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Biomedical subjects

B Rossi

Publications and source records attributed to B Rossi.

At least 127 records · Page 7Linked to original sources
PubMed1995

Myophosphorylase deficiency affects muscle mitochondrial respiration as shown by 31P-MR spectroscopy in a case with associated multifocal encephalopathy.

We report here a glycogen storage myopathy type V associated with multifocal encephalopathy. The patient, a 43-year-old male with increased serum CK, a heavy drinker and smoker, had been affected by generalized epilepsy since age 24, after a cranial injury. He had had a right hemiparesis 2 years before coming to our observation and a transient left hemiparesis the following year. CT and MRI of the brain showed multiple hemispheric lesions consistent with an ischemic process, as suggested by single photon emission tomography of the brain. Muscle biopsy showed a vacuolar myopathy, and myophosphorylase activity was 13% of the normal mean. Phosphorus magnetic resonance spectroscopy (31P-MRS) performed on resting calf muscles showed increased PCr to ATP and decreased PCr to P(i) ratios. During both aerobic and ischemic exercise 31P-MRS failed to show any cytosolic acidification and phosphomonoesters (PME) accumulation, two MRS findings in agreement with McArdle's syndrome diagnosis. Mitochondrial respiration was also affected as shown by a low PCr to P(i) ratio at rest and by a low rate of PCr re-synthesis during recovery from aerobic exercise. This latter finding in McArdle's disease can be explained by decreased mitochondrial substrate availability, which in turn can contribute to the phenotypic manifestations of the disease.

Adult↗
PubMed1995

Modulation of electrically elicited blink reflex components by visual and acoustic prestimuli in man.

The effects of a prestimulus on the electrically elicited blink reflex components were investigated in 20 healthy subjects. In the first group of 10 subjects (warned group), electric shocks were delivered in isolation or preceded, at an interstimulus interval (ISI) of 0.1 s, 1 s, or 10 s, by a visual or acoustic warning stimulus. In the second group of 10 subjects (unwarned group), the electric shocks were delivered either in isolation or preceded, at the same ISI, by visual or acoustic stimuli having no warning value. The modulation of the three blink reflex components was then analysed. Compared to the baseline condition, the R1 oligosynaptic component was enhanced at 0.1 s and 1 s ISI, in the warned group with the visual prestimulus, but only at 0.1 s after a visual and acoustic prestimulus in the unwarned group. On the contrary, the polysynaptic responses showed a different course: R2 was significantly reduced at the 0.1 s interval in the warned group with both the prestimuli, and only with the visual prestimulus in the unwarned group. The R3 was inhibited at all three intervals with the visual prestimulus, and at the 0.1 s and 1 s with the acoustic one in the warned group, and only at 0.1 s in the unwarned group, both after visual and acoustic prestimuli. The decrement in R2 and R3 observed with the shortest interval was probably related to the prepulse inhibition of startle reflex. Furthermore, only R3 was still inhibited at longer intervals, when the sustained processes of attention may have influenced this component. Perhaps this combination of events represents, in the warned group, the best preparation for voluntary reflex reaction.

Acoustic Stimulation↗
PubMed1995

Expression of the antigen recognized by mAb GB36 in normal skin and in skin tumours.

