Ultrastructural features of cocoon-shell globules in the vitelline cells of neoophoran platyhelminths

The Neoophora is characterized by the presence of complex female gonads composed of both germaria and vitellaria. The vitellaria are made up of vitelline cells that differentiate to produce and accumulate reserve substances (proteins, lipid, glycogen) and precursors of the egg capsule or cocoon shell (phenolic proteins). A comparative ultrastructural and cytochemical investigation of shell-forming globules from the vitelline cells of some neoophoran platyhelminths shows that the internal structures of the shellforming globules can be grouped into two (or three) main types. In the first, globule content is characterized by a more or less homogeneous electron-dense substructure evidently arising from repeated coalescence of small vesicles produced by the rough endoplasmic reticulum and Golgi complex. A variant of this type of globule shows intermingling (or concentric) electron-dense and translucent areas producing a pattern resembling brain convolutions (convoluted pattern). The second type of shell-globule structure shows a multigranular pattern, presumably resulting from repeated fusions of Golgian vesicles followed by incomplete coalescence of the electron-dense content. Comparison of shell-globule structure in different taxa could be useful in elucidating some complex and still-unclear phylogenetic relationships among Platyhelminthes.     

Isolated Bovine Spinal Motoneurons Have Specific Ganglioside Antigens Recognized by Sera from Patients with Motor Neuron Disease and Motor Neuropathy

The gangliosides GM1 and GD1b have recently been reported to be potential target antigens in human motor neuron disease (MND) or motor neuropathy. The mechanism for selective motoneuron and motor nerve impairment by the antibodies directed against these gangliosides, however, is not fully understood. We recently investigated the ganglioside composition of isolated bovine spinal motoneurons and found that the ganglioside pattern of the isolated motoneurons was extremely complex. GM1, GD1a, GD1b, and GT1b, which are major ganglioside components of CNS tissues, were only minor species in motoneurons. Among the various ganglioside species in motoneurons, several were immunoreactive to sera from patients with MND and motor neuropathy. One of these gangliosides was purified from bovine spinal cord and characterized as N-glycolylneuraminic acid-containing GM1 [GM1(NeuGc)] by compositional analysis, fast atom bombardment mass spectra, and the use of specific antibodies. Among seven sera with anti-GM1 antibody activities, five sera reacted with GM1(NeuGc) and two did not. Two other gangliosides, which were recognized by another patient's serum, appeared to be specific for motoneurons. We conclude that motoneurons contained, in addition to the known ganglioside antigens GM1 and GD1b, other specific ganglioside antigens that could be recognized by sera from patients with MND and motor neuropathy.     

Enzymatic synthesis of 12-ketoursodeoxycholic acid from dehydrocholic acid in a membrane reactor

Summary Dehydrocholic acid (3,7,12-trioxo-5-cholanic acid) (0.5% concentration) was completely and selectively reduced to 12-ketoursodeoxycholic acid (3, 7-dihydroxy-12-oxo- 5-cholanic acid) in a membrane reactor by means of 3-hydroxysteroid dehydrogenase and 7-hydroxysteroid dehydrogenase. Coenzyme regeneration was carried out with the glucose-glucose dehydrogenase system.     

Evidence for the deficiency of β-glucosidase-activating factor in fibroblasts of patients with I-cell disease

Summary Reduced activity of -glucosidase was shown in the cultured skin fibroblasts of four patients with I-cell disease when the enzyme was tested without the use of detergents. In the presence of taurocholate and triton X100 -glucosidase activity was normal. This suggested a deficiency of a -glucosidase-activating factor in I-cell fibroblasts rather than of the enzyme itself. The deficiency of -glucosidase activity was corrected to some extent by mixing cell lysates, and more effectively by cocultivation and fusion of I-cell disease and Gaucher fibroblasts. These results present evidence for the presence of a -glucosidase-activating factor in normal and Gaucher fibroblasts. In fibroblasts of patients with I-cell disease this activator is probably deficient, as is the case for most lysosomal enzymes.     

Nusinersen treatment and cerebrospinal fluid neurofilaments: An explorative study on Spinal Muscular Atrophy type 3 patients

The antisense oligonucleotide Nusinersen has been recently licensed to treat spinal muscular atrophy (SMA). Since SMA type 3 is characterized by variable phenotype and milder progression, biomarkers of early treatment response are urgently needed. We investigated the cerebrospinal fluid (CSF) concentration of neurofilaments in SMA type 3 patients treated with Nusinersen as a potential biomarker of treatment efficacy. The concentration of phosphorylated neurofilaments heavy chain (pNfH) and light chain (NfL) in the CSF of SMA type 3 patients was evaluated before and after six months since the first Nusinersen administration, performed with commercially available enzyme-linked immunosorbent assay (ELISA) kits. Clinical evaluation of SMA patients was performed with standardized motor function scales. Baseline neurofilament levels in patients were comparable to controls, but significantly decreased after six months of treatment, while motor functions were only marginally ameliorated. No significant correlation was observed between the change in motor functions and that of neurofilaments over time. The reduction of neurofilament levels suggests a possible early biochemical effect of treatment on axonal degeneration, which may precede changes in motor performance. Our study mandates further investigations to assess neurofilaments as a marker of treatment response.     

Variability of Hypericins and Hyperforin in Hypericum Species from the Sicilian Flora

Within Sicilian flora, the genus Hypericum (Guttiferae) includes 10 native species, the most popular of which is H. perforatum. Hypericum’s most investigated active compounds belong to naphtodianthrones (hypericin, pseudohypericin) and phloroglucinols (hyperforin, adhyperforin), and the commercial value of the drug is graded according to its total hypericin content. Ethnobotanical sources attribute the therapeutic properties recognized for H. perforatum, also to other Hypericum species. However, their smaller distribution inside the territory suggests that an industrial use of such species, when collected from the wild, would result in an unacceptable depletion of their natural stands. This study investigated about the potential pharmacological properties of 48 accessions from six native species of Hypericum, including H. perforatum and five ‘minor’ species, also comparing, when possible, wild and cultivated sources. The variability in the content of active metabolites was remarkably high, and the differences within the species were often comparable to the differences among species. No difference was enlightened between wild and cultivated plants. A carefully planned cultivation of Hypericum seems the best option to achieve high and steady biomass yields, but there is a need for phytochemical studies, aimed to identify for multiplication the genotypes with the highest content of the active metabolites.     

Bacillus subtilis as a host for mosquitocidal toxins production

Aedes albopictus transmits several arboviral infections. In the absence of vaccines, control of mosquito populations is the only strategy to prevent vector-borne diseases. As part of the search for novel, biological and environmentally friendly strategies for vector control, the isolation of new bacterial species with mosquitocidal activity represents a promising approach. However, new bacterial isolates may be difficult to grow and genetically manipulate. To overcome these limits, here we set up a system allowing the expression of mosquitocidal bacterial toxins in the well-known genetic background of Bacillus subtilis. As a proof of this concept, the ability of B. subtilis to express individual or combinations of toxins of Bacillus thuringiensis israelensis (Bti) was studied. Different expression systems in which toxin gene expression was driven by IPTG-inducible, auto-inducible or toxin gene-specific promoters were developed. The larvicidal activity of the resulting B. subtilis strains against second-instar Ae. albopictus larvae allowed studying the activity of individual toxins or the synergistic interaction among Cry and Cyt toxins. The expression systems here presented lay the foundation for a better improved system to be used in the future to characterize the larvicidal activity of toxin genes from new environmental isolates.