Timeline.

Clinical, neurophysiological, postural and cognitive assessment timeline for adolescents girls with AIS assisted with corrective postural exercises (AIS 1), adolescent girls with AIS treated with both corrective postural exercises and partial or full-time brace (AIS 2) and CTRL. Tables report for each group and for each phase (T0, T1, T2) which parameters will be recorded. The personal and clinical parameters and experimental protocol block descriptions are reported in the lower panel (1), (2), (3), (4), (5), (6), (7).</p

Guidelines to reproduce the experimental protocol.

Guidelines to reproduce the experimental protocol.</p

Blocks (iii)-(iv) graphical representation.

(A) target reaching; (B) forearm bisection task. The red circles symbolize the passive retroreflective markers. The subjects’ hand is represented in white and the experimenter’s hand is represented in black.</p

Three-Dimensional Reconstruction, by TEM Tomography, of the Ultrastructural Modifications Occurring in <i>Cucumis sativus</i> L. Mitochondria under Fe Deficiency

<div><p>Background</p><p>Mitochondria, as recently suggested, might be involved in iron sensing and signalling pathways in plant cells. For a better understanding of the role of these organelles in mediating the Fe deficiency responses in plant cells, it is crucial to provide a full overview of their modifications occurring under Fe-limited conditions. The aim of this work is to characterize the ultrastructural as well as the biochemical changes occurring in leaf mitochondria of cucumber (<i>Cucumis sativus</i> L.) plants grown under Fe deficiency.</p><p>Methodology/Results</p><p>Mitochondrial ultrastructure was investigated by transmission electron microscopy (TEM) and electron tomography techniques, which allowed a three-dimensional (3D) reconstruction of cellular structures. These analyses reveal that mitochondria isolated from cucumber leaves appear in the <i>cristae junction model</i> conformation and that Fe deficiency strongly alters both the number and the volume of cristae. The ultrastructural changes observed in mitochondria isolated from Fe-deficient leaves reflect a metabolic status characterized by a respiratory chain operating at a lower rate (orthodox-like conformation) with respect to mitochondria from control leaves.</p><p>Conclusions</p><p>To our knowledge, this is the first report showing a 3D reconstruction of plant mitochondria. Furthermore, these results suggest that a detailed characterization of the link between changes in the ultrastructure and functionality of mitochondria during different nutritional conditions, can provide a successful approach to understand the role of these organelles in the plant response to Fe deficiency.</p></div

Mitochondrial functionality of leaves from cucumber plants grown in control conditions (C) or under Fe deficiency (-Fe).

<p><b>(A)</b><i>In vivo</i> O₂ consumption rates expressed as μmol O₂ consumed min^-1 g FW^-1. The total O₂ consumption rate (IR), as well as the residual O₂ consumption, measured by using KCN (inhibiting cytochrome c oxidase) (IR + KCN) or KCN in combination with SHAM (inhibiting alternative oxidase) (IR + KCN +SHAM) were measured. Values represent mean ±SE of three independent measurements. <b>(B)</b> For each growth condition, the residual O₂ consumption (IR +KCN+SHAM), reported in (A), is shown as % value, with respect to correspondent IR. Mitochondrial respiration is the difference between IR and residual O₂ consumption. Student t-test was used to analyse statistical significance with respect to controls. *:p<0,05; **:p<0,01; ***:p<0,001. <b>(C)</b> Mitochondria (M) and chloroplasts (Cp) were purified from control plants by percoll gradient; the Cp and M fractions at 35/80% percoll and 23/40% percoll layers interfaces, respectively, are indicated with arrows in the gradient tubes. The purified Cp and M fractions were tested by western blot, by using antibodies against two protein markers: Toc33 and porin, specific for Cp, and M, respectively. 10 μg total proteins were loaded in each lane. <b>(D)</b> Western blot analysis of three different mitochondrial Fe-containing proteins (AOX, Rieske, cyt c) on mitochondria purified from leaves of control (C) or Fe-deficient (-Fe) plants. Porin was used as loading control; 10 μg total proteins were loaded in each lane. The results are representative of tree independent experiments.</p

Physiological characterization of leaves from cucumber plants grown in control condition (C) or under Fe deficiency (–Fe).

<p><b>(A)</b> Phenotype of 10 days old cucumber plants grown under control condition (C) or under Fe deficiency (-Fe). The following parameters have been evaluated, on expanded leaves of plants grown as in (A): <b>(B)</b> total chlorophyll concentration, expressed as mg chlorophyll g FW^-1; <b>(C)</b> O₂ evolution, expressed as μmol O₂ min^-1 g FW^-1; <b>(D)</b> net photosynthesis expressed as μmol CO₂ consumed s^-1 m^-2; <b>(E)</b> evapotranspiration, expressed as mmol H₂O s^-1m^-2; <b>(F)</b> stomatal conductance, expressed as mmol CO₂ s^-1m^-2; <b>(G)</b> manganese (Mn), iron (Fe), copper (Cu), zinc (Zn), molybdenum (Mo) concentrations, expressed as μg g DW^-1. Data are means ± SE of at least three independent experiments. Student t-test was used to analyse statistical significance with respect to controls. *:p<0,05; **:p<0,01; ***:p<0,001.</p

Determination of the cristae number <i>per</i> mitochondrion and the relative intracristae surface area (intracristae surface area <i>per</i> mitochondrion surface area, expressed as % ratio).

<p>Mean values ± SD are from analysis of one hundred mitochondria randomly selected from inclusions obtained from three independent biological samples. Student t-test was used to analyse statistical significance with respect to controls.</p><p>***:p<0,001</p><p>Determination of the cristae number <i>per</i> mitochondrion and the relative intracristae surface area (intracristae surface area <i>per</i> mitochondrion surface area, expressed as % ratio).</p

Transmission electron microscopy (TEM) analysis of leaf tissues from cucumber plants grown in control condition (C) or under Fe deficiency (-Fe).

<p>Control leaf tissues (<b>A-C</b>) and -Fe leaf tissues (<b>D-F</b>). All scale bars = 500 nm. M: mitochondria; C: chloroplast; V: vacuole.</p

Three-dimensional models of leaf mitochondria from cucumber plants grown in control condition (C) or under Fe deficiency (-Fe).

<p>Different colours were used for the rendering of the different suborganellar structures: magenta for inner membranes (IM), blue for outer membranes (OM), green for cristae and red for cristae junctions. <b>(A-D)</b> Tomographic slices of mitochondria from control (A,B) and Fe-deficient (C,D) plants; the inner membrane (IM), the outer membrane (OM), and the matrix enclosed by the IM are indicated; in particular, the rendering of the mitochondria structures from control (B) and Fe-deficient (D) plants, superimposed on the tomographic slices, is reported. <b>(E,F)</b> Three-dimensional models of the mitochondria from control (E) or Fe-deficient (F) plants. <b>(G,H)</b> Details of mitochondrial cristae junctions, in red, identified in mitochondrial from control (G) or Fe-deficient (H) plants. Scale bars 100 nm.</p