Amyloid β / PKC-dependent alterations in NMDA receptor composition are detected in early stages of Alzheimer´s disease

Amyloid beta (Aβ)-mediated synapse dysfunction is an early event in Alzheimer’s disease (AD) pathogenesis and previous studies suggest that NMDA receptor (NMDAR) dysregulation may contribute to these pathological effects. Although Aβ peptides impair NMDAR expression and activity, the mechanisms mediating these alterations in the early stages of AD are unclear. Here, we observed that NMDAR subunit NR2B and PSD-95 levels were aberrantly upregulated and correlated with Aβ₄₂ load in human postsynaptic fractions of the prefrontal cortex in early stages of AD patients, as well as in the hippocampus of 3xTg-AD mice. Importantly, NR2B and PSD95 dysregulation was revealed by an increased expression of both proteins in Aβ-injected mouse hippocampi. In cultured neurons, Aβ oligomers increased the NR2B-containing NMDAR density in neuronal membranes and the NMDA-induced intracellular Ca²⁺ increase, in addition to colocalization in dendrites of NR2B subunit and PSD95. Mechanistically, Aβ oligomers required integrin β1 to promote synaptic location and function of NR2B-containing NMDARs and PSD95 by phosphorylation through classic PKCs. These results provide evidence that Aβ oligomers modify the contribution of NR2B to NMDAR composition and function in the early stages of AD through an integrin β1 and PKC-dependent pathway. These data reveal a novel role of Aβ oligomers in synaptic dysfunction that may be relevant to early-stage AD pathogenesis.Subject terms: Alzheimer''s disease, Extracellular signalling molecules     

Rapid differentiation and in situ detection of 16 sourdough lactobacillus species by multiplex PCR

A two-step multiplex PCR-based method was designed for the rapid detection of 16 species of lactobacilli known to be commonly present in sourdough. The first step of multiplex PCR was developed with a mixture of group-specific primers, while the second step included three multiplex PCR assays with a mixture of species-specific primers. Primers were derived from sequences that specify the 16S rRNA, the 16S-23S rRNA intergenic spacer region, and part of the 23S rRNA gene. The primer pairs designed were shown to exclusively amplify the targeted rrn operon fragment of the corresponding species. Due to the reliability of simultaneously identifying Lactobacillus plantarum, Lactobacillus pentosus, and Lactobacillus paraplantarum, a previously described multiplex PCR method employing recA gene-derived primers was included in the multiplex PCR system. The combination of a newly developed, quick bacterial DNA extraction method from sourdough and this multiplex PCR assay allows the rapid in situ detection of several sourdough-associated lactobacilli, including the recently described species Lactobacillus rossii, and thus represents a very useful alternative to culture-based methodologies.     

Nitrate removal by immobilized cells of Phormidium uncinatum in batch culture and a continuous-flow photobioreactor

Cells of the non-N₂-fixing cyanobacterium Phormidium uncinatum were immobilized by adsorption into polyvinyl (PV-50) foam pieces. The effect of inoculum size as well as the initial inoculum/support ratio on the cell immobilization process was investigated. After 2 months of immobilization similar net O₂-exchange activity was measured in immobilized and free-living cells. Polyvinyl-adsorbed cells also showed similar nitrate uptake capacity to free-living cells. Nitrogen starvation promoted a remarkable increase in nitrate uptake rate of both free-living and immobilized cells. A lab-scale photobioreactor packed with polyvinyl foam pieces colonized in situ by cells was used for nitrate removal in a continuous mode. In the best working conditions found, nearly 90% of nitrate supplied in the influent (50 mg l^–1) was removed by cells having a residence time of 3–4 h. Correspondence to: J. L. Serra     

Production of biomass from untreated orange peel by Fusarium avenaceum

Summary The growth behaviour of Fusarium avenaceum (Sect. Roseum Wr.) in slurry fermentation systems using untreated orange peel as substrate was studied in a laboratory-fermenter scale to reproduce the results obtained in a shakenflask fermenter. The eventual effect of impeller speed on mechanical disruption of mycelial hyphae was then assessed by determining mycelial growth, total reducing sugars consumption, TOC reduction, carbon dioxide evolution and oxygen absorption rates. In particular, the main biomass yield coefficient, as well as the apparent specific growth rate, appeared to be independent of the impeller speed, at least within the experimental range of 450 and 900 min^–1 (equivalent to peripheral impeller speeds of 3.8–7.5 m sec^–1.     

