Excerpts from SUPPORT informed consent forms.

<p>^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155005#t002fn001" target="_blank">*</a>} A selection of statements extracted from the 22 institutional review board-approved SUPPORT consent forms that characterized the oxygen management interventions are displayed in a tabular format. Institutions are blinded in this table.</p

Usual care oxygen saturation (SpO₂) target ranges in 14 centers for preterm infants (24 to 27 weeks) compared to low and high SpO₂ target ranges in SUPPORT, COT and BOOST II.

<p>In panel A, the usual care SpO₂ intended target ranges from 14 neonatal intensive care units (NICUs) in the AVIOx study are plotted (dark grey vertical bars). On the X-axis, letters randomly assigned to each of the 14 NICUs in the AVIOx study are provided. NICUs are ordered on the x-axis from the lowest to the highest lower limit of the target range employed. The bar for all centers/patients combined delineates the medians of the upper limits and lower limits of the 14 ranges. The light grey-shaded area represents the low target range studied in the five clinical trials. Panel B shows the relationship of the lower limit of the target ranges (X-axis) to their total width or size (Y-axis) for the individual usual care neonatal intensive care units (NICUs) of the AVIOx study (open circles) and for the low (light grey triangle) and high (dark grey square) SpO2 target range arms of the clinical trials. Panel C shows the relationship of the lower limit (X-axis) to the upper limit (Y-axis) of the same target ranges. The ellipse represents the 95% prediction region for this relationship in the 14 usual care NICUs (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155005#sec006" target="_blank">METHODS</a>).</p

Usual care median achieved SpO₂ values in 14 care centers for preterm infants receiving oxygen therapy compared to low and high SpO₂ arms in COT and BOOST II.

<p>Panel A compares the median achieved SpO₂ values, and interquartile range, of each of these 14 usual care NICUs with the intended SpO₂ range established in the same NICUs (represented by dark grey vertical bars). Panel B compares median achieved SpO₂ values, and interquartile range, from these 14 NICUs to the target ranges of the low (lower grey-shaded area) and high (upper grey-shaded area) SpO₂ arms of the SUPPORT, BOOST II and COT trials. In panel C median achieved SpO₂ values are plotted in 7 low and 7 high SpO₂ arms during the BOOST II and COT trials, as well as in the 14 NICUs included in the AVIOx study; the latter are separated into 9 centers using a lower limit of the intended SpO₂ target range at or below 88% and 5 centers using a lower limit of the intended target SpO₂ range ≥90%. This separation was done to compare usual care to the clinical trial arms with comparable lower limits of the intended target SpO₂ ranges. For each of four compared groups, the median (thick horizontal line) and the mean (thin horizontal line) of the achieved SpO₂ values are plotted. The number of study arms is 7 for each target range because in three trials (BOOST II Australia and U.K. and COT) the data were provided separately from before and after recalibration of the Masimo pulse oximeters.</p

Usual care oxygen supplementation practices in preterm infants obtained from surveys, randomized controlled trials and observational studies.

<p>Data on usual care SpO₂ target ranges used and patient or subject characteristics are displayed when available from surveys, randomized controlled trials and observational trials.</p

Usual Care and Informed Consent in Clinical Trials of Oxygen Management in Extremely Premature Infants

<div><p>Objective</p><p>The adequacy of informed consent in the Surfactant, Positive Pressure, and Pulse Oximetry Randomized Trial (SUPPORT) has been questioned. SUPPORT investigators and publishing editors, heads of government study funding agencies, and many ethicists have argued that informed consent was adequate because the two oxygen saturation target ranges studied fell within a range commonly recommended in guidelines. We sought to determine whether each oxygen target as studied in SUPPORT and four similar randomized controlled trials (RCTs) was consistent with usual care.</p><p>Design/Participants/Setting</p><p>PubMed, EMBASE, Web of Science, and Scopus were searched for English articles back to 1990 providing information on usual care oxygen management in extremely premature infants. Data were extracted on intended and achieved oxygen saturation levels as determined by pulse oximetry. Twenty-two SUPPORT consent forms were examined for statements about oxygen interventions.</p><p>Results</p><p>While the high oxygen saturation target range (91 to 95%) was consistent with usual care, the low range (85 to 89%) was not used outside of the SUPPORT trial according to surveys and clinical studies of usual care. During usual care, similar lower limits (< 88%) were universally paired with higher upper limits (≥ 92%) and providers skewed achieved oxygen saturations toward the upper-end of these intended ranges. Blinded targeting of a low narrow range resulted in significantly lower achieved oxygen saturations and a doubling of time spent below the lower limit of the intended range compared to usual care practices. The SUPPORT consent forms suggested that the low oxygen saturation arm was a widely practiced subset of usual care.</p><p>Conclusions</p><p>SUPPORT does not exemplify comparative effectiveness research studying practices or therapies in common use. Descriptions of major differences between the interventions studied and commonly practiced usual care, as well as potential risks associated with these differences, are essential elements of adequate informed consent.</p></div

Percentage of time spent below an oxygen saturation (SpO₂) value of 85%, and below the intended SpO₂ range.

