Some physical and mechanical characterization of Tunisian planted Eucalytus loxophleba and Eucalyptus salmonophloia woods

After the independence in 1957 and with the support of the FAO117, Eucalyptus species were planted in Tunisia in different arboreta throughout the country for close observation and adaptation to climate and soil. The objective of this study is to evaluate the physical and mechanical properties of two species planted in marginal area in Sousse (arboretum El Hanya) in the east of Tunisia (Eucalytus loxophleba and Eucalyptus salmonophloia). The moisture content, specific gravity and volumetric shrinkage were measured. The Mechanical tests were performed to evaluate the hardness, the static bending and the resistance to compression parallel to fiber direction. Preliminary results showed that Eucalytusloxophleba and Eucalyptus salmonophloia have a low dimensional stability. During the drying period, woods showed signs of collapses. On the other hand, both species were highly resistant to compression strength while they were lower on the static bending. Eucalytus loxophleba and Eucalyptus salmonophloia characteristics established within this study were similar to other Eucalyptus species from Tunisia, Morocco, Australia and Brazil. This wood may be used in furniture, structural material and/or biomass energy. (Résumé d'auteur

Ascl1 is present in a subpopulation of Type-1 stem cells and Type-2 progenitors in adult hippocampus.

<p>(A–D) Ascl1 is weakly detected in Nestin::GFP⁺GFAP⁺ Type-1 stem cells (arrowhead) or strongly detected in Nestin::GFP⁺GFAP⁻ Type-2 progenitors (arrow) in SGZ of adult <i>Nestin::GFP</i> mice. (E) Percentage of Ascl1^High or Ascl1^Low cells that express the markers Nestin::GFP and GFAP (Type-1) (dark shaded bars) or just the marker Nestin::GFP (Type-2) (grey shaded bars). 50 Ascl1⁺ cells were counted per mouse, n = 3 <i>Nestin::GFP</i> mice. (F) Ascl1 is in Type-1 and early Type-2 cells based on a current model of adult hippocampal neurogenesis <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0018472#pone.0018472-Kempermann1" target="_blank">[14]</a>. Scale bar  = 20 µm.</p

A subset of Ascl1 lineage cells in adult SVZ have long term self renewing properties in the generation of olfactory bulb neurons.

<p>(A–D) Ascl1 is detected in Nestin::GFP⁺GFAP⁺ cells (B cells) in the SVZ (A–B′) and in Nestin::GFP⁺GFAP⁻ C cells in SVZ (A–B′) and RMS (C,C′) in 8 week old <i>Nestin::GFP</i> transgenic mice. (D) Percentage of Ascl1^High or Ascl1^Low cells that express the markers Nestin::GFP and GFAP (dark shaded bars) or just Nestin::GFP (grey shaded bars) in the RMS and the SVZ. 25 Ascl1⁺ cells per mouse were counted in the RMS; 60 Ascl1⁺ cells per mouse in the SVZ, n = 3 <i>Nestin::GFP</i> mice. (E–E′) mRNA in situ with Ascl1 (E) or Cre (E′) probes in the adult SVZ. (F–T′) Immunofluorescence in <i>Ascl1^CreERT2/+</i>;<i>R26R^YFP/YFP</i> mouse brain sections harvested 7, 30, or 180 days post-TAM demonstrates Ascl1 derived cells along the SVZ-RMS-OB pathway (F–N). 7 days post-TAM most YFP⁺ cells were located in the SVZ, or along the RMS (F–H) and express Sox2 (O–O′) or DCX (P–P′). 30 or 180 days post-TAM YFP⁺ cells mature into neurons in the granule cell layer or the periglomerular layer of the OB (I, L, R–R′, and data not shown). However, many YFP⁺ cells remain as Sox2⁺ or DCX⁺ progenitors in the RMS or SVZ (J–K, M–N, Q–Q′, S–T′). Scale bars  = 50 µm (F–N), 10 µm (O–T′).</p

A subset of Ascl1 lineage cells continue to produce new granule neurons 30 days after initial Ascl1 expression in adult hippocampus.