GB36, a mouse monoclonal antibody (mAb) raised against an epithelial antigen of the human trophoblast, reacts with the epithelial basement membrane of chorionic villi; it does not react with the invasive extravillous cytotrophoblast. Expression and characterization of the antigen of GB36 (designated GBA36) were investigated in normal keratinocytes by immunoprecipitation, immunofluorescence and immunoelectron microscopic studies. Immunoprecipitation experiments demonstrated that the proteins identified on keratinocytes by mAb GB36 and a rat mAb anti-integrin alpha 6 (GoH3) were the same. Using immunofluorescence and immunoelectron microscopic methods, GBA36 was localized on the cell membrane facing the epithelial basal lamina of basal keratinocytes. GBA36 distribution in benign and malignant skin tumours was evaluated by immunostaining methods (immunofluorescence and immunoperoxidase). Analysis of tumours revealed that whereas benign epithelial tumours and intradermal naevi displayed high levels of GBA36, the expression of this antigen decreased progressively in spinocellular and basal cell carcinomas, and in cutaneous melanomas in relation to invasiveness. During cell transformation, GBA36 undergoes quantitative alterations, and expression is down-regulated. Although the functional relevance of these changes remains unknown, the correlation of decreased GBA36 expression with tumour progression may indicate a role for altered integrin expression in tissue invasion by human skin carcinoma and melanoma.

Adult↗
PubMed1995

The cytoplasmic tail of CD4 is required for inhibition of human immunodeficiency virus type 1 replication by antibodies that bind to the immunoglobulin CDR3-like region in domain 1 of CD4.

Monoclonal antibodies (MAb) directed against the immunoglobulin complementary determining region 3 (CDR3)-like region of the CD4 molecule inhibit human immunodeficiency virus type 1 (HIV-1) transcription. We report here data showing that the cytoplasmic tail of CD4 is required for such inhibition to be achieved. To this aim, we studied the effect of MAb 13B8-2 treatment on (i) HIV-1 production in A2.01 cells, which express different forms of the CD4 gene, (ii) Tat-induced HIV-1 promoter activation, and (iii) mitogen-activated protein kinase (MAPK) activation, which is induced in CD4-positive cells by HIV-1 cross-linking of CD4. Inhibition of HIV production by 13B8-2 MAb treatment was consistently observed in cells expressing wild-type CD4 and cells expressing a hybrid CD4-CD8 molecule (amino acids 1 to 177 of CD4 fused to the hinge, transmembrane, and cytoplasmic domains of CD8). However, no delay in HIV-1 production was observed in cells expressing a truncated CD4 which lacks the cytoplasmic domain (CD4.401). Chloramphenicol acetyltransferase assays demonstrated that Tat-dependent activation of the HIV-1 long terminal repeat promoter was inhibited by MAb 13B8-2 in A2.01/CD4 and A2.01/CD4-CD8 but not in A2.01/CD4.401 cells. Finally, we found that MAb 13B8-2 treatment inhibited the activation of MAPK induced in A2.01/CD4 and A2.01/CD4-CD8 following cross-linking of CD4 by HIV-1.

Antibodies, Monoclonal↗
PubMed1995

[The scintigraphic prediction of residual lung function after lobectomy in patients with bronchial carcinoma].

The scintigraphic prediction of residual pulmonary function after pneumonectomy has been validated in a number of studies while scintigraphy is not standardized in case of lobectomy. This study was aimed at investigating the accuracy of the scintigraphic prediction of post-lobectomy lung function using Wernly method. We examined 43 patients with bronchial carcinoma: 20 of them underwent pneumonectomy and 23 underwent lobectomy. The pulmonary function data (vital capacity, CV, and forced expiratory volume in one second, VEMS) predicted by quantitative lung scan were compared with those observed in the postoperative follow-up. A good correlation between predicted and observed data was obtained in both the pneumonectomized group (r = 0.77 and 0.78 for CV and VEMS, respectively; p < 0.005) and the lobectomized group (r = 0.74 and 0.79 for CV and VEMS, respectively: p < 0.005). It can be concluded that the method used for the scintigraphy prediction of post-lobectomy pulmonary function is as accurate as the post-pneumonectomy method and can be used reliably in the clinical practice.

Aged↗
PubMed1995

Use of metaiodobenzylguanidine (MIBG) in pneumology.

