Foliar and wood chemistry of sugar maple along a gradient of soil acidity and stand health

The decline of sugar maple (Acer saccharum Marsh.) in forest of north-eastern North America is an important environmental issue. In this study, relationships between, soil, wood and foliar chemistry were assessed for 17 stands distributed within a large area of the Quebec sugar maple forest and that were growing on soils with a strong gradient of acidity and base saturation. There were many significant relationships between variables describing the acid-base status of the top-B soil (Ca and Mg concentrations, exchangeable acidity and base saturation) and Ca and Mn concentrations and Ca/Mn and Mg/Mn in tree tissues. Manganese was the element that showed the strongest inverse non-linear relationships with top-B soil base saturation with variance explanation of 71 and 65%, for wood and foliage, respectively. The 17 sites were divided in two groups according to their level of decline. The declining stands had significantly higher wood Mn and Mg concentrations and lower Ca/Mn ratios and significantly higher foliar Mn and lower Ca and Al concentrations. It was impossible to determine if these differences were a cause or a symptom of sugar maple health. However, the increase in Mn concentrations in tree tissues with increasing soil acidity, as well as the higher Mn concentrations in declining as compared to healthy stands suggest that Mn, as well as low Ca availability, could be an important contributing factor in the sugar maple decline.     

Evaluation of the <Emphasis Type="Italic">OPTC</Emphasis> gene in primary open angle glaucoma: functional significance of a silent change

Background  

We investigated the molecular basis of primary open-angle glaucoma (POAG) using Opticin (OPTC) as a candidate gene on the basis of its expression in the trabecular meshwork cells involved in the disease pathogenesis. Two hundred POAG patients and 100 controls were enrolled in this study. The coding sequence of OPTC was amplified by PCR from genomic DNA of POAG patients, followed by SSCP, DHPLC and DNA sequencing. Subsequent bioinformatic analysis, site-directed mutagenesis, quantitative RT-PCR and western blot experiments were performed to address the functional significance of a 'silent' change in the OPTC coding region while screening for mutations in POAG patients.     

Insect defoliation modulates influence of climate on the growth of tree species in the boreal mixed forests of eastern Canada

Increasing air temperatures and changing precipitation patterns due to climate change can affect tree growth in boreal forests. Periodic insect outbreaks affect the growth trajectory of trees, making it difficult to quantify the climate signal in growth dynamics at scales longer than a year. We studied climate‐driven growth trends and the influence of spruce budworm (Choristoneura fumiferana Clem.) outbreaks on these trends by analyzing the basal area increment (BAI) of 2058 trees of Abies balsamea (L.) Mill., Picea glauca (Moench) Voss, Thuja occidentalis L., Populus tremuloides Michx., and Betula papyrifera Marsh, which co‐occurs in the boreal mixedwood forests of western Quebec. We used a generalized additive mixed model (GAMM) to analyze species‐specific trends in BAI dynamics from 1967 to 1991. The model relied on tree size, cambial age, degree of spruce budworm defoliation, and seasonal climatic variables. Overall, we observed a decreasing growth rate of the spruce budworm host species, A. balsamea and P. glauca between 1967 and 1991, and an increasing growth rate for the non‐host, P. tremuloides, B. papyrifera, and T. occidentalis. Our results suggest that insect outbreaks may offset growth increases resulting from a warmer climate. The observation warrants the inclusion of the spruce budworm defoliation into models predicting future forest productivity.     

Activated satellite cells fail to restore myonuclear number in spinal cord transected and exercised rats

In this study, possible mechanisms underlying soleus muscleatrophy after spinal cord transection and attenuation of atrophy withcycling exercise were studied. Adult female Sprague-Dawley rats weredivided into three groups; in two groups the spinal cord was transectedby a lesion at T₁₀. One group wastransected and killed 10 days later, and another group was transectedand exercised for 5 days starting 5 days after transection. The third group served as an uninjured control. All animals received acontinuous-release 5'-bromo-2'-deoxyuridine pellet 10 daysbefore they were killed. Transection alone and transection withexercise lead to activation of satellite cells, but only the exercisegroup showed a trend toward an increase in the number of proliferatingsatellite cells. In all cases the number of activated satellite cellswas significantly higher than the number that divided. Although thenumber of cells undergoing proliferation increased with exercise, noincrease in fusion of satellite cells into muscle fibers was apparent. Spinal cord transection resulted in a 25% decrease in myonuclear number, and exercise was not associated with a restoration of myonuclear number. The number of apoptotic nuclei was increased aftertransection, and exercise attenuated this increase. However, thedecrease in apoptotic nuclei with exercise did not significantly affectmyonuclear number. We conclude that apoptotic nuclear loss likelycontributes to loss of nuclei during muscle atrophy associated withspinal cord transection and that exercise can maintain muscle mass, atleast in the short term, without restoration of myonuclear number.

     

Regulation of phospholipase D by phosphorylation-dependent mechanisms.

