Long Term Liver Engraftment of Functional Hepatocytes Obtained from Germline Cell-Derived Pluripotent Stem Cells

One of the major hurdles in liver gene and cell therapy is availability of ex vivo-expanded hepatocytes. Pluripotent stem cells are an attractive alternative. Here, we show that hepatocyte precursors can be isolated from male germline cell-derived pluripotent stem cells (GPSCs) using the hepatoblast marker, Liv2, and induced to differentiate into hepatocytes in vitro. These cells expressed hepatic-specific genes and were functional as demonstrated by their ability to secrete albumin and produce urea. When transplanted in the liver parenchyma of partially hepatectomised mice, Liv2-sorted cells showed regional and heterogeneous engraftment in the injected lobe. Moreover, approximately 50% of Y chromosome-positive, GPSC-derived cells were found in the female livers, in the region of engraftment, even one month after cell injection. This is the first study showing that Liv2-sorted GPSCs-derived hepatocytes can undergo long lasting engraftment in the mouse liver. Thus, GPSCs might offer promise for regenerative medicine.     

Immobilization of lipoxygenase in an alginate-silicate solgel matrix: formation of fatty acid hydroperoxides

A method for the immobilization of lipoxygenase (LOX) in an alginate-silicate gel matrix was developed. In this method, a mixture of calcium alginate beads and LOX in borate buffer are dispersed into a hexane solution of tetramethoxy-ortho-silicate (TMOS). Hydrolysis of the TMOS gives products that permeate and co-polymerize with the alginate gel to form a colloid within the beads that entraps the LOX. Optimum reaction conditions for sol-gel entrapment of LOX are at pH 9.0 in 0.2M borate buffer. The composite gel, after isolation and vacuum drying, had excellent protein retention that has good enzyme activity and stability at room temperature. The activity of the entrapped LOX was less than the activity of the free enzyme. However, the activity of the immobilized LOX can be restored by the addition of borate buffer and glycerol, or borate buffer saturated with an organic solvent. In contrast to the free enzyme in solution, which loses its activity in less than one day, sol-gel entrapped LOX retains its activity at ambient temperature for at least 25 days and can be recycled. This report demonstrates that the sol-gel entrapment method for immobilizing LOX can be useful in developing a process for the oxidation of polyunsaturated fatty acids.     

Altered Intracellular Localization of SOD1 in Leukocytes from Patients with Sporadic Amyotrophic Lateral Sclerosis

Several lines of evidence support the hypothesis of a toxic role played by wild type SOD1 (WT-SOD1) in the pathogenesis of sporadic amyotrophic lateral sclerosis (SALS). In this study we investigated both distribution and expression profile of WT-SOD1 in leukocytes from 19 SALS patients and 17 healthy individuals. Immunofluorescence experiments by confocal microscopy showed that SOD1 accumulates in the nuclear compartment in a group of SALS subjects. These results were also confirmed by western blot carried out on soluble nuclear and cytoplasmic fractions, with increased nuclear SOD1 level (p<0.05). In addition, we observed the presence of cytoplasmic SOD1 aggregates in agreement with an increased amount of the protein recovered by the insoluble fraction. A further confirmation of the overall increased level of SOD1 has been obtained from single cells analysis using flow cytometry as cells from SALS patients showed an higher SOD1 protein content (p<0.05). These findings add further evidence to the hypothesis of an altered WT-SOD1 expression profile in peripheral blood mononuclear cells (PBMCs) from patients with ALS suggesting that WT-SOD1 species with different degrees of solubility could be involved in the pathogenesis of the disease.     

Further surveys of Pseliodinium vaubanii Sournia (Pirrophyta) in the Adriatic sea

Abstract

The presence, in the North Adriatic sea and in the Chioggia and Marano lagoons, of Pseliodinium vaubanii a rare dinoflagellate species, is reported. Our results showed that Pseliodinium vaubanii is a very frequent and abundant species in these environments.