Tissue-selective alteration of ethanolamine plasmalogen metabolism in dedifferentiated colon mucosa

Human colon lipid analysis by imaging mass spectrometry (IMS) demonstrates that the lipid fingerprint is highly sensitive to a cell's pathophysiological state. Along the colon crypt axis, and concomitant to the differentiation process, certain lipid species tightly linked to signaling (phosphatidylinositols and arachidonic acid (AA)-containing diacylglycerophospholipids), change following a rather simple mathematical expression. We extend here our observations to ethanolamine plasmalogens (PlsEtn), a unique type of glycerophospholipid presenting a vinyl ether linkage at sn-1 position. PlsEtn distribution was studied in healthy, adenomatous, and carcinomatous colon mucosa sections by IMS. In epithelium, 75% of PlsEtn changed in a highly regular manner along the crypt axis, in clear contrast with diacyl species (67% of which remained constant). Consistently, AA-containing PlsEtn species were more abundant at the base, where stem cells reside, and decreased while ascending the crypt. In turn, mono?/diunsaturated species experienced the opposite change. These gradients were accompanied by a gradual expression of ether lipid synthesis enzymes. In lamina propria, 90% of stromal PlsEtn remained unchanged despite the high content of AA and the gradient in AA-containing diacylglycerophospholipids. Finally, both lipid and protein gradients were severely affected in polyps and carcinoma. These results link PlsEtn species regulation to cell differentiation for the first time and confirm that diacyl and ether species are differently regulated. Furthermore, they reaffirm the observations on cell lipid fingerprint image sensitivity to predict cell pathophysiological status, reinforcing the translational impact both lipidome and IMS might have in clinical research.     

α-Amylase and glucoamylase production by Schwanniomyces castellii

A chromogenic substrate (Cibachron blue-amylose), and soluble starch and maltose were used to characterize the amylolytic system from Schwanniomyces castellii 3754. The strain was able to produce inducible -amylase (EC 3.2.1.1) and glucoamylase (EC 3.2.1.3) when grown on different C sources. The effect of the C source was slightly different for -amylase and glucoamylase production. Melezitose, maltose and soluble starch enhanced both -amylase and glucoamylase synthesis to nearly the same extent; amylose, trehalose and cellobiose particularly induced -amylase synthesis. The optimal pH for the release of both amylases was 5.5–7.0; maximal -amylase synthesis, on the other hand, was observed in the medium buffered at pH 6.0. The optimal pH for -amylase and glucoamylase activity was in the range of 4.5–7.2 and 4.2–5.5, respectively. Temperatures allowing maximal activity were 45°C for -amylase and 45–52°C for glucoamylase; a rapid decline of both activities was observed just above these temperatures.The species Schwanniomyces castellii (together with Schw. alluvius) is now considered to be synonymous with Schw. occidentalis var. occidentalis (Kreger-Van Rij 1984).     

High-Throughput Mutation Profiling of Primary and Metastatic Endometrial Cancers Identifies KRAS,FGFR2 and PIK3CA to Be Frequently Mutated

Background

Despite being the most common pelvic gynecologic malignancy in industrialized countries, no targeted therapies are available for patients with metastatic endometrial carcinoma. In order to improve treatment, underlying molecular characteristics of primary and metastatic disease must be explored.

Methodology/Principal Findings

We utilized the mass spectrometric-based mutation detection technology OncoMap to define the types and frequency of point somatic mutations in endometrial cancer. 67 primary tumors, 15 metastases corresponding to 7 of the included primary tumors and 11 endometrial cancer cell lines were screened for point mutations in 28 known oncogenes. We found that 27 (40.3%) of 67 primary tumors harbored one or more mutations with no increase in metastatic lesions. FGFR2, KRAS and PIK3CA were consistently the most frequently mutated genes in primary tumors, metastatic lesions and cell lines.

Conclusions/Significance

Our results emphasize the potential for targeting FGFR2, KRAS and PIK3CA mutations in endometrial cancer for development of novel therapeutic strategies.