The antigen-antibody interaction algebraically interpreted as a relational process.

The antigen-antibody interaction occurring previous to the triggering of the immunological response is analyzed as a relational process in terms of lattices. Accordingly, this process is expressed as a lattice belonging to a pseudo-Boolean algebraic variety. The Heyting arrow operation, which appears in this kind of algebra, is used to analyze behaviors between non-comparable biological states expressed by the lattice. The resulting states coming from the arrows are connected with the influence of increasing and decreasing energies involved in the linking process.     

DNA condensation by an oxidizable cationic detergent. Interactions with lipid vesicles.

Cationic amphiphile-mediated delivery of plasmid DNA is the non-viral gene transfer method most often used. In the present work, we considered a new cysteine-detergent, ornithinyl-cysteinyl-tetradecylamide (C(14)-CO), able to convert itself, via oxidative dimerization, into a cationic cystine-lipid. By using fluorescence techniques, we first characterized the structure of complexes of plasmid DNA with C(14)-CO molecules either kept as monomers, or oxidized into dimers. Both forms are able to condense DNA, with the formation of hydrophobic micelle-like domains along the DNA chain. Domains with a larger molecular order were obtained with dimeric C(14)-CO/DNA complexes. In a second step, the interactions of these complexes with lipid vesicles considered as membrane models were investigated. In the presence of vesicles, we observed a decondensation of the DNA involved in complexes obtained with C(14)-CO monomers. With anionic vesicles, the DNA is released into the bulk solution, while with neutral vesicles, it remains bound to the vesicles via electrostatic interactions with inserted C(14)-CO molecules. In sharp contrast, the complexes with C(14)-CO dimers are unaffected by the addition of either neutral or anionic vesicles and show no interaction with them. These results may partly explain the low transfection efficiency of these complexes at the +/-charge ratios used in this study.     

Second-harmonic generation from organic molecules incorporated in sol-gel matrices

Orientation of optically nonlinear organic molecules inside sol-gel matrices upon application of an external D.C. electrical field is demonstrated for the first time. The quadratic nonlinear response of silicon oxide or transition metal oxide based gels containing organic molecules has been determined from Electric Field Induced Second Harmonic (EFISH) measurements. Large concentrations of Optically Nonlinear Organic Molecules (ONOM) have been either incorporated inside the macromolecular network or chemically bonded to the oxide backbone of the gels. These results demonstrate the feasibility of permanently poled doped sol-gel matrices. Moreover, EFISH measurements performed on organic molecules appear to be a useful tool for monitoring the changes occurring during sol-gel transformations.     

De novo-synthesized ceramide signals apoptosis in astrocytes via extracellular signal-regulated kinase.

Recent observations support the importance of ceramide synthesis de novo in the induction of apoptosis. However, the downstream targets of de novo-synthesized ceramide are unknown. Here we show that palmitate incorporated into ceramide and induced apoptotic DNA fragmentation in astrocytes. These effects of palmitate were exacerbated when fatty acid breakdown was uncoupled and were not evident in neurons, which show a very low capacity to take up and metabolize palmitate. Palmitate-induced apoptosis of astrocytes was prevented by L-cycloserine and fumonisin B1, two inhibitors of ceramide synthesis de novo, and by PD098059, an inhibitor of the extracellular signal-regulated kinase (ERK) cascade. Accordingly, palmitate activated ERK by a process that was dependent on ceramide synthesis de novo and Raf-1, but independent of kinase suppressor of Ras. Other potential targets of ceramide in the control of cell fate, namely, c-Jun amino-terminal kinase, p38 mitogen-activated protein kinase, and protein kinase B, were not significantly affected in astrocytes exposed to palmitate. Results show that the Raf-1/ERK cascade is the selective downstream target of de novo-synthesized ceramide in the induction of apoptosis in astrocytes and also highlight the importance of ceramide synthesis de novo in apoptosis of astrocytes, which might have pathophysiological relevance.     

Familial breast/ovarian cancer and BRCA1/2 genetic screening: the role of immunohistochemistry as an additional method in the selection of patients.

Only 20-25% of families screened for BRCA1/2 mutations are found positive. Because only a positive result is informative, we studied the role of BRCA1/2 immunohistochemistry as an additional method for patient selection. From 53 high-risk-affected probands, 18 (34%) had available paraffin blocks of their tumors and were selected for this study. Mutation screening was done by conformation-sensitive gel electrophoresis and multiplex ligation-dependent probe amplification. For immunohistochemistry, 21 neoplastic specimens (15 breast carcinomas, 5 ovary neoplasms, and 1 rectal adenocarcinoma) were analyzed with BRCA1 (monoclonal antibody, Ab-1, oncogene) and BRCA2 (polyclonal antibody, Ab-2, oncogene) antibodies. Absence of the BRCA1 protein was confirmed in negative tumors by Western blotting. Seven patients were positive for BRCA1/2 mutations: 5 for BRCA1 and 2 for BRCA2. Four out of five positive patients had tumors negative for BRCA1 immunostaining, and the remaining 13 BRCA1-negative patients had positive BRCA1 immunostaining in all tumor samples. Sensitivity to predict for BRCA1 mutation carriers was 80%, and specificity was 100%, with a positive predictive value of 100% and a negative predictive value of 93%. This correlation was statistically significant (p=0.001). No correlation was observed for BRCA2. If larger studies confirm these results, high-risk patients with BRCA1-negative tumors should be screened first for this gene.     

