Complexes of heparin with poly(alkylenimines): Competitive binding with methylene blue

The binding of heparin (Hep) and Hep fractions with oligo- and poly-(alkylenimines) having the general formula H₂N(CH₂-CHR-NH)_nH, where R = H or Me, has been investigated by spectroscopy, by evaluating the competition of the amines and Methylene Blue for the anionic sites of Hep. The strongest-binding was observed at pH 3.5, with the essentially linear triethylenetetramine and the slightly branched tetraethylenepentamine giving the most stable complexes. For N (number of nitrogen atoms per molecule) >5, a decrease of the binding ability of the amines was observed. The apparent stoichiometry of the complexes was a function of the relative concentration of Hep and the amine, indicating an equilibrium between different types of complexes. Beef-lung Hep and a Hep fraction consisting mainly of trisulphated disaccharide blocks gave stronger complexes than the more heterogeneous, pigmucosal Hep and a Hep fraction of lower sulphate content. The results are interpreted in terms of polyelectrolyte-type associations involving sulphate groups on adjacent residues of the Hep chain and sequences of charged nitrogen atoms on the polyamine.     

Animal Rennets as Sources of Dairy Lactic Acid Bacteria

The microbial composition of artisan and industrial animal rennet pastes was studied by using both culture-dependent and -independent approaches. Pyrosequencing targeting the 16S rRNA gene allowed to identify 361 operational taxonomic units (OTUs) to the genus/species level. Among lactic acid bacteria (LAB), Streptococcus thermophilus and some lactobacilli, mainly Lactobacillus crispatus and Lactobacillus reuteri, were the most abundant species, with differences among the samples. Twelve groups of microorganisms were targeted by viable plate counts revealing a dominance of mesophilic cocci. All rennets were able to acidify ultrahigh-temperature-processed (UHT) milk as shown by pH and total titratable acidity (TTA). Presumptive LAB isolated at the highest dilutions of acidified milks were phenotypically characterized, grouped, differentiated at the strain level by randomly amplified polymorphic DNA (RAPD)-PCR analysis, and subjected to 16S rRNA gene sequencing. Only 18 strains were clearly identified at the species level, as Enterococcus casseliflavus, Enterococcus faecium, Enterococcus faecalis, Enterococcus lactis, Lactobacillus delbrueckii, and Streptococcus thermophilus, while the other strains, all belonging to the genus Enterococcus, could not be allotted into any previously described species. The phylogenetic analysis showed that these strains might represent different unknown species. All strains were evaluated for their dairy technological performances. All isolates produced diacetyl, and 10 of them produced a rapid pH drop in milk, but only 3 isolates were also autolytic. This work showed that animal rennet pastes can be sources of LAB, mainly enterococci, that might contribute to the microbial diversity associated with dairy productions.     

Trisomy-8 mosaicism: Report of a case

Summary An 8-year-old male with mental retardation, speech difficulties, and minor congenital anomalies is presented. The clinical findings suggest the trisomy-8 syndrome. The karyotype indicates trisomy-8 mosaicism with trisomic as well as normal cell lines in blood lymphocytes.     

PRMT5 silencing selectively affects MTAP-deleted mesothelioma: In vitro evidence of a novel promising approach

Malignant mesothelioma (MM) is an aggressive asbestos-related cancer of the serous membranes. Despite intensive treatment regimens, MM is still a fatal disease, mainly due to the intrinsic resistance to current therapies and the lack of predictive markers and new valuable molecular targets. Protein arginine methyltransferase 5 (PRMT5) inhibition has recently emerged as a potential therapy against methylthioadenosine phosphorylase (MTAP)-deficient cancers, in which the accumulation of the substrate 5'-methylthioadenosine (MTA) inhibits PRMT5 activity, thus sensitizing the cells to further PRMT5 inhibition. Considering that the MTAP gene is frequently codeleted with the adjacent cyclin-dependent kinase inhibitor 2A (CDKN2A) locus in MM, we assessed whether PRMT5 could represent a therapeutic target also for this cancer type. We evaluated PRMT5 expression, the MTAP status and MTA content in normal mesothelial and MM cell lines. We found that both administration of exogenous MTA and stable PRMT5 knock-down, by short hairpin RNAs (shRNAs), selectively reduced the growth of MTAP-deleted MM cells. We also observed that PRMT5 knock-down in MTAP-deficient MM cells reduced the expression of E2F1 target genes involved in cell cycle progression and of factors implicated in epithelial-to-mesenchymal transition. Therefore, PRMT5 targeting could represent a promising new therapeutic strategy against MTAP-deleted MMs.     

Biotechnological approaches to overcome hybridization barriers and use of micropropagation tool for further improvement in Heliconia: a review

Plant Cell, Tissue and Organ Culture (PCTOC) - Heliconia, also known as ‘False-Bird-of-Paradise’, is a genus of flowering plants in the monotypic family of Heliconiaceae.The genus...     