Norepinephrine (NE) is one of the many biologically active substances that are metabolized by the lung. The norepinephrine uptake in the lung has been widely investigated in animal studies in order to detect abnormalities of the endothelial cells. 131I- or 123I-labelled metaiodobenzylguanidine (MIBG) is a radioactive tracer that shares several metabolic pathways with NE; it has been used instead NE or both in animal and in human studies for the same purposes. It has been definitely demonstrated that MIBG lung uptake following intravenous administration is a reliable marker of minimal endothelial cell lesions and it allows one to investigate several conditions involving damage to the lung vasculature such as adult respiratory distress syndrome, post-actinic lung fibrosis or chemotherapic lung toxicity. MIBG could also potentially be used as a tracer of the neuroadrenergic system of the airways, once it has been deposited in the whole branches of the bronchial tree. With this in mind we evaluated the feasibility of the administration of MIBG in radioaerosol form. We first performed a chromatographic study on [123I]MIBG aliquots aerosolized for variable lengths of time (2-30 minutes). Radiochemical purity was always over 96%, thus demonstrating that aerosolization does not significantly modify the stability of the tracer. We then compared both the lung clearance curves and the scintigraphic images in two groups of normal subjects who respectively underwent [123I]MIBG and 99mTc-DTPA radioaerosol dynamic scintigraphy. We found that MIBG clearance to be significantly lower than that of DTPA (135 +/- 32 minutes versus 69 +/- 27 minutes, p < 0.01), probably due to its being trapped in the adrenergic nerve endings of the airways. The pulmonary distribution of both tracers was homogeneous, as demonstrated by the correspondence of the values for the aerosol penetration index in both groups of subjects. Such results warrant further research and pharmacologic tests.

3-Iodobenzylguanidine↗
PubMed1994

Production and characterization of monoclonal and polyclonal antibodies to human alpha 2-HS: development of a two-site ELISA test.

A convenient and sensitive indirect sandwich ELISA test was developed for measuring both 63 kDa human alpha 2-HS secreted by human hepatoma cell lines and the 59 kDa alpha 2-HS species present in serum/plasma. Monoclonal and rabbit antibodies to plasma alpha 2-HS were produced and selected by immunoprecipitation techniques using iodinated alpha 2-HS or 35S-labeled alpha 2-HS. Various monoclonal antibodies recognizing both forms of the protein were coated onto microtiter plates and after binding of alpha 2-HS, biotinylated monoclonal antibodies with compatible binding or biotinylated immunopurified F(ab')2 fragments from the rabbit antiserum were added and subsequently revealed with avidin-biotin peroxidase complex. Formats using a rabbit detector antibody were the most sensitive and one was selected for the whole study. The test developed was capable of detecting plasma alpha 2-HS devoid of connecting peptide and HepG2 hepatoma cell line derived alpha 2-HS at the ng/ml level. The test has been used to measure levels of alpha 2-HS in both serum and supernatants from HepG2 cell lines.

Animals↗
PubMed1994

Insulin and interleukin-1 differentially regulate pp63, an acute phase phosphoprotein in hepatoma cell line.

We have reported previously that the phosphoprotein pp63, an acute phase protein, which has been recently identified as the rat fetuin, was capable of blocking the mitogenic effect of insulin on the rat Fao hepatoma cell line, without affecting metabolic effects of the hormone. Only the phosphorylated form of the protein has been shown to exhibit both anti-tyrosine kinase and growth inhibitory properties. In this study, we used the FTO-2B rat hepatoma cell line to analyze the mechanisms involved in the control of synthesis and/or phosphorylation of pp63. For this purpose, we investigated the action of effectors known to modulate hepatic functions, such as cytokines (interleukin (IL)-1 beta and IL-6), which regulate the production of acute phase proteins, and insulin, which elicits profound effects on hepatocyte metabolism. Here, we demonstrate that IL-1 beta diminished markedly the pp63 production by affecting its mRNA transcription and that the cytokine was able to modify the N-glycosylation process of the protein. In contrast, insulin did not affect the biosynthesis of pp63 but dramatically decreased its extent of phosphorylation.

