Venn diagram displaying the total number of variants in the targeted genes (coding exons and UTRs, with 10 bp of intronic flanking regions), which were identified by each enrichment method, in Patient #1 and Patient #2.

<p>The presence of mutations colored in red was not confirmed by Sanger sequencing while the presence of mutations colored in green was confirmed by Sanger sequencing (see also <b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0143373#pone.0143373.s001" target="_blank">S1 Fig</a></b>).</p

Percentage of base-pairs covered by 8, 20, 50 or 100 sequence reads in coding regions of targeted genes (with 10 bp of intronic flanking regions), according to each enrichment method.

<p>Percentage of base-pairs covered by 8, 20, 50 or 100 sequence reads in coding regions of targeted genes (with 10 bp of intronic flanking regions), according to each enrichment method.</p

Number of coding exons (with 10 bp of intronic flanking regions; <i>n</i>_<i>total</i> = 576) exhaustively covered by 8, 20, 50 or 100 sequence reads, according to each enrichment method.

<p>Number of coding exons (with 10 bp of intronic flanking regions; <i>n</i>_<i>total</i> = 576) exhaustively covered by 8, 20, 50 or 100 sequence reads, according to each enrichment method.</p

Percentage of base-pairs covered by 8, 20, 50 or 100 sequence reads in UTRs of targeted genes, according to each enrichment method.

<p>Percentage of base-pairs covered by 8, 20, 50 or 100 sequence reads in UTRs of targeted genes, according to each enrichment method.</p

Number of UTRs (<i>n</i>_<i>total</i> = 197) exhaustively covered by 8, 20, 50 or 100 sequence reads, according to each enrichment method.

<p>Number of UTRs (<i>n</i>_<i>total</i> = 197) exhaustively covered by 8, 20, 50 or 100 sequence reads, according to each enrichment method.</p

Role of oxidative stress on the ER stress markers expression induced by human oxidized LDL.

<p>Expression of <i>CHOP</i>/<i>Chop</i> and <i>P58IPK</i>/<i>p58IPK</i> in <b>(a)</b> MIN6 and <b>(b)</b> human islets cells exposed to hydrogen peroxide (H₂O₂) 150 μM at indicated times. The expression of the two genes was quantified in <b>(c)</b> MIN6 cells or <b>(d)</b> human islets that were co-incubated with vehicle (V), human native LDL (nLDL) or oxidized LDL (oxLDL) 2 mmol/l cholesterol supplemented by either DMSO (control, open bar) or N-acetylcystein (NAC, filled bar) 1 mmol/l for MIN6 and 10 mmol/l for human islets cells. The results were normalized against <i>Rplp0</i>/<i>RPLP0</i> and the expression levels from cells cultured with vehicle were set to 100%. Data are the mean ± SEM of 3 independent experiments performed in triplicate (***, P<0.001; **, P<0.01).</p

Activation of ER stress by human oxidized LDL.

<p><b>(a)</b> Western blotting analysis comparing changes in PERK, eIF2 and Ire1α and their phosphorylated forms (p). Total proteins were prepared from MIN6 cells cultured with 2 mmol/l cholesterol oxidized LDL (oxLDL) for the indicated times and 1 μmol/l thapsigargin (Thaps) for 6 h. The α-tubulin protein served as loading control. The figure is a representative experiment out of three. Measurement of <i>CHOP</i>/<i>Chop</i>, <i>P58IPK</i>/<i>p58IPK</i> and <i>ATF4</i>/<i>Atf4</i> mRNA levels in (<b>b)</b> and <b>(d)</b> MIN6 and (<b>c)</b> and <b>(e)</b> isolated human islets cells cultured with oxidized LDL. The mRNA level was quantified by quantitative real-time PCR in MIN6 or isolated human islets cells cultured for 48 h with vehicle (V), native (nLDL) or oxidized LDL (oxLDL). The PBA chemical chaperone 2.5 mmol/l were added in the cells cultured with oxidized LDL (oxLDL, <i>filled bar</i>). For <b>d</b>) and <b>e</b>), cells were cultured with oxidized LDL plus 1 mmol/l cholesterol HDL (<i>filled bar</i>). The mRNA level was normalized against the housekeeping acidic ribosomal phosphoprotein P0 gene (<i>RPLP0</i>/<i>Rplp0</i>) and the expression levels from cells cultured with vehicle were set to 100%. Data are the mean of ± SEM of 3 independent experiments performed in triplicate (***, P<0.001; **, P<0.01).</p

Effects of 4-BPA chemical chaperone on the loss of insulin expression caused by human oxidized LDL.

<p>The preproinsulin mRNA was quantified in MIN6 cells (<b>a)</b> and (<b>b</b>) human islets. Cells were exposed to vehicle (V), human native LDL (nLDL) or oxidized LDL (oxLDL) 2 mmol/l cholesterol, in the presence or absence of PBA 2.5 mmol/l (filled bars) for 48 h. The mRNA levels were normalized against the <i>Rplp0/RPLP0</i> and the expression levels from cells cultured with vehicle were set to 100%. Data are the mean of ± SEM of 3 independent experiments performed in triplicate (*, P<0.05).</p

Effects of Chop silencing in apoptosis caused by human oxidized LDL.

<p>MIN6 cells were transfected with a control RNA si-GFP duplex (Ctrl, <i>open bar</i>) or with siCHOP (<i>filled bar</i>). Thereafter, the cells were cultured for 72 h with vehicle (V) or oxidized LDL (oxLDL) 2 mmol/l cholesterol. The fraction of cells undergoing apoptosis was determined by scoring the percentage of cells displaying pyknotic nuclei. Data are the mean of ± SEM of 3 independent experiments performed in triplicate (**, P<0.01).</p

Schematic representation of mechanism coupling oxidized LDL to beta cell dysfunction and death.

<p>Schematic representation of mechanism coupling oxidized LDL to beta cell dysfunction and death.</p