Plasma Membrane-Associated Glycohydrolases Activation by Extracellular Acidification due to Proton Exchangers

In this paper, we show that the pH optimum for the plasma membrane (PM)-associated activity of four glycohydrolases (conduritol B epoxide sensitive β-glucosidase, β-glucosidase GBA2, β-hexosaminidase and β-galactosidase) measured on intact cells is acidic. Moreover, we show that drugs able to modify the efflux of protons across the PM, thus locally affecting the extracellular proton concentration close to the PM, are able to modulate the activities of these enzymes. These data strongly suggest that pH-dependent modulation of PM-associated glycohydrolases activities could be an effective way to locally modulate the cell surface glycoconjugate composition.     

Plasma Membrane-Associated Glycohydrolases Along Differentiation of Murine Neural Stem Cells

The activities of plasma membrane associated sialidase Neu3, total β-glucosidase, CBE-sensitive β-glucosidase, non-lysosomal β-glucosyl ceramidase GBA2, β-galactosidase, β-hexosaminidase and sphingomyelinase were determined at three different stages of differentiation of murine neural stem cell cultures, corresponding to precursors, commited progenitors, and differentiated cells. Cell immunostaining for specific markers of the differentiation process, performed after 7 days in culture in presence of differentiating agents, clearly showed the presence of oligodendrocytes, astrocytes and neurons. Glial cells were the most abundant. Sialidase Neu3 after a decrease from progenitors to precursors, showed an increase parallel to the differentiation process. All the other glycosidases increased their activity along differentiation. The activity of CBE-sensitive β-glucosidase and GBA2 were very similar at the precursor stage, but CBE-sensitive β-glucosidase increased 7 times while GBA2 only two in the differentiated cells. In addition, we analysed also sphingomyelinase as enzyme specifically associated to sphingolipids. The activity of this enzyme increased from precursors to differentiated cells.     

Evidence of host-associated populations of Cryptosporidium parvum in Italy

Recent studies have revealed extensive genetic variation among isolates of Cryptosporidium parvum, an Apicomplexan parasite that causes gastroenteritis in both humans and animals worldwide. The parasite's population structure is influenced by the intensity of transmission, the host-parasite interaction, and husbandry practices. As a result, C. parvum populations can be panmictic, clonal, or even epidemic on both a local scale and a larger geographical scale. To extend the study of C. parvum populations to an unexplored region, 173 isolates of C. parvum collected in Italy from humans and livestock (calf, sheep, and goat) over a 10-year period were genotyped using a multilocus scheme based on 7 mini- and microsatellite loci. In agreement with other studies, extensive polymorphism was observed, with 102 distinct multilocus genotypes (MLGs) identified among 173 isolates. The presence of linkage disequilibrium, the confinement of MLGs to individual farms, and the relationship of many MLGs inferred using network analysis (eBURST) suggest a predominantly clonal population structure, but there is also evidence that part of the diversity can be explained by genetic exchange. MLGs from goats were found to differ from bovine and sheep MLGs, supporting the existence of C. parvum subpopulations. Finally, MLGs from isolates collected between 1997 and 1999 were also identified as a distinct subgroup in principal-component analysis and eBURST analysis, suggesting a continuous introduction of novel genotypes in the parasite population.     

Accumulation and effects of sulfadimethoxine in Salix fragilis L. plants: a preliminary study to phytoremediation purposes

The application of manure to fertilize arable lands is one of the major means through which veterinary sulfonamides (SAs) enter the environment. Little is known about the capacity of woody plants to phytoremediate this class of antibiotics. To this purpose we performed preliminary studies to evaluate Salix fragilis L. response to sulfadimethoxine (SDM) by investigating both its ability to absorb and tolerate doses of SDM found in fresh faeces of treated calves. Forty cuttings were exposed to either 0, 0.5, 1, or 2 mM of SDM for one month. Decreases in photosynthetic electron transport rate and net CO2 assimilation after 25 days for the higher SDM concentrations were noticed. Moreover, alterations in root morphology of treated plants were observed and further investigated through electron microscopy. However, collected data revealed high root accumulation potential. These preliminary results are promising as they demonstrate that Salix fragilis L. can both absorb and tolerate high concentrations of SAs.     

Stem cell microvesicles transfer cystinosin to human cystinotic cells and reduce cystine accumulation in vitro

Cystinosis is a rare disease caused by homozygous mutations of the CTNS gene, encoding a cystine efflux channel in the lysosomal membrane. In Ctns knockout mice, the pathologic intralysosomal accumulation of cystine that drives progressive organ damage can be reversed by infusion of wildtype bone marrow-derived stem cells, but the mechanism involved is unclear since the exogeneous stem cells are rarely integrated into renal tubules. Here we show that human mesenchymal stem cells, from amniotic fluid or bone marrow, reduce pathologic cystine accumulation in co-cultured CTNS mutant fibroblasts or proximal tubular cells from cystinosis patients. This paracrine effect is associated with release into the culture medium of stem cell microvesicles (100-400 nm diameter) containing wildtype cystinosin protein and CTNS mRNA. Isolated stem cell microvesicles reduce target cell cystine accumulation in a dose-dependent, Annexin V-sensitive manner. Microvesicles from stem cells expressing CTNS(Red) transfer tagged CTNS protein to the lysosome/endosome compartment of cystinotic fibroblasts. Our observations suggest that exogenous stem cells may reprogram the biology of mutant tissues by direct microvesicle transfer of membrane-associated wildtype molecules.