Native serotonin 5-HT3 receptors expressed in Xenopus oocytes differ from homopentameric 5-HT3 receptors

Efficacies of the 5-hydroxytryptamine (serotonin) 5-HT3 receptor (5-HT3R) agonists 2-methyl-5-HT, dopamine, and m-chlorophenylbiguanide on 5-HT3R native to N1E-115 cells and on homopentameric 5-HT3R expressed in Xenopus oocytes were determined relative to that of 5-HT. Efficacies of 2-methyl-5-HT and dopamine on 5-HT3R native to differentiated N1E-115 cells are high (54 and 36%) as compared with their efficacies on homopentameric 5-HT3R-A(L) and 5-HT3R-A(S) receptors expressed in oocytes (4-8%). m-Chlorophenylbiguanide does not distinguish between 5-HT3R in N1E-115 cells and in oocytes. The distinct pharmacological profile of 5-HT3R native to differentiated N1E-115 cells is conserved when poly(A)+ mRNA from these cells is expressed in oocytes. The results indicate that, apart from the known 5-HT3R subunits, N1E-115 cells express additional proteins involved in 5-HT3R function.     

Demonstration of discrete secreted and membrane-bound ocular mucins in the dog

Our aims were to separate and characterize secreted canine ocular mucins, and to provide definitive evidence of membrane-bound mucins at the canine ocular surface. Mucus was collected by suction from the ocular surface of normal dogs and dispersed in guanidine hydrochloride and a cocktail of protease inhibitors. Caesium chloride density gradient centrifugation separated secreted mucins from membranes, which were collected from the top of the gradients. Membranes were extracted with octyl glucoside and screened using lectins and anti-mucin antibodies. Gradient fractions containing secreted mucins were constituted into pools on the basis of differential lectin and antibody staining. High molecular weight material from each pool was purified by gel filtration. This material, and the membrane extract, were reduced and alkylated. Vacuum blotting of separated materials after agarose gel electrophoresis was used to compare subunit structure. Density gradient profiles indicated three principal secreted glycoprotein peaks: one staining strongly with anti-mucin antibodies. Gel filtration demonstrated that each contained high molecular weight material. Vacuum blots demonstrated the presence of two secreted glycoproteins with differently sized subunits. On the basis of buoyant density, one of these may be lipid complexed. Membrane extracted material stained with anti-mucin antibodies, and vacuum blotting of this material provided evidence for two membrane-bound components. In conclusion, we have shown that normal canine ocular mucus contains two secreted mucins, each exhibiting different subunit structure; one of these mucins may undergo lipid complexation. Normal canine ocular mucus also contains two membrane-bound mucins: one of which is unique among membrane mucins in showing subunit structure.     

Presidential reflections

Two direct methods for the diagnosis of trichinellosis were compared: trichinoscopy and artificial digestion. Muscles from 17 wistar rats, orally infected with 500 Trichinella spiralis encysted larvae were examined. From each of the following muscles: diaphragm, tongue, masseters, intercostals, triceps brachialis and cuadriceps femoralis, 648,440 larvae from 1 g samples were recovered. The linear correlation between trichinoscopy and artificial digestion was very high and significant (r = 0.94, p < 0.0001), showing that both methods for the detection of muscular larvae did not differ significantly. In both methods, significant differences were found in the distribution of larvae per gramme of muscle.     

Effect of vanadium and chromium on the microstructural features of V–Cr–Mn–Ni spheroidal carbide cast irons

The objective of this investigation is to study the influence of vanadium（5.0wt%–10.0wt%） and chromium（0–9.0wt%） on the microstructure and hardness of Cr-V-Mn-Ni white cast irons with spheroidal vanadium carbides. The alloys＇ microstructural features are presented and discussed with regard to the distribution of phase elements. The structural constituents of the alloys are spheroidal VC, proeutectoid cementite, ledeburite eutectic, rosette-shaped carbide eutectic（based on M7C3）, pearlite, martensite, and austenite. Their combinations and area fraction（AF） ratios are reported to be influenced by the alloys＇ chemical composition. Spheroidized VC particles are found to be sites for the nucleation of carbide eutectics. Cr and V are shown to substitute each other in the VC and M7C3 carbides, respectively. Chromium alloying leads to the formation of a eutectic（γ-Fe ＋ М7С3）, preventing the appearance of proeutectoid cementite in the structure. Vanadium and chromium are revealed to increase the total carbide fraction and the amount of austenite in the matrix. Cr is observed to play a key role in controlling the metallic matrix microstructure.     