<p>(A) Targeting strategy for <i>Ascl1^CreERT2/+</i> knock-in mice. (B) Quantification of the percentage of YFP⁺ cells co-labeled with stage-specific markers in hippocampus of adult <i>Ascl1^CreERT2/+;R26R^YFP/YFP</i> mice 7, 30, or 180 days post-TAM. 150–500 YFP⁺ cells per mouse were counted for each marker, n = 2 <i>Ascl1^CreERT2/+;R26R^YFP/YFP</i> mice per time point. (C–F′) 7 days post-TAM YFP⁺ cells co-express GFAP (and have Type-1 morphology), Sox2, or NeuroD1, but not NeuN. (G–J′) 30 days post-TAM YFP⁺ cells overlap with NeuN, but also can co-express GFAP or NeuroD1. (K–N′) 180 days post-TAM a subpopulation of YFP⁺ cells are still Type-1 cells by morphology and express GFAP and Sox2, whereas the majority of YFP⁺ cells express NeuN but not NeuroD1. (O–V) Neurogenesis in the SGZ dramatically decreases between 12 weeks and 34 weeks of age as seen in the decrease in DCX (P,T), NeuroD1 (Q,U) and Ki67 (R,V). Arrowheads indicate the few cells positive for these markers in the 34 week old mice. Notably, Sox2 does not decrease (O,S) so may label quiescent Type-1 like cells. Scale bars  = 50 µm (C,G,K), 10 µm (D–F′, H–J′, I–V).</p

Inducible deletion of Cdk5 in nestin-lineage cells via Tam P7-P9 does not change YFP+ cell number in hippocampus or cortex.

<p>(<b>A</b>) Paradigm for Tam administration. Representative images of YFP-stained hippocampi from P14 (<b>B</b>, <b>C</b>) and P21 (<b>D</b>, <b>E</b>) show YFP+ cells in corpus callosum (cc), stratum oriens (Or) and <i>radiatum</i> (Rad) in WT (top) and iCdk5 littermates (bottom; scale bar, s.b.=100 µm). Stereological quantification of YFP+ cells in the Or (<b>F</b>), Rad (<b>G</b>), cortex (<b>H</b>) and cc (<b>I</b>).</p

Inducible deletion of Cdk5 in nestin-lineage cells results in fewer YFP+ cells in the hippocampus and corpus callosum.

<p>(<b>A</b>) Paradigm of Tam administration. Representative images of the brain at P14 (<b>B</b>, <b>C</b>) and P21 (<b>D</b>, <b>E</b>) at P14 in WT (top) and iCdk5 littermates (bottom) highlighting cortex, corpus callosum (cc), and hippocampus with stratum oriens (Or), CA1, stratum radiatum (Rad) and molecular layer (Mol, s.b.=100 µm). Total number of YFP+ cells in Or (<b>F</b>), Rad (<b>G</b>; inset shows typical YFP+ Rad cell at P14, s.b.=10 µm), and cc (<b>H</b>).</p

Inducible expression of DTA in nestin-lineage stem/progenitor cells decreases the number of Ki67+ and DCX+ cells 12 days (d) post-tamoxifen (TAM), but DCX+ cell number is normalized 30d post-TAM.

<p><b>(A)</b> Experimental design of immunohistochemical study. TAM was administered to 5–6 week-old control or Cre+DTA+ mice for 5 consecutive days, and brains were collected 12d and 30d post-TAM. <b>(B)</b> Representative photomicrographs of the dentate gyrus from control and Cre+DTA+ mice 12d post-TAM stained with an antibody against Ki67. Scale bar = 200 um <b>(B,</b> applies to <b>B, D). (C)</b> Stereological quantification of Ki67+ cell number in the DG granule cell layer (GCL) 12d (control N = 5, Cre+DTA+ N = 4) and 30d (control N = 6, Cre+DTA+ N = 9) post-TAM. <b>(D)</b> Representative photomicrographs of the DG from control and Cre+DTA+ mice 12d post-TAM stained with antibody against DCX. <b>(E)</b> Stereological quantification of DCX+ cells in the DG GCL 12d (control N = 4, Cre+DTA+ N = 5) and 30d (control N = 6, Cre+DTA+ N = 7) post-TAM. <b>(F)</b> High magnification images of the DG from control and Cre+DTA+ mice 12d post-TAM stained with an antibody against DCX+. Scale bar = 50um. <b>(G)</b> Stereological quantification of postmitotic DCX+ cells in the DG GCL 12d (control N = 4, Cre+DTA+ N = 5) and 30d (control N = 6, Cre+DTA+ N = 7). Data are mean±SEM,.**p<0.01, *p<0.05 by unpaired, two-tailed Student’s t-test.</p

Cre+DTA+ mice tested less than or more than 4 weeks post-TAM show similar levels of locomotion and the absence of baseline anxiety-related behaviors.

