Lamin A/C expression silencing leads to transformation.

<p><b>(A)</b> Lamin A/C was monitored by WB after shRNA depletion by SK-N-SH-lamin-A/C-shRNA or SK-N-SH-scramble-shRNA. Actin was used as a loading control. <b>(B)</b> Cumulative cell numbers of SK-N-SH-lamin-A/C-shRNA and SK-N-SH-scramble-shRNA, respectively. Three independent experiments were performed in triplicate (n = 9) using cells at less than eight passages, and error bars represent the s.d. Statistical significance was assessed using Student’s t-test;(p<0.01). Representative images showing differential cell growth on day 8 of the experiment. Scale bar, 100μm. <b>(C)</b> Wounding assay of confluent cell layers of SK-N-SH-lamin-A/C-shRNA or SK-N-SH-scramble-shRNA.Number of cells that migrated into a delimited wound area after 12h is plotted. Cells in three defined area per group per experiment were quantified in three independent experiments with three technical replicates. Error bars represent the s.d. Statistical significance was assessed using Student’s t-test; (*p<0.01). Representative images; scale bar, 100μM. <b>(D)</b> Quantification of the Matrigel chamber migration assay for SK-N-SH-lamin-A/C-shRNA and SK-N-SH-scramble-shRNA. Error bars represent the s.d. (n = 9). Statistical significance was assessed by Student’s t-test; (**p<0.01). Scale bar, 10μM. <b>(E)</b> SK-N-SH-lamin-A/C increased the number of colonies in soft agar compared with the SK-N-SH-scramble-shRNA control. The number of colonies per well was counted and plotted. Three independent experiments (n1 = 3) with 3 replicates per experiment (n2 = 9) were performed. Error bars represent the s.d. (n = 9). Statistical significance was assessed using Student’s t-test (**p<0.01). Scale bar, 100μM.</p

Progerin introduction in SK-N-DZ cells induces changes in cytoskeletal components and mechanical properties.

<p><b>(A)</b> Immunofluorescence staining showing Progerin. <b>(B)</b> Changes in β-actin, F-actin, vimentin filaments, and α-tubulin after Progerin introduction. Progerin is shown in green; β-actin, F-actin, vimentin, and α-tubulin are shown in red; and DNA is stained in blue (DAPI). Scale bar, 10μm. <b>(C)</b> Representative AFM images of height measurement in SK-N-DZ-ev cells. In red and blue height profile across lines in different cells from height AFM image are shown Scale bar, 10μm <b>(D)</b>The AFM tip is positioned directly above the lamellar region. Scale bar 10μm <b>(E)</b> Normalized histogram of the data obtained from both groups for the lamellar region. Each histogram was fitted to a Gaussian curve to obtain the mean value and the standard deviation of Young’s Modulus. Three independent experiments (n1 = 3) with 3 replicates per experiment (n2 = 9) were performed. A Student’s t-test confirmed that the difference was significant (**p<0.01).</p

Lamin A/C reintroduction induces reorganizational changes in the different cytoskeletal components.

<p>Immunofluorescence staining showing changes in SK-N-DZ-lamin-A/C compared with SK-N-DZ-ev cells in <b>(A)</b> β-actin filaments, <b>(B)</b> F-actin filaments, <b>(C)</b> Vimentin filaments, and <b>(D)</b> α<b>-</b>tubulin. Lamin A/C is shown in green; β -actin, F-actin, vimentin, and α-tubulin are shown in red.DNA is stained in blue (DAPI). Scale bar, 10μM.</p

Methylation analysis of the Lamin A/C gene in neuroblastoma patients.

<p>Degree of methylation in neuroblastoma tumours at Lamin A/C locus is visualized as colour scale from black(very low methylation) to yellow (fully methylated) in a heatmap, where rows represent CpGs represented on the array (with array probe ID) and columns represent patients. As indicated by coloured bars on the right (green/red), the heatmap illustrates the genomic locations of the Lamin A/C gene from top to bottom, while array locations marked in green represent positions upstream of the TSS, and red marked locations represent the coding sequence In addition, the promoter region -37 to +333 is highlighted in blue. The column tree vizualizes the hierachical clustering on patients. On top of the heatmap clinical parameters: risk (white = low risk, yellow = intermediate risk, blue = high risk) and international neuroblastoma staging system (Inns) stage are given (see colour legend for 6 stages).</p