Investigation of the inclusion of the herbicide metobromuron in native cyclodextrins by powder X-ray diffraction and isothermal titration calorimetry

We report the formation of inclusion complexes between the phenylurea herbicide metobromuron [3-(p-bromophenyl)-1-methoxy-1-methylurea] and β- and γ-cyclodextrin in the solid state. Formation of crystalline inclusion complexes by the kneading method was confirmed by powder X-ray diffraction and further structural characterization using the principles of isostructurality followed. In addition, ΔH°, ΔS°, ΔG° and the association constants (K) at 298 K were determined for complex formation in solution using isothermal titration calorimetry. The magnitudes of K for the formation of 1:1 complexes between metobromuron and α-, β- and γ-CD were estimated as 598, 310 and 114, respectively.     

The Vesicular Acetylcholine Transporter Is Required for Neuromuscular Development and Function

The vesicular acetylcholine (ACh) transporter (VAChT) mediates ACh storage by synaptic vesicles. However, the VAChT-independent release of ACh is believed to be important during development. Here we generated VAChT knockout mice and tested the physiological relevance of the VAChT-independent release of ACh. Homozygous VAChT knockout mice died shortly after birth, indicating that VAChT-mediated storage of ACh is essential for life. Indeed, synaptosomes obtained from brains of homozygous knockouts were incapable of releasing ACh in response to depolarization. Surprisingly, electrophysiological recordings at the skeletal-neuromuscular junction show that VAChT knockout mice present spontaneous miniature end-plate potentials with reduced amplitude and frequency, which are likely the result of a passive transport of ACh into synaptic vesicles. Interestingly, VAChT knockouts exhibit substantial increases in amounts of choline acetyltransferase, high-affinity choline transporter, and ACh. However, the development of the neuromuscular junction in these mice is severely affected. Mutant VAChT mice show increases in motoneuron and nerve terminal numbers. End plates are large, nerves exhibit abnormal sprouting, and muscle is necrotic. The abnormalities are similar to those of mice that cannot synthesize ACh due to a lack of choline acetyltransferase. Our results indicate that VAChT is essential to the normal development of motor neurons and the release of ACh.Cholinergic neurotransmission has key functions in life, as it regulates several central and peripheral nervous system outputs. Acetylcholine (ACh) is synthesized in the cytoplasm by the enzyme choline acetyltransferase (ChAT) (16). Choline supplied by the high-affinity choline transporter (CHT1) is required to maintain ACh synthesis (52). A lack of ChAT (4, 35) or the high-affinity choline transporter (21) in genetically modified mice is incompatible with life. ACh plays an important role in wiring the neuromuscular junction (NMJ) during development (38, 43). Embryonic synthesis of ACh is fundamental for the development of proper nerve-muscle patterning at the mammalian NMJ, as ChAT-null mice present aberrant nicotinic ACh receptor (nAChR) localization and increased motoneuron (MN) survival, axonal sprouting, and branching (4, 35).The vesicular ACh transporter (VAChT) exchanges cytoplasmic ACh for two vesicular protons (37, 41). Previously reported electrophysiological studies showed that quantal size is decreased by vesamicol, an inhibitor of VAChT, but only in nerve terminals that have been electrically stimulated (19, 59, 60, 63). VAChT overexpression in developing Xenopus MNs increases both the size and frequency of miniature-end-plate currents (54). In Caenorhabditis elegans, mutations in VAChT affect behavior (65). Moreover, a decrease in VAChT expression has functional consequences for mammals, as mutant mice with a 70% reduction in the expression levels of this transporter (VAChT knockdown [KD^HOM] mice) are myasthenic and have cognitive deficits (47). Hence, vesicular transport activity is rate limiting for neurotransmission “in vivo” (18, 47).Exocytosis of synaptic vesicle contents is the predominant mechanism for the regulated secretion of neurotransmitters (55). However, alternative mechanisms of secretion have been proposed (20, 56, 61). Quantal ACh release, comparable to that seen in developing nerve terminals, has been detected in myocytes and fibroblasts in culture, which presumably do not express VAChT (14, 24). More recently, it was found that the correct targeting of Drosophila photoreceptor axons is disrupted in flies with null mutations in ChAT (64). Remarkably, the inactivation of VAChT did not produce the same result (64). The result suggests that the release of ACh during development is not dependent on VAChT, perhaps because it is nonvesicular or because vesicular storage can occur without VAChT.To test if the VAChT-independent secretion of ACh has any physiological role in the mammalian nervous system, we generated a mouse line in which the VAChT gene is deleted. These mice lack the stimulated release of ACh from synaptosomes, die after birth, and show several alterations in neuromuscular wiring consistent with a severe decrease in the cholinergic input to muscles during development. These experiments indicate that VAChT has an important role in maintaining activity-dependent ACh release that supports life and the correct patterning of innervation at the NMJ.     

