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Biomedical subjects

A Merli

Publications and source records attributed to A Merli.

At least 19 recordsLinked to original sources
PubMed1992

Structural study and preliminary crystallographic data for the hemoglobin from reindeer (Rangifer tarandus tarandus)

The ferric form of reindeer hemoglobin (Rangifer tarandus tarandus) has been crystallized in an orthorhombic crystalline form from polyethylene glycol solutions, at pH 8.2. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit cell edges a = 84.2 A, b = 59.9 A, c = 119.5 A; one hemoglobin tetramer is contained in the asymmetric unit. The crystals diffract X-rays to a limit spacing of 3.0 A. Inspection of amino acid sequences in the N-terminal region of beta-chains, and analysis of hemoglobin three-dimensional models, allows one to rationalize, on a molecular basis, the reduced O2 affinity and the decreased effect of organic phosphates observed in ruminant hemoglobins. By analogy, the analysis is extended to birds and reptiles, whose hemoglobin beta-chains display, as in ruminants, the deletion of the N-terminal residue and a methionine at the NA2 position.

2,3-Diphosphoglycerate↗
PubMed1991

Crystallization and preliminary data for the ferric form of Lucina pectinata hemoglobin I.

Cytoplasmic monomeric hemoglobin I from the bacteria-harboring gill of the bivalve mollusc Lucina pectinata has been crystallized in a form suitable for atomic resolution X-ray structural investigations. The crystals have been grown at pH 4.8, in 0.05 M-acetate buffer, using 2.6 M-ammonium sulfate as precipitating agent. The crystals belong to the monoclinic space group P2(1), with unit cell constants a = 50.0 A, b = 38.6 A, c = 42.1 A, beta = 107.1 degrees, and contain one molecule (14,000 Mr) in the asymmetric unit. By means of single crystal microspectrophotometry it has been shown that the crystals contain the ferric form of L. pectinata "sulfide reactive" hemoglobin I. On the other hand, by careful control of the buffering medium composition, it has been possible to obtain stable crystals of the deoxy, oxy and sulfide forms of the protein.

Animals↗
PubMed1991

Analysis at the clonal level of T-cell phenotype and functions in severe aplastic anemia patients.

The aim of this study was to analyze at the clonal level the phenotype and functions of T cells from patients with severe aplastic anemia (SAA). For this purpose we studied 175 T-cell clones obtained from peripheral blood (PB) and bone marrow (BM) of four SAA patients and 97 clones from two healthy controls. The percentage of CD8+ T-cell clones obtained from the patients' PB and BM was higher, but not significantly (P = .07 and P = .14, respectively), than that obtained in controls. A higher proportion of T-cell clones from SAA patients exhibited lectin-dependent cytolytic activity and especially natural killer-like activity when compared with controls (PB: P less than .01, P less than .05; BM: P less than .05, P less than .01, respectively). Lymphokine release was tested before and after mitogen stimulation. A number of patients' clones were able to release interferons (IFNs) spontaneously (PB: 28.6% v 0%, P less than .05; BM: 28.6% v 0%, P less than .10). After mitogen stimulation, patients' BM T-cell clones produced IFNs in greater proportions (90.9% v 46.7%, P less than .01) and in greater quantities (PB: 25.5 arbitrary units [AU]/mL v 5.7 AU/mL, P less than .03; BM: 26 AU/mL v 9.1 AU/mL, P = .011) as compared with controls. Tumor necrosis factor (TNF) activity was not found in supernatants of unprimed T-cell clones. After mitogen stimulation, PB T-cell microcultures produced TNF alpha in greater proportions (97.9% v 72.2%, P less than .01) and, also in this case, in greater quantities (PB: 7.2 AU/mL v 1.5 AU/mL, P = .007; BM: 9.9 AU/mL v 1.5 AU/mL, P = .003) than controls. In conclusion, T-cell clones from SAA patients exhibit predominantly a CD8+ phenotype, a greater cytotoxic activity, and can be shown to produce greater quantities of suppressor lymphokines when compared with controls.

Adolescent↗
PubMed1990

Identification of four subsets of human CD3-CD16+ natural killer (NK) cells by the expression of clonally distributed functional surface molecules: correlation between subset assignment of NK clones and ability to mediate specific alloantigen recognition.