Characterization of a monoclonal antibody to human proacrosin and its use in acrosomal status evaluation.

Among the monoclonal antibodies (MAb) selected after immunization of mice with a detergent-insoluble fraction from human spermatozoa, MAb 4D4 was found to stain in immunofluorescence the principal part of the acrosome of human spermatozoa. Acrosome reaction induced decreased and spotty 4D4 immunofluorescence staining. Immunoelectron microscopy before or after embedding revealed that the epitope defined by MAb 4D4 was sequestered in the anterior acrosomal matrix and, after the acrosome reaction, remained partly bound on matrix elements attached to the inner acrosomal membrane. Western blot analysis of sperm extracts showed that the epitope defined by MAb 4D4 was located on a 55 KD polypeptide in whole cells and on 55 and 50 KD polypeptides in non-ionic detergent fractions. Human proacrosin-enriched fraction obtained by FPLC purification exhibited several proteolytic activities against gelatin in gel enzymography: a 50 KD major band and two minor bands in the 20-30 KD area; the 50 KD polypeptide reacted with MAb 4D4 in Western blots. Furthermore, the 4D4-immunoprecipitated polypeptide from sperm extract showed that the 50 KD band exhibited proteolytic activity with an optimal pH at 8.0 that was strongly inhibited by soybean trypsin inhibitor and ZnCl2. MAb 4D4 also reacted with the acrosome of the monkey Macaca fascicularis but not with the acrosome of any of the other non-primate mammalian species examined so far. Various shape defects of the acrosomal principal region were revealed by 4D4 labeling of spermatozoa with head anomalies from infertile patients. MAb 4D4 also recognized proacrosin in paraffin-embedded human testis sections. These data make the monoclonal antiproacrosin antibody 4D4 an efficient tool for evaluation of the acrosomal status of human spermatozoa and spermatids.     

Antigenic study of the differentiation of the human kidney using monoclonal antibodies

The production of monoclonal antibodies against human embryonic renal cells allowed to display on the adult human kidney some antigens typical of certain structures or tissues: the proximal convoluted tubule for EG 9-11 and EG 19-6 monoclonal antibodies, the glomerular basement membrane for EG 14-1, the urothelium for EE 24-6, the connective tissue for EK 8-1 and EK 17-1 and probably the capsular and tubular basement membranes for EK 8-1. Simultaneously, we could follow the spatial and temporal repartition of the antigens during the renal development. One of them (EI 16-1) seemed to disappear in the adult and might correspond to a foetal type-antigen.     

Enzymes as molecular automata: a reflection on some numerical and philosophical aspects of the hypothesis.

Enzymes, by means of their properties of specific recognition and allosteric modulation, are able to integrate many separate processes into systemic units with coherent functions; in a sense, they have to be considered as the true organizers of the cytoplasmic processes. In this respect, the present article describes a simple model, based on binary variables and automata theory, which simulates the basic regulatory performance of the modulated enzyme. The model admits a variety of modifications and improvements; it also suggests some original lines of thought on which to reflect about the organization and collective phenomena of the networks of enzymes. In discussing the connection of this 'molecular automata' hypothesis with other areas of present-day theoretical biology, a fertile panorama of initiatives appear. A special partnership between Information Science (computation) and Biology is developing.     

An Na+-stimulated Mg2+-transport system in human red blood cells

The initial rate of net Mg2+ efflux was measured in human red blood cells by atomic absorption. In fresh erythrocytes incubated in Na+,K+-Ringer's medium this rate was 7.3 +/- 2.8 mumol/l cells per h (mean +/- S.D. of 14 subjects) with an energy of activation of 13 200 cal/mol. Cells with total Mg2+ contents ([ Mg]i) ranging from 1.8 to 24 mmol/l cells were prepared by using a modified p-chloromercuribenzenesulphonate method. Mg2+ efflux was strongly stimulated by increases in [Mg]i and in external Na+ concentrations ([ Na]o). A kinetic analysis of Mg2+ efflux as a function of [Mg]i and [Na]o revealed the existence of two components: an Na+-stimulated Mg2+ efflux, which exhibited a Michaelian-like dependence of free internal Mg2+ content (apparent dissociation constant = 2.6 +/- 1.4 mmol/l cells; mean +/- S.D. of six subjects) and on external Na+ concentration (apparent dissociation constant = 20.5 +/- 1.9 mM; mean +/- S.D. of four subjects) and a variable maximal rate ranging from 35 to 370 mumol/l cells per h, and an Na+-independent Mg2+ efflux, which showed a linear dependence on internal Mg2+ content with a rate constant of (6.6 +/- 0.7) X 10(-3) h-1. Fluxes catalyzed by the Na+-stimulated Mg2+ carrier were partially dependent on the ATP content of the cells and completely inhibited by quinidine (IC50 = 50 microM) and by Mn2+ (IC50 = 0.5-1.0 mM).