The rapid production of phosphatidic acid following receptor stimulation has been demonstrated in a wide range of mammalian cells. Virtually every cell uses phosphatidylcholine as substrate to produce phosphatidic acid in a controlled reaction catalyzed by specific PLD isoforms. Considerable effort has been directed at studying the regulation of PLD activities and subsequent work has characterized a family of proteins including PLD1 and PLD2. Whereas both PLD enzymes are dependent on phosphatidylinositol 4, 5-bisphosphate for activity only the PLD1 isoform was strongly stimulated by the small GTPases ARF and RhoA and by protein kinase Calpha as well. A role for tyrosine kinase activities in the membrane recruitment of small GTPases, in the synthesis of phosphatidylinositol 4,5-bisphosphate and tyrosine phosphorylation of PLD1 and PLD2 has been uncovered. However, it still not clear exactly how tyrosine phosphorylation of proteins contributes to PLD activation in cells. Here we review the data linking tyrosine phosphorylation of proteins to the activation of PLD and describe recent finding on the sites and possible mechanisms of action of tyrosine kinases in receptor-mediated PLD activation. Finally, a model illustrating the potential complex interplay linking these signaling events with the activation of PLD is presented.     

Rates and Genomic Consequences of Spontaneous Mutational Events in Drosophila melanogaster

Because spontaneous mutation is the source of all genetic diversity, measuring mutation rates can reveal how natural selection drives patterns of variation within and between species. We sequenced eight genomes produced by a mutation-accumulation experiment in Drosophila melanogaster. Our analysis reveals that point mutation and small indel rates vary significantly between the two different genetic backgrounds examined. We also find evidence that ∼2% of mutational events affect multiple closely spaced nucleotides. Unlike previous similar experiments, we were able to estimate genome-wide rates of large deletions and tandem duplications. These results suggest that, at least in inbred lines like those examined here, mutational pressures may result in net growth rather than contraction of the Drosophila genome. By comparing our mutation rate estimates to polymorphism data, we are able to estimate the fraction of new mutations that are eliminated by purifying selection. These results suggest that ∼99% of duplications and deletions are deleterious—making them 10 times more likely to be removed by selection than nonsynonymous mutations. Our results illuminate not only the rates of new small- and large-scale mutations, but also the selective forces that they encounter once they arise.     

Mapping of a Chromosome 12 Region Associated with Airway Hyperresponsiveness in a Recombinant Congenic Mouse Strain and Selection of Potential Candidate Genes by Expression and Sequence Variation Analyses

In a previous study we determined that BcA86 mice, a strain belonging to a panel of AcB/BcA recombinant congenic strains, have an airway responsiveness phenotype resembling mice from the airway hyperresponsive A/J strain. The majority of the BcA86 genome is however from the hyporesponsive C57BL/6J strain. The aim of this study was to identify candidate regions and genes associated with airway hyperresponsiveness (AHR) by quantitative trait locus (QTL) analysis using the BcA86 strain. Airway responsiveness of 205 F2 mice generated from backcrossing BcA86 strain to C57BL/6J strain was measured and used for QTL analysis to identify genomic regions in linkage with AHR. Consomic mice for the QTL containing chromosomes were phenotyped to study the contribution of each chromosome to lung responsiveness. Candidate genes within the QTL were selected based on expression differences in mRNA from whole lungs, and the presence of coding non-synonymous mutations that were predicted to have a functional effect by amino acid substitution prediction tools. One QTL for AHR was identified on Chromosome 12 with its 95% confidence interval ranging from 54.6 to 82.6 Mbp and a maximum LOD score of 5.11 (p = 3.68×10⁻³). We confirmed that the genotype of mouse Chromosome 12 is an important determinant of lung responsiveness using a Chromosome 12 substitution strain. Mice with an A/J Chromosome 12 on a C57BL/6J background have an AHR phenotype similar to hyperresponsive strains A/J and BcA86. Within the QTL, genes with deleterious coding variants, such as Foxa1, and genes with expression differences, such as Mettl21d and Snapc1, were selected as possible candidates for the AHR phenotype. Overall, through QTL analysis of a recombinant congenic strain, microarray analysis and coding variant analysis we identified Chromosome 12 and three potential candidate genes to be in linkage with airway responsiveness.     

Transendothelial migration of colon carcinoma cells requires expression of E-selectin by endothelial cells and activation of stress-activated protein kinase-2 (SAPK2/p38) in the tumor cells.

