Corticosteroids and other anti-Inflammatory strategies in pediatric heart surgery:a national survey of practice

Background: The role of steroids to mitigate the deleterious effects of pediatric cardiopulmonary bypass (CPB) remains a matter of debate; therefore, we aimed to assess preferences in administering corticosteroids (CSs) and the use of other anti-inflammatory strategies in pediatric cardiac surgery. Methods: A 19-question survey was distributed to consultants in pediatric cardiac anesthesia from 12 centers across the United Kingdom and Ireland. Results: Of the 37 respondents (37/60, 62%), 24 (65%) use CSs, while 13 (35%) do not use steroids at all. We found variability within 5 (41%) of the 12 centers. Seven consultants (7/24, 29%) administer CSs in every case, while 17 administer CSs in selected cases only (17/24, 71%). There was variability in the dose of steroid administration. Almost all consultants (23/24, 96%) administer a single dose at induction, and one administers a two-dose regimen (1/24, 4%). There was variability in CS indications. Most consultants (24/37, 66%) use modified ultrafiltration at the conclusion of CPB. Fifteen consultants (15/32, 47%) report the use of aprotinin, while only 3 use heparin-coated circuits (3/24, 9%). Conclusions: We found wide variability in practice in the administration of CSs for pediatric cardiac surgery, both within and between units. While most anesthetists administer CSs in at least some cases, there is no consensus on the type of steroid, the dose, and at which patient groups this should be directed. Modified ultrafiltration is still used by most of the centers. Almost half of consultants use aprotinin, while heparin-coated circuits are infrequently used. </jats:sec

STAT3 Regulates Monocyte TNF-Alpha Production in Systemic Inflammation Caused by Cardiac Surgery with Cardiopulmonary Bypass

BACKGROUND: Cardiopulmonary bypass (CPB) surgery initiates a controlled systemic inflammatory response characterized by a cytokine storm, monocytosis and transient monocyte activation. However, the responsiveness of monocytes to Toll-like receptor (TLR)-mediated activation decreases throughout the postoperative course. The purpose of this study was to identify the major signaling pathway involved in plasma-mediated inhibition of LPS-induced tumor necrosis factor (TNF)-α production by monocytes. METHODOLOGY/PRINCIPAL FINDINGS: Pediatric patients that underwent CPB-assisted surgical correction of simple congenital heart defects were enrolled (n = 38). Peripheral blood mononuclear cells (PBMC) and plasma samples were isolated at consecutive time points. Patient plasma samples were added back to monocytes obtained pre-operatively for ex vivo LPS stimulations and TNF-α and IL-6 production was measured by flow cytometry. LPS-induced p38 mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB activation by patient plasma was assessed by Western blotting. A cell-permeable peptide inhibitor was used to block STAT3 signaling. We found that plasma samples obtained 4 h after surgery, regardless of pre-operative dexamethasone treatment, potently inhibited LPS-induced TNF-α but not IL-6 synthesis by monocytes. This was not associated with attenuation of p38 MAPK activation or IκB-α degradation. However, abrogation of the IL-10/STAT3 pathway restored LPS-induced TNF-α production in the presence of suppressive patient plasma. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that STAT3 signaling plays a crucial role in the downregulation of TNF-α synthesis by human monocytes in the course of systemic inflammation in vivo. Thus, STAT3 might be a potential molecular target for pharmacological intervention in clinical syndromes characterized by systemic inflammation

Post-perfusion plasma suppresses LPS-induced TNF-α production by monocytes.

<p><b>A</b>. Percentage of TNF-α producing cells in the monocyte population after <i>ex vivo</i> LPS stimulation (100 ng/mL) of patient PBMC isolated at various time points (n = 4). <b>B</b>. Reduced TNF-α synthesis by monocytes after LPS (10 ng/mL) stimulation in whole blood assays with patient samples obtained at the indicated time points (n = 5). <b>C</b>. Experimental setup for experiments shown in D,E,G-I. In short, patient PBMC obtained before surgery (Pre-op) were mixed with control (pooled AB plasma from healthy donors) or autologous patient plasma samples obtained at indicated time points, followed by LPS (100 ng/mL) stimulation for 4 h. Monocyte populations (CD14/SSC gate) were then analyzed for intracellular TNF-α and IL-6 synthesis. <b>D</b>. Significantly reduced production of TNF-α by monocytes after LPS stimulation in the presence of plasma samples from different sources (n = 13). Shown are percentages of TNF-α producing monocytes relative to control (100%). *<i>P</i><0.05, **<i>P</i><0.001 vs. control (ANOVA). <b>E</b>. Percentages of IL-6 producing monocytes as in D. **<i>P</i><0.001 vs. control (ANOVA). <b>F</b>. Dexamethasone levels in patient plasma samples as measured by radio-immunoassay (n = 9). Median ± interquartile range. *<i>P</i><0.05 vs. pre-op (ANOVA). <b>G</b>. Production of TNF-α and IL-6 by monocytes after LPS stimulation in the presence of dexamethasone-free plasma samples (n = 4). *<i>P</i><0.05 vs. control (ANOVA). <b>H</b>. Mean fluorescence intensities (MFI) of TNF-α and IL-6 in monocytes after LPS stimulation in different plasma milieus (n = 7). *<i>P</i><0.05, **<i>P</i><0.001 vs. control (ANOVA). <b>I</b>. Representative flow cytometry results (contour plots) of the LPS-induced TNF-α production by monocytes in the presence of control or patient plasma (Pre-op, End-CPB, 4 h or 24 h post-perfusion plasma from a No-dexamethasone patient). Isotype control: mouse IgG1. Data represented as mean ± SEM, unless otherwise indicated.</p

Patient characteristics.

