Neurotoxicity studies with the monoamine oxidase B substrate 1-methyl-3-phenyl-3-pyrroline

The neurotoxic properties of the parkinsonian inducing agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) are dependent on its metabolic activation in a reaction catalyzed by centrally located monoamine oxidase B (MAO-B). This reaction ultimately leads to the permanently charged 1-methyl-4-phenylpyridinium species MPP(+), a 4-electron oxidation product of MPTP and a potent mitochondrial toxin. The corresponding 5-membered analogue, 1-methyl-3-phenyl-3-pyrroline, is also a selective MAO-B substrate. Unlike MPTP, the MAO-B-catalyzed oxidation of 1-methyl-3-phenyl-3-pyrroline is a 2-electron process that leads to the neutral 1-methyl-3-phenylpyrrole. MPP(+) is thought to exert its toxic effects only after accumulating in the mitochondria, a process driven by the transmembrane electrochemical gradient. Since this energy-dependent accumulation of MPP(+) relies upon its permanent charge, 1-methyl-3-phenyl-3-pyrrolines and their pyrrolyl oxidation products should not be neurotoxic. We have tested this hypothesis by examining the neurotoxic potential of 1-methyl-3-phenyl-3-pyrroline and 1-methyl-3-(4-chlorophenyl)-3-pyrroline in the C57BL/6 mouse model. These pyrrolines did not deplete striatal dopamine while analogous treatment with MPTP resulted in 65-73% depletion. Kinetic studies revealed that both 1-methyl-3-phenyl-3-pyrroline and its pyrrolyl oxidation product were present in the brain in relatively high concentrations. Unlike MPP(+), however, 1-methyl-3-phenylpyrrole was cleared from the brain quickly. These results suggest that the brain MAO-B-catalyzed oxidation of xenobiotic amines is not, in itself, sufficient to account for the neurodegenerative properties of a compound like MPTP. The rapid clearance of 1-methyl-3-phenylpyrroles from the brain may contribute to their lack of neurotoxicity.     

Conformational stabilization at the active site of molluskan (Rapana thomasiana) hemocyanin by a cysteine-histidine thioether bridge A study by mass spectrometry and molecular modeling

In some type-3 copper proteins (molluskan hemocyanin, catechol oxidase and fungal tyrosinase) one of the histidine residues, liganding the Cu(A) atom of the dinuclear copper active site, is covalently linked to a cysteine residue by a thioether bridge. The purpose of this study was to disclose the function of this bridge. Mass spectral analysis of a peptide, isolated from Rapana thomasiana (gastropodan mollusk) hemocyanin, indicated a stabilization of the peptide structure in the region of the bridge. Molecular modeling of three thioether containing type-3 copper proteins using the dead-end elimination method showed that the concerned histidine would be very flexible if not linked to the cysteine. Also, the side chain orientation of the histidine is rather exceptional, as evidenced by statistical data from the protein databank. It is suggested that the role of the bridge is to fix the histidine in an orientation that is optimal for coordination of the Cu(A) atom.     

Fracture prevention in COPD patients; a clinical 5-step approach

Although osteoporosis and its related fractures are common in patients with COPD, patients at high risk of fracture are poorly identified, and consequently, undertreated. Since there are no fracture prevention guidelines available that focus on COPD patients, we developed a clinical approach to improve the identification and treatment of COPD patients at high risk of fracture. We organised a round-table discussion with 8 clinical experts in the field of COPD and fracture prevention in the Netherlands in December 2013. The clinical experts presented a review of the literature on COPD, osteoporosis and fracture prevention. Based on the Dutch fracture prevention guideline, they developed a 5-step clinical approach for fracture prevention in COPD. Thereby, they took into account both classical risk factors for fracture (low body mass index, older age, personal and family history of fracture, immobility, smoking, alcohol intake, use of glucocorticoids and increased fall risk) and COPD-specific risk factors for fracture (severe airflow obstruction, pulmonary exacerbations and oxygen therapy). Severe COPD (defined as postbronchodilator FEV₁ < 50% predicted) was added as COPD-specific risk factor to the list of classical risk factors for fracture. The 5-step clinical approach starts with case finding using clinical risk factors, followed by risk evaluation (dual energy X-ray absorptiometry and imaging of the spine), differential diagnosis, treatment and follow-up. This systematic clinical approach, which is evidence-based and easy-to-use in daily practice by pulmonologists, should contribute to optimise fracture prevention in COPD patients at high risk of fracture.     