In previous studies we identified a surface molecule (termed GL183) capable of mediating cell activation and selectively expressed by a subset of human CD3-CD16+ natural killer (NK) cells. In this study we analyzed whether other subset-specific functional molecules were expressed in GL183- NK cells. To this end, mice were immunized with the PE29 (CD3-CD16+GL183-) NK clone. Monoclonal antibodies (mAbs) were selected by screening the hybridoma supernatants for their ability to trigger the cytolytic activity of clone PE29 against the human myelomonocytic leukemia U937. The EB6 mAb (IgG1) triggered the PE29 clone, but not a GL183+ clone used as a control. EB6+ cells ranged between 1 and 13% of peripheral blood lymphocytes and were largely included in the CD3-CD16+CD56+ cell populations (only less than 2% of EB6+ cells were CD3+). Analysis of resting or activated CD3-CD16+ populations, or clones for the expression of EB6 or GL183 mAbs, allowed us to identify four distinct, phenotypically stable, NK subsets (EB6+GL183-; EB6+GL183+; EB6-GL183+; EB6-GL183-). Similar to GL183 mAb, the EB6 mAb selectively triggered the NK subset expressing the corresponding surface antigen to lyse human tumor cell lines including U937, IGROV-I, M14, and A549. In addition, EB6 mAb sharply inhibited the cytolytic activity of EB6+ clones against P815, M12, and P3U1 murine target cells. In EB6+GL183+ ("double-positive") clones both EB6 and GL183 mAb inhibited the redirected killing of P815 cells induced by anti-CD16, anti-CD2 mAbs and phytohemagglutinin (PHA). Similar to GL183 molecules, molecules precipitated by EB6 mAb were represented by either single 58-kD chain or double chains of 55 and 58 kD (with no detectable differences in EB6+GL183- or EB6+GL183+ clones). In sequential immunoprecipitation experiments using the double-positive clones CEG52 and CA25.50, preclearing of cell lysates with EB6 or GL183 mAb removed only EB6 or GL183 molecules, respectively, thus indicating that the two antigenic determinants are carried by two distinct molecules. Peptide map analysis indicated that EB6 (or GL183) molecules precipitated from double-positive clones were identical to the corresponding molecules isolated from single-positive ones. On the other hand, comparison of the EB6 and GL183 maps revealed peptides that were unique to each molecule, although most of the major peptides migrated to identical positions. We further investigated whether correlation existed between the phenotypic assignment of NK clones and their ability to mediate specific lysis of normal allogeneic cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1990

A novel surface antigen expressed by a subset of human CD3- CD16+ natural killer cells. Role in cell activation and regulation of cytolytic function.

The GL183 mAb was obtained by immunizing BALB/c mice with the E57 clone (CD7+CD2+CD3-CD16+CD56+) derived from human peripheral blood NK cells. In human peripheral blood, GL183-reactive cells ranged between 2 and 12% (mean 6.5%) in 10 different donors. Double fluorescence and FACS analysis showed that GL183+ cells were consistently included in the CD56+ or CD16+ cell populations. Moreover, since only a fraction of CD56+ or CD16+ cells (approximately 40%) coexpressed GL183 surface antigen, reactivity with GL183 mAb appears to define two subsets within the CD3- lymphocyte population expressing NK cell markers. Although, the majority of GL183+ cells were CD3-, approximately 1% expressed CD3 surface antigens. As shown by clonal analysis, these infrequent CD3+GL183+ cells coexpressed CD56 and CD16 antigens. Cloning of CD3-GL183+ or CD3-GL183- cell populations under limiting dilution conditions yielded clonal progenies that maintained their original surface phenotype. Therefore, expression or lack of expression of GL183 surface antigens represents a stable phenotypic property of a subset of human CD3- NK cells. Immunoprecipitation experiments and two-dimensional PAGE analysis indicated that GL183-reactive molecules were represented in different clones either by a single 58-kD chain or, more frequently, by two chains of approximately 55 and approximately 58 kD, respectively. Analysis of GL183+ or GL183- NK clones for their ability to lyse human (IGROV I) or murine (P815) tumor target cells indicated that GL183- clones were, on average, fivefold more efficient in inducing target cell lysis. GL183+ and GL183- clones produced comparable levels of TNF-alpha in response to PHA plus PMA or anti-CD16 mAb plus PMA. Importantly, production of TNF-alpha was also induced by stimulation of GL183+ clones with GL183 mAb plus PMA. These data indicated that GL183 antigen could mediate cell triggering. This concept was confirmed by the analysis of Ca2+ mobilization, as GL183 mAb induced (in GL183+ clones) increments of [Ca2+]i comparable with those induced by PHA. Moreover, GL183 mAb, or its F(ab')2 fragments, strongly enhanced the cytolytic activity of GL183+ clones against a panel of human tumor target cells, including U937, Raji, IGROV I, M14, and A549. In contrast, GL183 mAb, but not the F(ab')2 fragments, sharply inhibited the cytolytic activity of the same clones against P815, M12, and P3U1 murine target cells. In this case, the effect of GL183 mAb (inhibition) was opposite that of PHA or of stimulatory anti-CD2 or anti-CD16 mAbs, which consistently enhanced the target cell lysis.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1989

[Knowing pemphigus].

