Flow cytometric approach to the study of erythrophagocytosis: evidence for an alternative immunoglobulin-independent pathway in agammaglobulinemic mice

Neuraminidase treatment of red blood cells (RBCs) is believed to induce changes similar to RBC senescence, and leads to a rapid clearance of RBCs from the circulation in vivo. The objective of this study using immunodeficient SCID mice and the lipophilic fluorescent probe PKH-26 was to ascertain whether antibodies are required as the final signal allowing the phagocytosis of neuraminidase-treated murine RBCs. All of the methods we applied are based on flow cytometry analysis using fluorescent probes: fluoresceinyl isothiocyanate (FITC)-labeled lectins for membrane carbohydrate identification and PKH-26-labeled RBCs for in vitro phagocytosis and in vivo clearance studies. The results can be summarized as follows: (i) the rate of neuraminidase-induced desialylation of RBCs from normal and immunodeficient mice is identical as ascertained with FITC-labeled lectins (wheat germ agglutinin (WGA) and Ricinus communis agglutinin (RCA(120))); (ii) the rate of clearance of enzyme-treated RBCs from both types of mice is also similar, as is their localization in spleen, liver and lung; (iii) the rates of in vitro phagocytosis of untreated and neuraminidase-treated PKH-26-labeled RBCs from both species of mice are very similar in the presence of homologous sera. In the absence of serum or in the presence of heterologous sera, the rate of phagocytosis is markedly decreased but not totally abolished. These data suggest that neuraminidase-treated RBCs can be cleared via an alternative pathway that is antibody-independent. This pathway exists in immunocompetent mice but with a very low activity and is the only one active in immunodeficient mice. In accordance with results reported by Connor et al. [J. Biol. Chem. 269 (1994) 2399], it is possible that this antibody-independent mechanism is involved in the clearance of circulating senescent RBCs. Finally, the methods described here may also be of interest for the investigation of the mechanisms involved in the phagocytosis of apoptotic cells.     

Deletion of the mental retardation gene Gdi1 impairs associative memory and alters social behavior in mice

Non-specific mental retardation (NSMR) is a common human disorder characterized by mental handicap as the only clinical symptom. Among the recently identified MR genes is GDI1, which encodes alpha Gdi, one of the proteins controlling the activity of the small GTPases of the Rab family in vesicle fusion and intracellular trafficking. We report the cognitive and behavioral characterization of mice carrying a deletion of Gdi1. The Gdi1-deficient mice are fertile and anatomically normal. They appear normal also in many tasks to assess spatial and episodic memory and emotional behavior. Gdi1-deficient mice are impaired in tasks requiring formation of short-term temporal associations, suggesting a defect in short-term memory. In addition, they show lowered aggression and altered social behavior. In mice, as in humans, lack of Gdi1 spares most central nervous system functions and preferentially impairs only a few forebrain functions required to form temporal associations. The general similarity to human mental retardation is striking, and suggests that the Gdi1 mutants may provide insights into the human defect and into the molecular mechanisms important for development of cognitive functions.     

The sensitivity of posturographic parameters to acquisition settings

The objective of this study was to evaluate the sensitivity of posturographic parameters (PP) to changes in acquisition settings. A group of eight young adults underwent a set of typical orthostatic posture trials, and selected PP were then calculated from a set of centre of pressure (CoP) displacement time series obtained by applying different cut-off frequencies to the same set of raw data. Four PP out of 11 showed significant changes with respect to cut-off frequency. Statistical mechanics parameters exhibited smaller sensitivity than summary measures. On the basis of the results obtained, a proposal for a standard cut-off frequency and a sampling rate value is embodied in the paper together with some suggestions on measurement settings, with a view to standardized use of instrumentation for quantitative analysis in orthostatic posturography.     

Allergologic exploration of germins and germin-like proteins,a new class of plant allergens

BACKGROUND: Germins and the related germin-like proteins (GLPs) are glycoproteins expressed in many plants in response to biotic and abiotic stress. To test the potential impact of germins and GLPs, recombinant germin from Triticum aestivum (tGermin) and GLPs from Arabidopsis thaliana (tGLP), both produced in transformed tobacco plants, were used. METHODS: Sera from 82 patients with type I allergy to birch, grass or mugwort pollen and/or wheat were tested in immunoblot for IgE binding to tGermin and tGLP, and the IgE reactivity after chemical and enzymatic deglycosylation was analysed. The biological activity of tGermin and tGLP was determined in a histamine release assay and in skin prick testing (SPT). RESULTS: In an immunoblotting assay, 24 out of 82 tested sera (29.26%) from allergic patients showed IgE-binding to tGermin, and 18 of these sera (21.95%) displayed also IgE-binding to tGLP. The deglycosylation experiments indicated that glycan moieties contribute significantly to the IgE-binding of tGermin and tGLP. Both tGermins and tGLP induced specifically histamine release in an in vitro assay as well as in SPT. CONCLUSION: Our in vitro and in vivo findings demonstrate that germin and GLPs are capable to bind IgE most likely via carbohydrate determinants, and represent allergenic molecules.     

Über die Grenzbedingungen von Zelleben und spezifischer Zellfunktion am isolierten Muskel

Zusammenfassung 1. Es werden an quergestreiften Muskeln (Zwerchfell, Herzvorhof von Warmblütern, Froschherz) unter den Bedingungen von Substratund Sauerstoffmangel Funktion und histologischer Zustand verglichen.2. Drosselung der Energiezufuhr führt in jedem Falle zu negativer Inotropie und diastolischem Stillstand. Dies führt zu dem zwingenden Schluß, daß die Kontraktion und nicht die Erschlaffung den energieverbrauchenden Teil des Kontraktionsvorganges darstellt.3. Gleichzeitiger Glucose- und Sauerstoffmangel führt erst mit dem Erlöschen der Glykogenolyse zur Kontraktur, die nach völligem Glykogenschwund irreversibel wird (Totenstarre).4. Die Totenstarre wird als Modellkontraktion abgelehnt.5. Aus den untersuchten Mangelbedingungen läßt sich eine Reihenfolge der Energieausnützung für den quergestreiften Muskel ableiten, wobei wesentliche Unterschiede zwischen Warmblüter und Kaltblüter bestehen.Mit 7 Textabbildungen     

Immunohistochemically detectable p53 and mdm-2 oncoprotein expression in colorectal carcinoma: prognostic significance

Aims—To investigate the correlation between the expression of the p53 and mdm-2 oncoproteins and to assess their prognostic value in colorectal cancer.