<p>The mean ± SE of the percentage of time spent below a true SpO₂ value of 85% for 7 low SpO₂ arms from the BOOST II and COT trials (light grey bar) is plotted <i>versus</i> the time spent below 85% for 7 high SpO₂ arms from the same trials (dark grey bar) <i>versus</i> the time spent below the lower limit of the intended range for the nine usual care neonatal intensive care units from the AVIOx study that had with comparable lower limits of the intended range (median lower limit of 88%; white bar). The number of low and high SpO₂ arms is 7 because in three trials (BOOST II Australia and U.K. and COT), the data were provided separately for before and after recalibration of the Masimo pulse oximeters.</p

Late Multiple Organ Surge in Interferon-Regulated Target Genes Characterizes Staphylococcal Enterotoxin B Lethality

<div><p>Background</p><p>Bacterial superantigens are virulence factors that cause toxic shock syndrome. Here, the genome-wide, temporal response of mice to lethal intranasal staphylococcal enterotoxin B (SEB) challenge was investigated in six tissues.</p><p>Results</p><p>The earliest responses and largest number of affected genes occurred in peripheral blood mononuclear cells (PBMC), spleen, and lung tissues with the highest content of both T-cells and monocyte/macrophages, the direct cellular targets of SEB. In contrast, the response of liver, kidney, and heart was delayed and involved fewer genes, but revealed a dominant genetic program that was seen in all 6 tissues. Many of the 85 uniquely annotated transcripts participating in this shared genomic response have not been previously linked to SEB. Nine of the 85 genes were subsequently confirmed by RT-PCR in every tissue/organ at 24 h. These 85 transcripts, up-regulated in all tissues, annotated to the interferon (IFN)/antiviral-response and included genes belonging to the DNA/RNA sensing system, DNA damage repair, the immunoproteasome, and the ER/metabolic stress-response and apoptosis pathways. Overall, this shared program was identified as a type I and II interferon (IFN)-response and the promoters of these genes were highly enriched for IFN regulatory matrices. Several genes whose secreted products induce the IFN pathway were up-regulated at early time points in PBMCs, spleen, and/or lung. Furthermore, IFN regulatory factors including Irf1, Irf7 and Irf8, and Zbp1, a DNA sensor/transcription factor that can directly elicit an IFN innate immune response, participated in this host-wide SEB signature.</p><p>Conclusion</p><p>Global gene-expression changes across multiple organs implicated a host-wide IFN-response in SEB-induced death. Therapies aimed at IFN-associated innate immunity may improve outcome in toxic shock syndromes.</p></div

Pulmonary pathology: hematoxylin and eosin (H&E) stain, TUNEL assay and immunohistochemistry staining for nitrotyrosine and polyADP-ribose.

<p>Compared to control animals at 24 h, staphylococcal enterotoxin B (SEB) challenge caused a multifocal, minimal to mild perivascular, peribronchiolar, interstitial and subpleural lymphohistiocytic inflammatory infiltrate. At 48 h a coalescing, neutrophil-predominant infiltrate was seen in SEB exposed animals that now extended into alveoli. Multiple vessel walls 48 h after SEB exposure contained neutrophilic fragments (arrows) consistent with vasculitis (H&E inset, SEB 48 h). Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay demonstrated an increase in bronchiolar apoptotic cells (arrows) after SEB challenge compared to control that was significant at 24 h post-exposure (2.93±0.12 <i>versus</i> 0.06±0.06 cells/HPF; <i>p<0.001</i>). Immunohistochemistry for nitrotyrosine was not different comparing SEB to control with all specimens showing faint staining (arrows) of alveolar epithelium, small vessel endothelium and alveolar macrophages. In contrast, immunohistochemistry for polyADP-ribose (PAR) showed increased staining associated with SEB exposure that was mostly proportional to the increase in inflammatory cellularity. At 48 h, hypertrophied alveolar epithelial cells (arrows) stained prominently for PAR.</p

Quantitative real-time PCR (qRT-PCR) confirmation of tissue-wide changes in gene expression.

<p>Nine genes were quantitated by qRT-PCR across all 6 tissues at 24 h. (A) Scatter plot of all genes and tissues tested comparing microarray and qRT-PCR fold-change from control. As shown by the line of identify (x  =  y), qRT-PCR typically returned higher fold-change results than microarray. Gene specific results, colored by tissue (see Legend), are shown as follows: (B) Cxcl11; (C) Herc6; (D) Irf1; (E) Irf8; (F) Irgm1; (G) Parp12; (H) Stat1; (I) Xaf1; and (J) Zbp1. All qRT-PCR results met the >1.5 fold-change cut-off for gene selection, except for measurements of Irf8 in PBMCs and spleen. However, Irf8 similarly failed selection by microarray in these tissues at 24 h. Four samples were tested per tissue. Each PBMC sample represented a pool of multiple mice while each organ sample came from an individual mouse.</p

Thematic analysis, interferon (IFN) response subtype classification, and promoter analysis for binding matrices responsive to IFN.

<p>(A) Canonical pathways significantly associated with the all-tissue response to staphylococcal enterotoxin B (SEB) challenge. Seventy-nine unique genes were recognized by the Ingenuity Pathway Analysis® (IPA®) database and mapped to IFN signaling, antigen presentation, and activation of IFN regulatory factor (IRF) by cytosolic pattern recognition receptors, among the other canonical pathways shown. (B) Classification of genes significantly up-regulated across all tissues by IFN response subtype. Note that for <i>Mus musculus</i>, the Interferome v2.01 database contained 1655 Type I genes, 1413 Type II genes, and no Type III genes. (C) IFN pathway-driven regulatory binding sites identified in the promoters of genes regulated across all tissues. Of 81 promoter regions analyzed (from +500 to −1500 bp), 68 were found to contain IFN-driven regulatory matrices as shown. Results generated by Interferome v2.01 using TRANSFAC® Professional (2012) matrices and the MATCH™ algorithm.</p