Ethylene emission and abscission of immature ‘Montmorency’ sour cherry fruitlets vary with ethephon concentration,phenology stage and ambient temperatures following application

Plant Growth Regulation - Ethylene is an endogenous plant hormone associated with natural senescence and abscission of plant organs. Ethylene release from the degradation of 2-chloroethylphosphonic...     

Routine Clinical-Pathologic Correlation of Pigmented Skin Tumors Can Influence Patient Management

Background

Several studies have demonstrated the benefit of integrating clinical with pathologic information, to obtain a confident diagnosis for melanocytic tumors. However, all those studies were conducted retrospectively and no data are currently available about the role of a clinical-pathologic correlation approach on a daily basis in clinical practice.

Aim of the Study

In our study, we evaluated the impact of a routine clinical-pathologic correlation approach for difficult skin tumors seen over 3 years in a tertiary referral center.

Results

Interestingly, a re-appraisal was requested for 158 out of 2015 (7.7%) excised lesions because clinical-pathologic correlation was missing. Of note, in 0.6% of them (13 out of 2045) the first histologic diagnosis was revised in the light of clinical information that assisted the Pathologist to re-evaluate the histopathologic findings that might be bland or inconspicuous per se.

Conclusion

In conclusion, our study demonstrated that an integrated approach involving clinicians and pathologists allows improving management of selected patients by shifting from a simply disease-focused management (melanoma versus nevus) to a patient-centered approach.     

Different Stress-Induced Calcium Signatures Are Reported by Aequorin-Mediated Calcium Measurements in Living Cells of Aspergillus fumigatus

Aspergillus fumigatus is an inhaled fungal pathogen of human lungs, the developmental growth of which is reliant upon Ca²⁺-mediated signalling. Ca²⁺ signalling has regulatory significance in all eukaryotic cells but how A. fumigatus uses intracellular Ca²⁺ signals to respond to stresses imposed by the mammalian lung is poorly understood. In this work, A. fumigatus strains derived from the clinical isolate CEA10, and a non-homologous recombination mutant ΔakuB ^KU80, were engineered to express the bioluminescent Ca²⁺-reporter aequorin. An aequorin-mediated method for routine Ca²⁺ measurements during the early stages of colony initiation was successfully developed and dynamic changes in cytosolic free calcium ([Ca²⁺]_c) in response to extracellular stimuli were measured. The response to extracellular challenges (hypo- and hyper-osmotic shock, mechanical perturbation, high extracellular Ca²⁺, oxidative stress or exposure to human serum) that the fungus might be exposed to during infection, were analysed in living conidial germlings. The ‘signatures’ of the transient [Ca²⁺]_c responses to extracellular stimuli were found to be dose- and age-dependent. Moreover, Ca²⁺-signatures associated with each physico-chemical treatment were found to be unique, suggesting the involvement of heterogeneous combinations of Ca²⁺-signalling components in each stress response. Concordant with the involvement of Ca²⁺-calmodulin complexes in these Ca²⁺-mediated responses, the calmodulin inhibitor trifluoperazine (TFP) induced changes in the Ca²⁺-signatures to all the challenges. The Ca²⁺-chelator BAPTA potently inhibited the initial responses to most stressors in accordance with a critical role for extracellular Ca²⁺ in initiating the stress responses.     

Interaction between Calpain-1 and HSP90: New Insights into the Regulation of Localization and Activity of the Protease

Here we demonstrate that heat shock protein 90 (HSP90) interacts with calpain-1, but not with calpain-2, and forms a discrete complex in which the protease maintains its catalytic activity, although with a lower affinity for Ca²⁺. Equilibrium gel distribution experiments show that this complex is composed by an equal number of molecules of each protein partner. Moreover, in resting cells, cytosolic calpain-1 is completely associated with HSP90. Since calpain-1, in association with HSP90, retains its proteolytic activity, and the chaperone is displaced by calpastatin also in the absence of Ca²⁺, the catalytic cleft of the protease is not involved in this association. Thus, calpain-1 can form two distinct complexes depending on the availability of calpastatin in the cytosol. The occurrence of a complex between HSP90 and calpain-1, in which the protease is still activable, can prevent the complete inhibition of the protease even in the presence of high calpastatin levels. We also demonstrate that in basal cell conditions HSP90 and calpain-1, but not calpain-2, are inserted in the multi-protein N-Methyl-D-Aspartate receptor (NMDAR) complex. The amount of calpain-1 at the NMDAR cluster is not modified in conditions of increased [Ca²⁺]_i, and this resident protease is involved in the processing of NMDAR components. Finally, the amount of calpain-1 associated with NMDAR cluster is independent from Ca²⁺-mediated translocation. Our findings show that HSP90 plays an important role in maintaining a given and proper amount of calpain-1 at the functional sites.