The classification, pathogenesis, clinical features and treatment of pemphigus are examined with emphasis' on the histopathological similarities of the elementary intraepithelial lesion and the histotopographical differences at epithelial level that differentiate the various types of pemphigus. In a detailed review of the pathogenesis of the condition, the circumstances propitious for the onset of pemphigus are identified as an HLA phenotype and the autoantigenic denaturation of the glycocalyx. The condition is described as vesicolobullous and is differentiated from similar bullous and non-bullous dermatoses. The most significant pictures of oral pemphigus are specified as are the basic elements of general and local treatment.

Autoimmune Diseases↗
PubMed1989

[Candida-host relationship: physiology and pathology].

The Authors give a detailed analysis of the potential virulence factors and possible pathogenic processes associated with candida. They qualify the interactions between local, general, natural and acquired defences, setting out the possible deficiency involved in the emergence of secondary candidiasis. They make a classificatory revision of forms of candidiasis on the basis of topographic, anatomo-pathologic criteria and of the course of the disease. They investigate the most significant aspects of the situations described especially in oral localization. An evaluation is made of chronic mucocutaneous candidiasis as a pathological affection in itself and in its correlation with H.I.V. infection.

Acquired Immunodeficiency Syndrome↗
PubMed1988

An in vitro model for Toxoplasma infection in man. Interaction between CD4+ monoclonal T cells and macrophages results in killing of trophozoites.

The cell interactions that take place between Toxoplasma gondii trophozoites and the human immune system have been investigated by using an in vitro model of infection. PBMC were co-cultured with live, appropriately attenuated, trophozoites. When cells from immune (seropositive) donors were used, a proliferative response was observed. At the same time, the proliferating T cells proved capable of controlling the growth of live trophozoites. By contrast, cells from seronegative donors failed to mount a proliferative response and intracellular overgrowth of trophozoites with subsequent cell injury occurred. Actively proliferating T cells were expanded in continuous cell lines with IL-2 and periodical restimulation with Ag in the presence of autologous irradiated mononuclear cells. From some of the lines obtained, clones were also derived. Ten clones were selected for further studies. They proliferated in response to trophozoites but not to unrelated Ag. Their response required the presence of autologous monocytes-macrophages isolated from peripheral blood on Percoll density gradients. B cells that were obtained from the same donors and immortalized by EBV infection proved inefficient as APC. These data suggest that live trophozoites have to be processed by macrophages in order to be presented to T cells. Upon appropriate antigen stimulation, all of the clones produced IL-2 and IFN-gamma, a finding that was consistent with both their CD4+ surface phenotype and their helper capacity on B cell proliferation and differentiation in vitro. The supernatants of all of the stimulated clones released a factor that activated macrophages to kill intracellular trophozoites as well as an unrelated pathogen, Listeria monocytogenes. This factor was identified as IFN-gamma because it was neutralized by specific anti-IFN-gamma antibodies. The present in vitro model of response to live protozoa may prove suitable to assess the role of both T lymphocytes and macrophages in intracellular parasite infections in man. Furthermore, this experimental system may be applied to detect specific lesions of cell mediated immunity in a number of immunodeficiency syndromes.

Adult↗
PubMed1986

Deacylation kinetics of gamma-chymotrypsin in solution and in the crystal.

The rate of catalyzed hydrolysis of the acyl enzyme analogs indolacryloyl-gamma-chymotrypsin and furylacryloyl-gamma-chymotrypsin in the crystal has been measured by single crystal microspectrophotometry and compared with the rate of catalyzed hydrolysis of acyl-gamma-chymotrypsin in solution and acyl-alpha-chymotrypsin both in solution and in the crystal. The maximal deacylation rate is the same for both species and independent of the physical state. However, the pH dependence of the deacylation rate of crystalline acyl-gamma-chymotrypsin shows a 0.9 unit shift in the pK of the catalytic system which is unique and probably consequent to specific lattice interactions.

Acylation↗
PubMed1986

T cell nature of some lymphokine-activated killer (LAK) cells. Frequency analysis of LAK precursors within human T cell populations and clonal analysis of LAK effector cells.

