FFA) levels were raised in healthy volunteers by the administration of a fatty meal and epinephrine infusion (0.15 mg/kg per min), to test the hypothesis that enhanced lipolysis might lead to changes in lipoprotein distribution and to the formation of lipoprotein complexes, also impairing the interconversion of high density lipoproteins (HDL). The study was carried out in double-blind conditions in volunteers pre-treated with either placebo or with acipimox, a nicotinic acid analogue with a long-lasting activity. Lipolysis was effectively induced; the treatment with acipimox prevented the rise of free fatty acids (FFA), and it also blunted the triglyceride (TG) increase occurring during the test. Whereas the mean low density lipoprotein (LDL) particle size did not change, the HDL particle distribution showed a progressive shift to smaller particles, both after placebo and after acipimox, the changes in size being maximal 3-7 h after the meal. Evaluation of HDL interconversion in plasma samples incubated at 37 degrees C for 6 h showed the expected accumulation of HDL2a particles, with a parallel decrease of HDL3a; however, this conversion was not affected by the presence of elevated FFA levels and no difference was noted in subjects taking either placebo or acipimox. These clinical data fail to confirm the hypothesis that enhanced lipolysis may lead to dramatic changes in plasma lipoprotein distribution and/or in aggregation or fusion of lipoprotein particles, as reported from in vitro experiments. This study, however, successfully achieved a useful model of exaggerated lipolysis and confirmed the important activity of a low dose nicotinic acid analogue in inhibiting lipolysis.

Adult↗
PubMed1993

Immunohistochemical study of p53 in human lung carcinomas.

Immunohistochemical analysis of p53 protein was carried out on 95 lung carcinomas from all histological types, including 60 primary tumors, 35 lymph node metastases, and 36 corresponding nude mice xenografts, using four antibodies: PAb240 specific for some mutant conformations; PAb421, PAb1801, and CM1 reactive with most of the forms of p53. Nuclear staining with at least two of those four antibodies revealed the presence of an accumulated protein, considered as indicative of a missense mutation in the p53 gene, in 50% of primary tumors of all histological types, except carcinoids. Some defect of messenger RNA expression was detected by Northern blot analysis in an additional 26% of tumors. p53 immunophenotype of the original tumor was fairly maintained on nude mice. p53 accumulation was not correlated with survival, but with disease extension (P = 0.01). Finally, immunohistochemical analysis allowed the recognition of p53 mutant immunophenotype in 41% of tumors where p53 DNA and messenger RNA were apparently normal, using standard molecular biology. Thus, this method provides a rapid and efficient approach for studying p53 mutations leading to an accumulated protein in lung tumors cells.

Animals↗