Acute-Phase Proteins↗
PubMed1994

Monomeric 55-kDa guanidinobenzoatase switches to a serine proteinase activity upon tetramerization. Tetrameric proteinase SP 220 K appears as the native form.

Guanidinobenzoatases are cell surface enzymes present in cells capable of migration or remodeling. The guanidinobenzoatase purified to homogeneity from human renal carcinoma did not display gelatinase activity under the 55-kDa form (Poustis-Delpont, C., Descomps, R., Auberger, P., Delque-Bayer, P., Sudaka, P., and Rossi, B. (1992) Cancer Res. 52, 3622-3628). We bring new insights into the structure-activity relationships of this enzyme using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, [3H]diisopropyl fluorophosphate labeling, gelatin zymography, and immunodetection using a polyclonal antibody raised against the 55-kDa entity. Upon aggregation into a 220-kDa form, the enzyme exhibited [3H]diisopropyl fluorophosphate labeling and diisopropyl fluorophosphate-inhibitable gelatinase activity whereas its capability to cleave p-nitrophenyl p'-guanidinobenzoate as a substrate was abolished. Thus, the guanidinobenzoatase property appears as a feature of a 55-kDa inactive form of a serine proteinase subunit. After boiling in the presence of sodium dodecyl sulfate (3% w/v), the 220-kDa entity subjected to SDS-polyacrylamide gel electrophoresis could be dissociated into a 55-kDa protein as shown by silver staining. The resulting 55-kDa band remained [3H]diisopropyl fluorophosphate-labeled and reacted with anti-55-kDa guanidinobenzoatase antibodies, strongly suggesting that the 220-kDa proteinase was a noncovalently associated tetramer. Interestingly, Triton X-100 extracts of renal carcinoma plasma membranes exhibited a 220-kDa serine proteinase activity, as expressed in gelatin zymography, which was barely detectable in the non-tumoral counterpart. It is noteworthy that an anti-55-kDa guanidinobenzoatase reactive 220-kDa species was also observed in renal carcinoma plasma membranes extracts as assessed by Western blot, whereas it was hardly visible in the non-tumoral counterpart. No signal was immunodetected at M(r) 55,000 in renal carcinoma and kidney cortex membranes in Western blot experiments. Taken together, our data support the idea that the enzyme is expressed under its tetrameric form in the membrane. The purified enzyme is able to exhibit a serine proteinase activity when it recovers its native tetrameric form. This high molecular weight tetrameric proteinase SP 220 K appears as a new member of the cell surface serine protease family.

Animals↗
PubMed1994

Immunofluorescent quantification of tyrosine phosphorylation of cellular proteins in whole cells by flow cytometry.

Tyrosine phosphorylation of proteins, a major event in the transduction of mitogenic signals, was analysed by flow cytometry with a fluorescent antiphosphotyrosine monoclonal antibody, on formaldehyde-fixed, permeabilized cells. We have used this method (PY-Facs) to study activation of normal human T lymphocytes and cells of a leukemic T-cell line: Jurkat. In contrast to normal T cells, Jurkat cells as well as three other leukemic cell lines display a higher constitutive level of tyrosine phosphorylation. This level of tyrosine phosphorylation results from an equilibrium that can be up-regulated by the tyrosine phosphatase inhibitor, vanadate peroxide, and down-regulated by the tyrosine kinase inhibitors, genistein and staurosporine. We have also observed an increased tyrosine phosphorylation of proteins after mitogenic stimulation of Jurkat cells via T-cell receptor triggering. In addition, the entry of normal purified T cells from G0 phase into the cell cycle after co-stimulation with a phorbol ester and an anti-receptor antibody is correlated with a pronounced increase in tyrosine phosphorylation. We thus confirmed that this biochemical event was tightly associated with the activation status of the cells. The rapidity and sensitivity of the method we describe here make it particularly convenient for routine use and processing of a large number of samples, e.g., during analysis of human tumors. Moreover, because it retains sufficiently the integrity of treated cells and does not alter expression of membrane antigens, this method is suitable for multiparametric analysis, particularly for simultaneous studies associating the measure of tyrosine phosphorylation levels with possible modifications of membrane or intracellular structures as well as with cell cycle status.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1994

Thrombin and thrombin receptor agonist peptide induce early events of T cell activation and synergize with TCR cross-linking for CD69 expression and interleukin 2 production.