Type XVII collagen (BP180) can function as a cell−matrix adhesion molecule via binding to laminin 332

Collagen XVII (COL17) is a transmembrane glycoprotein that is expressed on the basal surface of basal epidermal keratinocytes. Previous observations have led to the hypothesis that an interaction between COL17 and laminin 332, an extracellular matrix protein, contributes to the attachment of the basal keratinocyte to the basement membrane. In order to isolate and manipulate COL17 interactions with ECM components, we induced COL17 expression in two cells lines, SK-MEL1 and K562, that exhibit little or no capacity to attach to our test substrates, including laminin 332, types I and IV collagen, and fibronectin. Cells expressing high levels of COL17 preferentially adhered to a laminin 332 matrix, and, to a lesser extent, type IV collagen, while showing little or no binding to type I collagen or fibronectin. A quantitative analysis of cell adhesive forces revealed that, compared with COL17-negative cells, COL17-positive cells required over 7-fold greater force to achieve 50% detachment from a laminin 332 substrate. When a cell preparation (either K562 or SK-MEL1) with heterogeneous COL17 expression levels was allowed to attach to a laminin 332 matrix, the COL17-positive and COL17-negative cells differentially sorted to the bound and unbound cell fractions, respectively. COL17-dependent attachment to laminin 332 could be reduced or abolished by siRNA-mediated knock-down of COL17 expression or by adding to the assay wells specific antibodies against COL17 or laminin 332. These findings provide strong support for the hypothesis that cell surface COL17 can interact with laminin 332 and, together, participate in the adherence of a cell to the extracellular matrix.     

Review of scientific contributions by the Belgian medical centers concerned with human in vitro fertilization and embryo transfer (IVF).

In vitro fertilization and embryo transfer (IVF) may be considered as a particular application of modern medical therapeutics linked to human reproduction. The treatment of human sterility therefore involves some fundamental human values such as life, love and death. The quality of this highly technological treatment with fast knowledge of outcome at the end of the patient's menstrual cycle has been evaluated since the early 80s. It is a typically multidisciplinary team effort involving medical doctors, biologists, laboratory technicians, nurses and clerks that is representative of modern medical practice. IVF covers much more than just embryology, as this review will explain. IVF developed in close relation with clinical and experimental research protocols, which are the major topics of this paper. The newness of the techniques used led to the necessary interactions between clinicians and biologists working on animal experimental embryology.     

The instantly released Drosophila immune proteome is infection-specific

In this study, we analyzed the hemolymph proteome of Drosophila third instar larvae, which were induced with a suspension of Gram-positive bacteria or yeast. Profiling of the hemolymph proteins of infected versus non-infected larvae was performed by two-dimensional difference gel electrophoresis. Infection with Micrococcus luteus or Saccharomyces cerevisiae induced, respectively, 20 and 19 differential protein spots. The majority of the spots are specifically regulated by one pathogen, whereas only a few spots correspond to proteins altered in all cases of challenging (including after challenge with lipopolysaccharides). All of the upregulated proteins can be assigned to specific aspects of the immune system, as they did not increase in the hemolymph of sterile pricked larvae. Next to known immune proteins, unannotated proteins were identified such as CG4306 protein, which has homologues with unknown function in all metazoan genome databases available today.     

Discrimination by male dogwood borer, Synanthedon scitula (Lepidoptera: Sesiidae), among traps baited with commercially available pheromone lures

The response of male dogwood borer, Synanthedon scitula (Harris) (Lepidoptera: Sesiidae), to four commercially available pheromone lures and to pheromone source concentration was evaluated in field trapping studies in commercial apple orchards in Virginia and West Virginia Male peachtree borer, Synanthedon exitiosa (Say), lesser peachtree borer, Synanthedon pictipes (Grote & Robinson), and lilac borer, Podosesia syringae (Harris), were also captured in traps baited with all lures tested. Among the lures used, the Scenturion dogwood borer lure was the most attractive to and selective for dogwood borer. Male dogwood borer showed a concentration-dependent response to traps baited with different source concentrations of the pheromone from Scenturion. Chemical analyses revealed that all lures contained principally (Z,Z)-3,13-octadecadien-1-ol acetate (ODDA) but that a significantly higher percentage of (Z,E) plus (E,Z)-3,13-ODDA was present in the Scenturion lure, compared with the others. Weekly collections of fresh male and female dogwood borer pupal exuviae from April until October in 2002 and 2003 suggested that traps baited with Scenturion lures significantly underestimated the size of populations in commercial apple orchards.     

Effect of a single in ovo injection of 2,3,7,8-tetrachlorodibenzo-p-dioxin on protein expression in liver and ovary of the one-day-old chick analyzed by fluorescent two-dimensional difference gel electrophoresis and mass spectrometry

The polyhalogenated aromatic hydrocarbon 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is an ubiquitously distributed environmental pollutant which can induce a broad spectrum of toxic responses in animals, including birds. In this study, we investigated the impact of 0 or 20 ng TCDD injections into the yolk of chicken eggs before start of development, on liver and ovarian protein expression in hatchlings using fluorescent two-dimensional difference gel electrophoresis (2-D-DIGE) under a pH range of 4-7, combined with MS. Despite considerable interindividual variability, exposure to TCDD prior to the start of embryonic development resulted in significant changes in expression of a small set of proteins. Expression of fibrinogen gamma chain precursor in the liver and 60 kDa heat shock protein in the ovary were significantly higher as a result of the very early exposure to TCDD. NADH ubiquinone oxidoreductase (42 kDa subunit) and regucalcin expression was decreased by early TCDD treatment in the liver and ovary, respectively. These proteins could not be directly linked with drug metabolism per se but are involved in blood clotting, oxidative stress, electron transport, and calcium regulation. It remains to be elucidated how these changes in the hatchling might be linked to the observed long-term consequences during posthatch life of the chicken.