<p>Age, CPB, ACC and PICU durations represented as median ± SD. ACC: aortic crossclamping, ASD: atrial septum defect, AVSD: atrioventricular septum defect, CoA: Coarctation aorta, CPB: cardiopulmonary bypass, Extracardiac conduit change due to stenosis after Fontan procedure, PICU: pediatric intensive care unit, VSD: ventricular septum defect. No significant differences were found between both patient groups (Mann-Whitney test).</p

Post-perfusion plasma does not interfere with p38 MAPK or NF-κB activation.

<p>Representative examples (<b>A</b>) and densitometric analyses (<b>B–C</b>) of LPS-induced p38 MAPK and IκB-α phosphorylation in monocytes in the presence of 24 h (control) or 4 h post-surgery plasma. Tubulin: loading control. Mean ± SEM (n = 4). *<i>P</i><0.05 vs. 0 min (ANOVA).</p

STAT3 signaling is required for the suppressive effects of post-perfusion plasma on TNF-α production.

<p><b>A</b>. Pre-treatment of 4 h post-surgery plasma samples with anti-IL-10 partially restored TNF-α production by patient monocytes in response to LPS (n = 10). Control: plasma from healthy donors. <b>B</b>. Activation of STAT3 in monocytes by incubation with suppressive (4 h post-perfusion) but not control (24 h post-perfusion) plasma. Cells were incubated in the absence or presence of LPS to match the experimental setup as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035070#pone-0035070-g002" target="_blank">Fig. 2</a>. <b>C</b>. Pre-treatment of patient PBMC with active STAT3 inhibitor (pY-STAT3i) but not control peptide (STAT3i) before LPS stimulation in the presence of post-surgery plasma restored TNF-α synthesis (left panel), in contrast to IL-6 (right panel). Shown are percentages of TNF-α and IL-6 producing monocytes normalized to control (24 h post-surgery) plasma (n = 8). <b>D</b>. TNF-α and IL-6 levels measured in supernatants of LPS-stimulated mononuclear cells after pre-treatment with STAT3 inhibitor or control peptide, in the presence of 4 h post-surgery plasma (n = 8). Cytokine levels were normalized to LPS stimulation in control plasma from healthy donors due to interassay variability. All results are depicted as mean ± SEM. *<i>P</i><0.05 vs. control condition (ANOVA), ns: not significant.</p

Inflammatory events induced by CPB surgery.

<p>Increased mean neutrophil (<b>A</b>) and monocyte (<b>B</b>) counts after on-pump cardiac surgery (n = 21 and n = 24, respectively). <b>C</b>. Increased numbers of circulating CD14+CD16+ monocytes after CPB surgery (n = 14). <b>D</b>. Increased mean C-reactive protein (CRP) levels in patient blood samples post-surgery (n = 22). <b>E</b>. Lymphopenia was observed 4 h post-surgery (n = 27). Box-and-whiskers plots. *<i>P</i><0.01, **<i>P</i><0.001 vs. pre-op (ANOVA). <b>F</b>. Cyto- and chemokine color profiles of plasma samples (n = 12) obtained at indicated time points, represented as % change compared to baseline. MIF: Macrophage migration inhibitory factor.</p

Long-term restoration of the human T-cell compartment after thymectomy during infancy : a role for thymic regeneration?

Thymectomy during early childhood is generally thought to have serious consequences for the establishment of the T-cell compartment. In the present study, we investigated the composition of the T-cell pool in the first 3 decades after thymectomy during infancy due to cardiac surgery. In the first 5 years after thymectomy, naive and total CD4(+) and CD8(+) T-cell numbers in the blood and T-cell receptor excision circle (TREC) levels in CD4(+) T cells were significantly lower than in healthy age-matched controls. In the first years after thymectomy, plasma IL-7 levels were significantly elevated and peripheral T-cell proliferation levels were increased by ∼ 2-fold. From 5 years after thymectomy onward, naive CD4(+) and CD8(+) T-cell counts and TRECs were within the normal range. Because TREC levels are expected to decline continuously in the absence of thymic output, we investigated whether normalization of the naive T-cell pool could be due to regeneration of thymic tissue. In the majority of individuals who had been thymectomized during infancy, thymic tissue could indeed be identified on magnetic resonance imaging scans. Whereas thymectomy has severe effects on the establishment of the naive T-cell compartment during early childhood, our data suggest that functional regrowth of thymic tissue can limit its effects in subsequent years