Thrombin stimulation of the T leukemic cell line Jurkat induced a transient increase in [Ca2+]i. Proteolytic activity of the enzyme was required for this effect since diisopropyl fluorophosphate-thrombin failed to increase [Ca2+]i. Furthermore, hirudin and anti-thrombin III inhibited the thrombin-induced [Ca2+]i rise in Jurkat T cells. A synthetic thrombin receptor agonist peptide (TRP) of 7 residues (SFLLRNP) was found to be as effective as thrombin for [Ca2+]i mobilization, and both agonists induced Ca2+ release exclusively from internal stores. Thrombin stimulated tyrosine phosphorylation of several proteins of molecular mass 40, 42, 70, 120, and 130 kDa. There was a good correlation between thrombin-induced tyrosine phosphorylation of the latter three proteins and Ca2+ mobilization. Thrombin and TRP also caused translocation of protein kinase C from the cytosol to the plasma membrane. As a likely consequence of these events, thrombin activated the nuclear factor NF-kB. Several cell lines of hematopoietic origin including the leukemic T cell line HPB.ALL and the erythroleukemic cell line K562 were responsive to thrombin, whereas others such as THP1, a myelomonocytic cell line, and BL2, a Burkitt lymphoma were refractory to thrombin or TRP stimulation. The magnitude of the thrombin response in the different cell types paralleled the expression of the thrombin receptor mRNA. We found that activation of Jurkat T cells by a combination of phytohemagglutinin and phorbol 12-myristate 13-acetate led to a dramatic inhibition of thrombin receptor mRNA expression and to a concomitant loss of the thrombin response. Finally, we demonstrate that thrombin and TRP enhanced CD69 expression and interleukin 2 production induced by T cell receptor cross-linking in both Jurkat T cells and peripheral blood lymphocytes. These findings highlight the role of thrombin as a potential regulator of T lymphocyte activation.

Amino Acid Sequence↗
PubMed1994

Induction of tyrosine phosphorylation and T-cell activation by vanadate peroxide, an inhibitor of protein tyrosine phosphatases.

Rapid tyrosine phosphorylation of key cellular proteins is a crucial event in the transduction of activation signals to T-lymphocytes. The regulatory role of protein tyrosine phosphatases (PTPases) in this process was explored by studying the effects of a powerful PTPase inhibitor, vanadate peroxide (pervanadate), on the activation cascade of Jurkat human leukaemic T-cells. Pervanadate induced activation of the tyrosine kinases lck and fyn (4- and 3-fold respectively) and a dramatic increase in tyrosine phosphorylation of cellular proteins, notably phospholipase C gamma 1. After this event, we observed a rise in intracellular Ca2+ concentration, corresponding to an influx. This effect required surface expression of the CD45 PTPase and was not observed in CD45-deficient variants of Jurkat cells. In the CD45-negative variant, the effect of pervanadate on tyrosine phosphorylation was globally decreased and some phosphorylated substrates were specifically missing. Pervanadate also stimulated transcription of the c-fos gene and accumulation of its mRNA as well as several other hallmarks of T-lymphocyte activation such as surface expression of the CD69 antigen and the interleukin 2 receptor alpha-chain (CD25). Pervanadate synergized with signals delivered by T-cell antigen receptor engagement or by a phorbol ester to induce interleukin 2 production. Pervanadate activated NF-kappa B, as shown by an increase in DNA-binding activity of this transcription factor. We thus conclude that PTPases play a crucial role in the negative regulation of signal transduction culminating in T-lymphocyte activation. Moreover, induction of tyrosine phosphorylation appears sufficient per se to initiate a complete activation programme.