<p><b>(A, B)</b> Locomotor activity (LM) in both short (<b>A,</b> Group 1, control N = 8, Cre+DTA+ N = 5) and long (<b>B,</b> Group 2, control N = 33, Cre+DTA+ N = 13) TAM-beh interval groups. Insets <b>A, B</b>: total beam breaks over 2 hr. Main panels <b>A, B</b>: beam breaks over 2 hr presented in 5 min bins. X axis * = main effect of time. Posthoc analysis (Bonferroni) revealed all points in main panels were significantly different than the initial locomotor activity data point. However, individual data point asterisks are omitted for clarity, as there was no main effect of genotype or interaction of time X genotype for either Group 1 or Group 2. <b>(C-F)</b> Time spent in the center <b>(C, E)</b> and periphery <b>(D, F)</b> during an open field test (OF) in short (<b>C-D,</b> Group 1, control N = 30, Cre+DTA+ N = 9) and long (<b>E-F,</b> Group 2, control N = 42, Cre+DTA+ N = 17) TAM-beh interval groups. <b>(G-J)</b> Number of transitions between light and dark chambers <b>(G, I)</b> and latency to enter the dark chamber <b>(H, J)</b> in the light/dark test (L/D test) in both short (<b>G-H,</b> Group 1, control N = 30, Cre+DTA+ N = 9) and long (<b>H-I,</b> Group 2, control N = 38, Cre+DTA+ N = 17) TAM-beh groups. Data are mean±SEM. ^<i>d</i>p<0.0001, two-way ANOVA with repeated measures and Bonferroni posthoc <b>(A, B)</b>. *p<0.05, unpaired two-tailed Student’s t-test (<b>insets A, B,</b> and <b>C-H</b>).</p

Experimental design of behavioral study.

<p>TAM was administered to 5–6 week-old control or Cre+DTA+ mice for 5 consecutive days. Behavioral testing began 12d (Group 1) or 33d post-TAM (Group 2), and continued as indicated through day 27 (Group 1, TAM-behavioral [TAM-beh] interval less than 4 weeks) or day 52 (Group 2, TAM-beh interval more than 4 weeks) post-TAM. Both groups were examined in the open field test (OF), locomotor test (LM), novelty induced hypophagia (NIH), light/dark test (L/D), juvenile social interaction test (JI), and tail suspension test (TST). Specifically for Groups 1 and Groups 2, OF was performed 12d or 33d post-TAM, LM 15d or 37d post-TAM, NIH 17-19d or 39-41d post-TAM, L/D 21d or 43d post-TAM, JI 22-25d or 44-47d post-TAM, and TST 27d or 52d post-TAM.</p

Relative to control mice, stress induced anxiety-like and depressive-like behavior are evident in Cre+DTA+ mice tested less than–but not more than– 4 weeks post-TAM.

<p><b>(A, B)</b> Latency to feed in the novelty induced hypophagia test (NIH) in control vs. Cre+DTA+ mice at short (<b>A,</b> Group 1, control N = 31, Cre+DTA+ N = 10) and long (<b>B,</b> Group 2, control N = 39, Cre+DTA+ N = 16) TAM-behavior intervals. <b>(C, D)</b> Total immobility time in the tail suspension test (TST) in control vs. Cre+DTA+ mice at short (<b>C,</b> Group 1, control N = 27, Cre+DTA+ N = 9) and long (<b>D,</b> Group 2, control N = 38, Cre+DTA+ N = 15) TAM-behavior intervals. <b>(E, F)</b> Interaction time during juvenile interaction training and test sessions in control vs. Cre+DTA+ mice at short (<b>E,</b> Group 1, control N = 30, Cre+DTA+ N = 9) and long (<b>F,</b> Group 2, control N = 40, Cre+DTA+ N = 17) TAM-beh intervals. Data = mean±SEM. *p<0.05, unpaired two-tailed Student’s t-test (A-D). ^<i>b</i>p<0.01,^<i>c</i>p<0.005, ^<i>d</i>p<0.0001, two-way ANOVA with repeated measures and Bonferroni posthoc.</p