Adhesion and migration of tumor cells on and through the vascular endothelium are critical steps of the metastatic invasion. We investigated the roles of E-selectin and of stress-activated protein kinase-2 (SAPK2/p38) in modulating endothelial adhesion and transendothelial migration of HT-29 colon carcinoma cells. Tumor necrosis factor alpha (TNF alpha) strongly increased the expression of E-selectin in human umbilical vein endothelial cells (HUVEC). This effect was independent of the activation of SAPK2/p38 induced by TNF alpha. Adhesion of HT-29 cells on a monolayer of HUVEC pretreated with TNF alpha was dependent on E-selectin expression but was independent of SAPK2/p38 activity of both HUVEC and tumor cells. The adhesion of HT-29 cells to E-selectin-expressing HUVEC led to the activation of SAPK2/p38 in the tumor cells as reflected by the increased phosphorylation of the actin-polymerizing factor HSP27 by mitogen-activated protein kinase 2/3, a direct target of SAPK2/p38. Moreover, a recombinant E-selectin/Fc chimera quickly increased the activation of SAPK2/p38 in HT-29 cells. Blocking the increased activity of SAPK2/p38 of HT-29 cells by SB203580 or by expressing a dominant negative form of SAPK2/p38 inhibited their transendothelial migration. Similarly, HeLa cells stably expressing a kinase-inactive mutant of SAPK2/p38 showed a decreased capacity to cross a layer of HUVEC. Overall, our results suggest that the regulation of transendothelial migration of tumor cells involves two essential steps as follows: adhesion to the endothelium through adhesion molecules, such as E-selectin, and increased motogenic potential through adhesion-mediated activation of the SAPK2/p38 pathway.     

Towards the preparation of radiolabeled 1-aryl-3-benzyl ureas: Radiosynthesis of [(11)C-carbonyl] AR-A014418 by [(11)C]CO(2) fixation

The highly selective glycogen synthase kinase-3 (GSK-3) inhibitor N-(4-methoxybenzyl)-N'-(5-nitro-1,3-thiazol-2-yl)urea (AR-A014418) was radiolabeled with carbon-11 ((11)C; half-life=20.4min) at the urea moiety via [(11)C]CO(2) fixation. Reaction of [(11)C]CO(2) with 4-methoxybenzylamine in the presence of a CO(2) fixating base was followed by dehydration with POCl(3) and addition of 2-amino-5-nitrothiazole to prepare [(11)C-carbonyl] AR-A014418. This reaction resulted in an 8% uncorrected radiochemical yield, based on [(11)C]CO(2), with high specific activity (4Ci/μmol) within 30min. An in vitro GSK-3β enzyme activity assay revealed that AR-A014418 (K(i)=770nM) is not as potent as previously claimed. The [(11)C]CO(2) fixation methodology described herein should prove generally applicable to preparing 1-aryl-3-benzyl-[(11)C-carbonyl] ureas as radiotracers for positron emission tomography.     

Development of new carbon-11 labelled radiotracers for imaging GABAA- and GABAB-benzodiazepine receptors

Two quinolines identified as positive allosteric modulators of γ-aminobutyric acid (GABA)(A) receptors containing the α(2) subunit, 9-amino-2-cyclobutyl-5-(6-methoxy-2-methylpyridin-3-yl)-2,3-dihydro-1H-pyrrolo[3,4-b]quinolin-1-one (4) and 9-amino-2-cyclobutyl-5-(2-methoxypyridin-3-yl)-2,3-dihydro-1H-pyrrolo[3,4-b]quinolin-1-one (5), were radiolabelled at the methoxy position with carbon-11 (half-life=20.4 min). These quinolines represent a new class of potential radiotracers for imaging the benzodiazepine site of GABA(A) receptors with positron emission tomography (PET). Both radiotracers were reliably isolated following reaction of their respective pyridinone/pyridinol tautomeric precursors with [(11)C]CH(3)I in clinically useful, formulated quantities (2.9% and 2.7% uncorrected radiochemical yield, respectively, relative to [(11)C]CO(2)) with high specific activities (>70 GBq μ mol(-1); >2 Ci μ mol(-1)) and high radiochemical purities (>95%). The radiosyntheses reported herein represent rare examples of selectively isolating radiolabelled compounds bearing [(11)C]2-methoxypyridine moieties. Although both radiotracers demonstrated promising imaging characteristics based on preliminary ex vivo biodistribution studies in conscious rodents, higher brain uptake was observed with [(11)C]5 and therefore this radiotracer was further evaluated. Carbon-11 labelled 5 readily penetrated the brain (>1 standard uptake value in cortical regions at 15 min post-injection of the radiotracer), had an appropriate regional brain distribution for GABA(A) receptors that appeared to be reversible, and did not show any appreciable radiometabolites in rat brain homogenates up to 15 min post-injection. Preadministration of flumazenil (1, 10 mg kg(-1)) or 5 (5 mg kg(-1)) effectively blocked >50% of [(11)C]5 binding to the GABA(A) receptor-rich regions, thereby suggesting that this radiotracer is worthy of further evaluation for imaging GABA(A) receptors. Additionally (R,S)-N-(1-(3-chloro-4-methoxyphenyl)ethyl)-3,3-diphenylpropan-1-amine, 6, an allosteric modulator of GABA(B) receptors, was efficiently labelled in one step using [(11)C]methyl iodide. Ex vivo biodistribution studies in conscious rats showed low brain uptake, therefore, efforts are underway to discover alternative radiotracers to image GABA(B). In conclusion, [(11)C]5 is worthy of further evaluation in higher species for imaging GABA(A) receptors in the central nervous system.