Base Sequence↗
PubMed1994

Expression of the complement regulatory protein CD59 on human spermatozoa: characterization and role in gametic interaction.

Protectin (CD59) is a complement regulatory protein which blocks the membrane attack complex during complement activation. CD59 was identified on the human sperm surface by means of H19, an IgG1 anti-protectin mouse monoclonal antibody. Using indirect immunofluorescence, flow cytometry and immunoperoxidase, CD59 was found to be present on the whole plasma membrane including the head and tail of fresh ejaculated, capacitated and acrosome-reacted spermatozoa. Immunoperoxidase staining of normal testicular sections indicated that this protein was already present on intraluminal germ cells. Analysis of this sperm protein by gel electrophoresis and immunoblotting revealed that its molecular weight of 20 kDa was comparable to that of CD59 expressed on peripheral blood cells (erythrocytes, lymphocytes) and that it was bound to the membrane through a glycophospholipid tail which could be released after treatment with phosphatidylinositol-specific phospholipase C. Associated to membrane cofactor protein (CD46) and decay accelerating factor (CD55) located in the acrosomal membranes, CD59 may participate to the protection of male gametes against complement-mediated damage as they travel through the female genital tract. Moreover CD59, known as an adhesion molecule involved in lymphocyte rosettes, may also participate in cell to cell adhesion during gametic interaction since H19 inhibited sperm binding and reduced the penetration rate and index during the hamster egg penetration test.

Animals↗
PubMed1994

High expression of the antigen recognized by the monoclonal antibody GB24 on human breast carcinomas: a preventive mechanism of malignant tumor cells against complement attack?

GB24 is a mouse monoclonal antibody raised against a common trophoblast-lymphocyte cross-reactive antigen. GB24 detects the membrane cofactor protein (MCP, CD46), a member of the complement regulatory protein family, which serves as a cofactor for factor 1 mediated cleavage of C3b. This study investigated the reactivity of GB24 on 38 breast carcinomas and 34 normal/benign breast tissues by immunochemistry as well as the reactivity of F2B7-2, an antibody specific to the decay accelerating factor (DAF, CD55) of the complement. GB24 staining was present on both normal tissue and benign lesions, but very strong diffuse reactivity was observed on carcinomas. This reactivity increased with the tumor grade. By contrast, malignant tumor cells lacked DAF expression. F2B7-2 antibody reacted weakly with benign epithelial cells. Results were studied by computer assisted image analysis to accurately define the mean optical densities. The densitometric analysis of MCP positive carcinomas showed a high intensity of the staining. Expression of MCP and DAF on MCF-7 cell lines was analyzed by flow cytometry. MCF-7 cell lines were strongly stained by mAb GB24 only. These data suggest that selectively enhanced expression of the antigen recognized by GB24 is associated with malignant breast disorders. This high expression, which may reflect a protective mechanism by which tumor cells survive complement activation, may prove useful as a marker of malignant transformation.

Adenocarcinoma↗
PubMed1994

Orienting of attention in deafferented patients.

Visual attentional processes were compared in two deafferented patients and 11 normal subjects. Two consecutive stimuli were presented in rapid succession in one of two locations. A peripheral cue first oriented attention to one location where a response was requested. After 100 msec, a second response was required at either the same or opposite location (valid vs invalid cue). Four probabilities of valid cue occurrence were presented: 100, 80, 50 and 20%. Results showed (1) faster reaction times for the second response on cued than on uncued signals; (2) greater attentional effects with increased cue probability; (3) smaller attentional effects in patients. These findings suggest that the patients adopted a cost-minimizing strategy.

Adult↗