Differential gene expression in controls due to medication.

<p>The x-axis the average change in expression following the use of statins and aspirin depicted as 2expΔΔCT. Values are given as mean + SD. A relative fold change of ≤1 or −1 indicates no effect. For ADRB2 and ABCG1 the changes were significant; p = 0.04 and p<0.001 respectively.</p

Differential gene expression in patients versus controls determined qPCR in the replication set.

<p>The X-axis depicts the relative fold changes in gene expression in the patients compared to controls. depicted as the qPCR 2exp ΔΔCT change. Values are given as mean + SD. A relative fold change of ≤1 or −1 indicates no effect.</p

Differential gene expression in patients versus controls validated by qPCR in the discovery set.

<p>The X-axis depicts the relative fold changes in gene expression in the patients compared to controls. For each gene the first (pale grey) bar depicts the fold change in microarray expression levels, the second (mid grey) bar shows the change in microarray intensities and the final (black) bar shows the qPCR 2exp ΔΔCT change. Values are given as mean + SD. A relative fold change of ≤1 or −1 indicates no effect.</p

Baseline characteristics Patients and Controls.

<p>Baseline characteristics Patients and Controls.</p

Characteristics of the core pedigree included in linkage analysis.

<p>Age is expressed as mean ± standard deviation and data are expressed as number (N). Lipid values are expressed as median with interquartile range (IQR). Pedigree members with an event use lipid lowering medication. LDL = low density lipoprotein; HDL = high density lipoprotein.</p

Clinical characteristics of the core pedigree included in linkage analysis.

<p>AMI = Acute Myocardial Infarction; AP = Angina Pectoris; PTCA = Percutaneous Transluminal Coronary Angioplasty; CABG = Coronary Artery Bypass Graft; CVA = Cerebrovascular Accident. Simva = Simvastatin, Rosuva = Rosuvastatin, Prava = Pravastatin and Atorva = Atorvastatin. BMI = body mass index (kg/cm²).</p>†<p> = index case. Subjects were considered smokers if they were current smokers or when they quitted smoking within the last 5 years.</p

Structures obtained from molecular dynamic simulations of the mutant p.Ser307Cys KERA protein.

<p>Molecular dynamic simulations were performed as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098289#pone.0098289.s001" target="_blank">File S1</a>. The residue Cys303 is highlighted in yellow, Cys343 in green, Ser307 in cyan and Cys307 residue in blue. <b>A</b>, Structure of wild-type KERA. <b>C</b> Structure of the KERA mutant p.Ser307Cys. <b>B</b>, A detailed view of the C-terminal part of wild type KERA highlighting the Cys303–Cys343 disulphide bond. <b>D</b>, Possible structural effects of the substitution of a serine for a cysteine at residue 307 showing a favourable Cys303–Cys307 disulphide bond. Consequently, Cys343 is available for binding with cysteine residues of other proteins.</p

Co-localization of KERA with CXCL1 and CD3 positive type I helper T lymphocytes was assessed in human plaque segments.

<p><b>A</b>: triple stain with KERA (blue), CXCL1 (red) and CD3 positive cells (brown). <b>B</b>: Spectral Imaging of triple staining. <b>C</b>: The yellow staining demonstrates that these cells are positive for all three components.</p

Expression of KERA in atherosclerotic tissue in <i>Apoe^−/−</i> mice after induction of atherosclerosis by collar placement.

<p><b>A–B</b>, Early (week = 2, A) and advanced (week = 8, B) atherosclerotic tissue from murine carotid arteries were stained for KERA (brown) and hematoxylin (blue). While present mainly near endothelial cells in early lesion, KERA is predominantly present in the matrix of the plaque at more advanced lesions. <b>C–D</b>, KERA expression overtime in <i>Apoe^−/−</i> mice with collar placement show significant correlation with plaque size (r² = 0.69, P<0.0001).</p

KERA is expressed in atherosclerotic but not in non-diseased arteries.

<p><b>A</b>, KERA is not expressed in tonsil tissue; <b>B</b>: KERA is expressed in corneal tissue <b>C</b>: KERA is not expressed in mammary artery. <b>D</b>: KERA is expressed in arterial segments obtained from a patient with the <i>KERA</i> mutation.</p