The cell lineage of the lymphokine-activated killer (LAK) cells has been reinvestigated. Both T and non-T cells, isolated on the basis of rosette formation with sheep erythrocytes (E), generated LAK activity after 3-4 days of culture in recombinant interleukin 2 (rIL 2) in 8 different individuals tested. By applying a microculture technique which allows clonal expansion of virtually all E rosetting T cells, we further analyzed the frequency of clonogenic LAK precursors within T cell populations. Approximately 1 of 25 T cells was found to be a LAK precursor. Moreover, microcultures with LAK activity lysed both the natural killer-sensitive K562 cell line and the P815 target cells in the presence of phytohemagglutinin (PHA). Since cytolytic T lymphocytes capable of lysing P815 cells in the PHA-dependent assay were approximately 1/3, it is evident that only a minor subset of cytolytic T lymphocyte precursors can acquire LAK activity even in the presence of large amounts of IL 2. Several LAK clones obtained by limiting dilution were further expanded and analyzed for their phenotypic and functional properties. Twelve out of 14 clones analyzed expressed the T3+ T11+ phenotype whereas 2 were T3- T11+. All had maintained their original cytolytic pattern; moreover, the large majority of the T3+ clones produced IL 2 and interferon-gamma following PHA stimulation.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1985

Enzyme immunoassay for evaluation of Toxoplasma gondii growth in tissue culture.

An enzyme-linked immunosorbent assay (ELISA) for the evaluation of growth kinetics of Toxoplasma gondii in tissue cultures was developed. Tissue culture microplates (96 wells) were seeded with Vero cells, infected with a virulent strain of T. gondii, and incubated for different time periods. The ELISA was performed with anti-T. gondii antibodies on the infected cells. The method was simple, rapid, and accurate, and very good correlations between the ELISA results and the percentage of T. gondii rosettes in infected cells, the number of free T. gondii in the supernatant, and the amount of T. gondii antigen in the supernatant were observed.

Animals↗
PubMed1985

In vitro production of different interferon types by cloned human NK cells.

Human peripheral blood null cells were conjugated in vitro with K-562 cells and expanded into continuous cell lines using IL-2 containing medium (CM) and periodical restimulation with phytohaemagglutinin (PHA). Most of these lines were made up of granular blasts expressing high natural killer (NK) activity. When analysed for different surface markers, the large majority of the blasts were E rosette+, T3+, Tac+, DR+, Leu7+ with a variable proportion of cells expressing T8 and M1 antigens (range: 20-80%). In contrast, T4 antigen was expressed by the majority of cells of the control cell lines originated in the absence of K-562 cells. Twenty-nine clones were obtained from one of the above lines using the limiting dilution technique and subsequently maintained in CM for 4 months or more. The majority of these clones maintained their cytotoxic potential and were able to produce different interferon (IFN) types (IFN-alpha, IFN-gamma or both) when growing in CM. In addition in a number of selected clones, simultaneous stimulation with PHA and K-562 cells was able to induce or support the production of both IFN types.

Antigens, Surface↗
PubMed1977

Polypeptides in the succinate-coenzyme Q reductase segment of the respiratory chain.

Complex II (succinate-coenzyme Q reductase) was resolved into ten different polypeptides by polyacrylamide gel electrophoresis. Four polypeptides, CII-1, CII-2, CII-3, and CII-4 with molecular weights of 70 000, 24 000, 13 500, and 7000, were present in large amounts in all preparations examined. CII-1 and CII-2 are the flavoprotein and iron-sulfur protein, respectively, of succinate dehydrogenase; CII-3 and CII-4 have not been functionally indentified. Six polypeptides were present in much smaller amoumts as judged by staining intensity, and each of these comigrated with components in complex III. The amino acid compositions of several of the minor components in complex II were identical with that of an equivalently migrating polypeptide in complex III. We conclude that succinate-coenzyme Q reductase contains four different polypeptides and is contaminated with variable amounts of complex III when isolated as complex II.

Amino Acids↗
PubMed1976

[Frontal vertebral cleft in the newborn].

In the newborn and foetus in the last months of gestation it is possible to see in lateral radiography of the spine, a longitudinal radiotransparent band which may divide the bone nucleus of the vertebral body into two portions. This front vertebral separation should be considered an expression of a reversible abnormality rather than a malformation to the extent that it almost always concerns a differentiation delay in the vertebral structures. The literature and personal cases show, in fact, that the finding disappears more or less rapidly.

Diagnosis, Differential↗