Yield stress distribution in injection‐moulded glassy polymers

A methodology for structural analysis simulations is presented that incorporates the distribution of mechanical properties along the geometrical dimensions of injection‐moulded amorphous polymer products. It is based on a previously developed modelling approach, where the thermomechanical history experienced during processing was used to determine the yield stress at the end of an injection‐moulding cycle. Comparison between experimental data and simulation results showed an excellent quantitative agreement, both for short‐term tensile tests as well as long‐term creep experiments over a range of strain rates, applied stresses, and testing temperatures. Changes in mould temperature and component wall thickness, which directly affect the cooling profiles and, hence, the mechanical properties, were well captured by the methodology presented. Furthermore, it turns out that the distribution of the yield stress along a tensile bar is one of the triggers for the onset of the (strong) localization generally observed in experiments. © 2015 Society of Chemical Industry     

Structure-activity relationship study of 16 a-thiocamptothecins: an integrated in vitro and in silico approach

The instability of the hydroxylactone E ring represents a critical drawback of camptothecins, because the lactone ring is recognized to be essential for stabilization of topoisomerase I‐mediated DNA cleavage. In an attempt to investigate the effect of the thiopyridone pharmacophofore on the molecular and pharmacological features of the drug, we prepared a series of novel 16 a‐thiocamptothecin analogues. Due to the sulfur atom, a destabilization of the hydrogen bond between the hydroxy group in position 17 of the opened E ring and the carbonyl of the pyridone moiety is predicted, thus shifting the equilibrium toward the closed lactone form and increasing the lipophilic properties of the compounds. This feature was associated with superior antiproliferative potency, with reduced interaction with the human serum albumin and with substantial increase of the persistence of the topoisomerase I–DNA cleavable complex. These effects were prominent for thio‐SN38, the most active compound of the series. The favorable interactions at the molecular and cellular level of the reported thiocamptothecins confer promising features, and these compounds warrant preclinical development.     

Easy mass spectrometry for metabolomics and quality control of vegetable and animal fats

Easy ambient sonic spray ionization (EASI), a novel desorption/ionization technique for ambient mass spectrometry analysis, is shown to permit the characterization of different types of vegetable and animal fats. The taxonomic markers of the oils, mainly fatty acids and phenols, are extracted with methanol/water (1:1) solution and made alkaline with NH₄OH, then placed and dried on the surface of a glass tip and directly analyzed by EASI‐MS in the negative‐ion mode. EASI provides a supersonic cloud of charged droplets that causes efficient desorption and ionization of the oil markers directly from the surface of the glass tip. As proof‐of‐principle cases, EASI(?)‐MS was applied to genuine samples of olive oil, hazelnut oil, soybean oil, grape seed oil, canola oil, butter, and lard. Characteristic metabolomics EASI(?)‐MS profiles of fatty acids and eventually phenols were obtained.     

BRAF Gene and Melanoma: Back to the Future

As widely acknowledged, 40–50% of all melanoma patients harbour an activating BRAF mutation (mostly BRAF V600E). The identification of the RAS–RAF–MEK–ERK (MAP kinase) signalling pathway and its targeting has represented a valuable milestone for the advanced and, more recently, for the completely resected stage III and IV melanoma therapy management. However, despite progress in BRAF-mutant melanoma treatment, the two different approaches approved so far for metastatic disease, immunotherapy and BRAF+MEK inhibitors, allow a 5-year survival of no more than 60%, and most patients relapse during treatment due to acquired mechanisms of resistance. Deep insight into BRAF gene biology is fundamental to describe the acquired resistance mechanisms (primary and secondary) and to understand the molecular pathways that are now being investigated in preclinical and clinical studies with the aim of improving outcomes in BRAF-mutant patients.     

Prolonged neuromuscular blockade with mivacurium in a newborn

Recent reports have demonstrated that puerperal psychosis is preceded by prodromic signs. These signs must be recognized in a multidisciplinary context, opening the way to improved diagnosis, therapy and prognosis. On the basis of our clinical experience, we stress the important of early management in women who present prodromic signs of puerperal psychosis. Specific management relies on intensive psychotherapy with daily sessions centered on filiation. With this approach, the emergence of delusions can generally be prevented, usually allowing an uninterrupted mother-infant relationship. We hypothesize that this approach has a favorable influence on later mother-child interactions and cause profound changes in the mother's psychic organization, resulting in a better long-term prognosis. We illustrate our point with a case report.