Clinical Characteristics of NEC, SIP and Surg-CTL Infants.

<p>Results are expressed as number (%) or median (interquartile range). Standard antibiotic regimens in our neonatal unit for treatment of NEC and SIP, included: (i) vancomycin, aminoglycoside or 3rd generation cephalosporin and metronidazole, or (ii) vancomycin and meropenem, depending on the severity of intra-abdominal pathology and microbial culture/sensitivity. Bold values indicate statistical significance (<i>P</i><0.016) after the Bonferroni correction.</p><p>^{<b><i>#</i></b>} Comparison not significant after the Bonferroni correction.</p><p>Clinical Characteristics of NEC, SIP and Surg-CTL Infants.</p

miRNA and mRNA Targets in Regulatory Networks of NEC and SIP.

<p>qPCR-confirmed significant over-expression (↑) and under-expression (↓) in NEC or SIP tissues compared with Surg-CTL tissues.</p><p>*Significant inverse correlation (<i>P</i><0.05) of miRNA/mRNA pairs in same NEC or SIP specimens.</p><p>miRNA and mRNA Targets in Regulatory Networks of NEC and SIP.</p

Pathway traced network of miRNA/mRNA pairs in NEC.

<p>A functional network on the association of selected miRNAs and potential target mRNAs was generated by the MetaCore software and sequence-based prediction. These miRNA/mRNA pairs were regulated at opposite directions and/or exhibited significant correlation in same samples. qPCR-validated expression changes of miRNAs or mRNAs are shown as red and blue color circles, representing up- and down-regulation. Green, red and grey arrows between target genes represent positive, negative and unspecified interactions, respectively. Big arrows between miRNA/mRNA pairs indicate experimentally proven relationships. Transcription factors and functional categories, including angiogenesis, arginine metabolism, cell adhesion, chemotaxis and inflammation, ECM remodeling, hypoxia/oxidative stress, and muscle contraction are highlighted in different colors. miRNAs which exhibited significant inverse correlation with specific mRNAs are underlined.</p

Principal component analysis of miRNA expression profiles in NEC, SIP and Surg-CTL small bowel tissues.

<p>miRNA expression profiles of NEC (n = 4), SIP (n = 4) and Surg-CTL (n = 4) tissues were analyzed by principal component analysis using the Partek Genomics Suite. The ellipsoids represent 95% confidence intervals of NEC (red), SIP (blue) and Surg-CTL (green) clusters. Each dot represents an experiment dataset. The axes correspond to principal component 1 (PC1; x-axis), PC2 (y-axis) and PC3 (z-axis).</p

Differentially Expressed miRNAs in Small Bowel Tissues from Infants with NEC (n = 10), SIP (n = 10) and Surg-CTL (n = 10) by qPCR Assay.

<p>Bold values indicate statistical significance <i>(P</i> < 0.0167).</p><p>^{<b><i>#</i></b>} Comparison not significant after Bonferroni correction.</p><p>Differentially Expressed miRNAs in Small Bowel Tissues from Infants with NEC (n = 10), SIP (n = 10) and Surg-CTL (n = 10) by qPCR Assay.</p

Clinical characteristics of NEC, SIP and surgical control patients recruited for mRNA analysis.

<p>Results are expressed as % or median (interquartile range).</p><p>Note: All surgical specimens were of ileal origin, except 1 SIP specimen was from the descending colon.</p

mRNA expression levels of target genes in resected intestinal tissues of NEC and SIP infants.

<p>Expression levels of IL-6, Ang-2, ErbB3, IL1-RII, and uPAR in resected intestinal tissues from NEC (n = 7) and SIP (n = 6) infants were quantified by qPCR and compared with surgical control tissues (n = 6). Results showed that IL-6, IL1-RII and uPAR were significantly higher (<i>P</i><0.01) in NEC tissues, compared with surgical control tissues (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036977#pone-0036977-g003" target="_blank">Figure 3A, D and E</a>). Results are presented as median, interquartile range and range of expression levels relative to β-actin.</p

Clinical characteristics of NEC and SIP patients recruited for plasma analyses.

<p>Results are expressed as % or median (interquartile range).</p><p>Note: N/A = not applicable.</p

mRNA expression levels of target genes in FHs-74 Int cell line in response to LPS and PAF.

<p>FHs-74 Int cells were cultured in the presence of LPS (100 ng/mL) or PAF (25 mM) for 6 h individually or in combination. qPCR analysis of target mRNA expressions showed that combined treatment with LPS and PAF significantly increased levels of Ang-2, IL1-RII and uPAR (n = 4; <i>P</i><0.05), whereas single stimulant, LPS or PAF, did not alter the expression levels. Results are presented as mean and SEM of expression levels relative to β-actin.</p

Comparison of plasma levels of target proteins in NEC, SIP and respective control infants by ELISA.

<p>Levels of IL-6, Ang-2, sErbB3, sIL1-RII, and suPAR in NEC (n = 13) and SIP (n = 8) infants were quantified by ELISA and compared with those in respective gestational age-matched control (CTL) infants (NEC CTL, n = 13; SIP CTL, n = 8). Levels of IL-6, Ang-2, sIL1-RII and suPAR were significantly higher in NEC infants compared with NEC-CTL (<i>P</i><0.01) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036977#pone-0036977-g002" target="_blank">Figure 2A, B, D, E</a>) and SIP infants had significantly higher level of sErbB3 compared with NEC infants (P<0.05) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036977#pone-0036977-g002" target="_blank">Figure 2C</a>). Results are presented as median, interquartile range and range.</p