Structural details and composition of <Emphasis Type="Italic">Trichomonas vaginalis</Emphasis> lipophosphoglycan in relevance to the epithelial immune function

Trichomonas vaginalis causes the most common non-viral sexually transmitted infection linked to increased risk of premature birth, cervical cancer and HIV. This study defines molecular domains of the parasite surface glycoconjugate lipophosphoglycan (LPG) with distinct functions in the host immunoinflammatory response. The ceramide phospho-inositol glycan core (CPI-GC) released by mild acid had Mr of ∼8,700 Da determined by MALDI-TOF MS. Rha, GlcN, Gal and Xyl and small amounts of GalN and Glc were found in CPI-GC. N-acetyllactosamine repeats were identified by endo-β-galactosidase treatment followed by MALDI-MS and MS/MS and capLC/ESI-MS/MS analyses. Mild acid hydrolysis led to products rich in internal deoxyhexose residues. The CPI-GC induced chemokine production, NF-κB and extracellular signal-regulated kinase (ERK)1/2 activation in human cervicovaginal epithelial cells, but neither the released saccharide components nor the lipid-devoid LPG showed these activities. These results suggest a dominant role for CPI-GC in the pathogenic epithelial response to trichomoniasis.     

Oxidative Stress in Cerebrospinal Fluid of Patients with Aseptic and Bacterial Meningitis

This study aimed to determine whether patients with aseptic and bacterial meningitis presented alterations in oxidative stress parameters of cerebrospinal fluid (CSF). A total of 30 patients were used in the research. The CSF oxidative stress status has been evaluated through many parameters, such as lipid peroxidation through thiobarbituric acid reactive substances (TBARS) and antioxidant defense systems such as superoxide dismutase (SOD), glutathione S-transferase (GST), reduced glutathione (GSH) and ascorbic acid. TBARS levels, SOD and GST activity increase in aseptic meningitis and in bacterial meningitis. The ascorbic acid concentration increased significantly in patients with both meningitis types. The reduced glutathione levels were reduced in CSF of patients with aseptic and bacterial meningitis. In present study we may conclude that oxidative stress contributes at least in part to the severe neurological dysfunction found in meningitis.     

Initiation of Oligodendrocyte Progenitor Cell Migration by a PDGF-A Activated Extracellular Regulated Kinase (ERK) Signaling Pathway

During CNS development, oligodendrocyte progenitor (OP) cells migrate from germinal zones to presumptive white matter tracts to generate myelinating oligodendrocytes. In vitro and in vivo studies indicate that platelet-derived growth factor-A (PDGF-A) is a potent chemoattractant for OP cells and important for normal distribution throughout the developing CNS. However, PDGF-A does not localize in concentration gradients corresponding to OP migratory pathways, as would be expected for a chemoattractant to direct migration. Therefore, the mechanism by which PDGF-A regulates OP distribution remains to be clarified. Here we show that PDGF-A induces OP migration and continuous exposure to PDGF-A is not required to maintain migration. Using pharmacological inhibitors, we show that a self-sustaining extracellular-regulated-kinase signaling pathway drives OP migration for up to 72 hours after the initial PDGF stimulus. These findings indicate PDGF-A may act to mobilize OP cells that then respond to distinct directional signals to distribute appropriately within the CNS. Special issue article in honor of Dr. George DeVries.     

Synthesis and antibacterial activity evaluation of a novel cotton fiber (Gossypium barbadense) ampicillin derivative

We prepared cellulose cotton fibers containing ampicillin moieties and evaluated their antibacterial activity. In spite of recent progress in experimental and clinical medicine, the problem of chronic wounds treatment remains to be solved. In fact conventional methods are based on solutions of antibiotics and antiseptics and ointment bandages but the efficacy of this method is low and so the idea to use modified cotton gauzes would have to prevent infections insorgence during wounds healing. Ampicillin, a large spectrum antibiotic, was covalently coupled to cellulose backbone of hydrophilic cotton fibers by a heterogeneous synthesis to produce a functionalized biopolymer with a satisfactory degree of substitution (DS) and antibacterial activity. The obtained biopolymer was characterized by infrared spectroscopy (FT-IR). Finally, the antibacterial activity in inhibiting microorganism growth in Petri dishes, was evaluated. The results suggested that these biomaterials posses an excellent “in vitro” antibacterial activity and so they can be efficiently employed in biomedical fields for chronic wounds management to ensure a valid protection against infections and contaminations. Biopolymers so functionalized were found to be very efficient to contrast sensible bacteria growth.     

Generation of large droplet aerosols within microbiological containment using a novel flow-focussing technique

Flow-focussing technology was harnessed to enable generation of large droplet aerosols within high-level microbiological containment. The Collison nebuliser and flow-focussing aerosol generator (FFAG) produced aerosols from distilled water with average mass median aerodynamic diameters (MMAD) of 4.19 and 11.93 μm, respectively. The medium type [water, phosphate buffered saline (PBS) or microbiological broth] had minimal effect on the droplet size generated by the FFAG. The FFAG can be modulated to generate reproducible aerosols with a wide range of MMADs (9–105 μm). The number of particulates (i.e. fluospheres) contained within the droplets increased as the MMAD increased from 12 to 50 μm. The technology described can be used for the exposure of small-animal models to large droplet aerosols (>10 μm) and has applications in healthcare, pharmaceutical, agricultural and biodefence environments.     

Prolactin induces chitotriosidase expression in human macrophages through PTK,PI3‐K,and MAPK pathways

We previously reported that prolactin (PRL) induces chitotriosidase (CHIT‐1) mRNA expression in human macrophages. In this investigation we determined the signaling pathways involved in CHIT‐1 induction in response to PRL. The CHIT‐1 induction PRL‐mediated was reduced by wortmannin and LY‐294002, inhibitors of phosphatidylinositol 3‐kinase (PI3‐K) and by genistein an inhibitor of protein tyrosine kinase (PTK). Pre‐treatment of macrophages with SB203580, a specific inhibitor of the mitogen‐activated kinases (MAPK) p38, or with U0126, an inhibitor of MAPK p44/42, prevented both basal and exogenous PRL‐mediated CHIT‐1 expression. No significant effects on CHIT‐1 induction PRL‐mediated were observed with a protein kinase C inhibitor (PKC), rottlerin, or with an Src inhibitor, PP2, or with JAK2 inhibitor, AG490. In addition, PRL induced a phosphorylation of AKT that was prevented both by the two MAPK inhibitors SB203580 and U0126 and by the PI3‐K inhibitors wortmannin and LY‐294002. In conclusion, our results indicate that PRL up‐regulated CHIT‐1 expression via PTK, PI3‐K, MAPK, and signaling transduction components. J. Cell. Biochem. 107: 881–889, 2009. © 2009 Wiley‐Liss, Inc.     

Oral health and oral implant status in edentulous patients with implant‐supported dental prostheses who are receiving long‐term nursing care

Aim: The aim of this study was to investigate oral health and oral implant status in a group of edentulous patients receiving long‐term residential or nursing care (LTC), all of whom had implant‐supported fixed or removable dental prostheses. Material and methods: A dental examination was performed on a total of 3310 patients receiving LTC and from this population 35 edentulous patients in whom dental implants had been placed formed the cohort for this study. All examinations were performed by a specialist in hospital dentistry and took place in the patients’ own home environment. Oral health was assessed by means of a protocol which evaluated oral hygiene status, possible oral mucosal inflammation and oral mucosal friction levels. Any problems with the implant‐supported prosthesis, implant mobility or other complications were also assessed. In addition, patients were asked about any oral symptoms and their usual oral hygiene procedures. Results: About half of the subjects (17/35) were registered as having no/mild inflammation with 18 of 35 having moderate/severe inflammation. Twelve of the 35 patients had good/acceptable oral hygiene and 23 of 35 had poor/bad oral hygiene. Twenty‐one of the 35 patients depended on help from the nursing personnel for their daily oral hygiene procedures. Obvious problems with food impaction were noted in 11 patients. A total of 229 implants had been placed in 43 jaws supporting 40 full arch‐fixed prostheses and three implant‐borne overdentures. There was no evidence of mobility or fractures of either the implants or the prostheses. Fifteen implants showed some exposed screw threads. Pus was exuding from one implant site and general peri‐implant gingival hyperplasia was noted in two patients. Twenty‐four patients were completely satisfied with the function and appearance of their implant‐supported prostheses. Two patients were totally dissatisfied. Conclusion: This study indicates that oral implant therapy can be considered as a treatment of choice in elderly patients, even if oral hygiene is sub‐optimal.