Helen Betts

Genome-wide association study identifies variants in the CFH region associated with host susceptibility to meningococcal disease

Meningococcal disease is an infection caused by Neisseria meningitidis. Genetic factors contribute to host susceptibility and progression to disease, but the genes responsible for disease development are largely unknown. We report here a genome-wide association study for host susceptibility to meningococcal disease using 475 individuals with meningococcal disease (cases) and 4,703 population controls from the UK. We performed, in Western European and South European cohorts (consisting of 968 cases and 1,376 controls), two replication studies for the most significant SNPs. A cluster of complement factor SNPs replicated independently in both cohorts, including SNPs within complement factor H (CFH) (rs1065489 (p.936D<E), P = 2.2 × 10−11) and in CFH-related protein 3 (CFHR3)(rs426736, P = 4.6 × 10−13). N. meningitidis is known to evade complement-mediated killing by the binding of host CFH to the meningococcal factor H–binding protein (fHbp). Our study suggests that host genetic variation in these regulators of complement activation plays a role in determining the occurrence of invasive disease versus asymptomatic colonization by this pathogen.

A functional microsatellite of the macrophage migration inhibitory factor gene associated with meningococcal disease

Macrophage migration inhibitory factor (MIF) is an abundantly expressed proinflammatory cytokine playing a critical role in innate immunity and sepsis and other inflammatory diseases. We examined whether functional MIF gene polymorphisms (–794 CATT5–8 microsatellite and –173 G/C SNP) were associated with the occurrence and outcome of meningococcal disease in children. The CATT5 allele was associated with the probability of death predicted by the Pediatric Index of Mortality 2 (P=0.001), which increased in correlation with the CATT5 copy number (P=0.04). The CATT5 allele, but not the —173 G/C alleles, was also associated with the actual mortality from meningoccal sepsis [OR 2.72 (1.2‐6.4), P=0.02]. A family‐based association test (i.e., transmission disequilibrium test) performed in 240 trios with 1 afflicted offspring indicated that CATT5 was a protective allele (P=0.02) for the occurrence of meningococcal disease. At baseline and after stimulation with Neisseria meningitidis in THP‐1 monocytic cells or in a whole‐blood assay, CATT5 was found to be a low‐expression MIF allele (P= 0.005 and P=0.04 for transcriptional activity; P=0.09 and P=0.09 for MIF production). Taken together, these data suggest that polymorphisms of the MIF gene affecting MIF expression are associated with the occurrence, severity, and outcome of meningococcal disease in children.—Renner, P., Roger, T., Bochud, P.‐Y., Sprong, T., Sweep, F. C. G. J., Bochud, M., Faust, S. N., Haralambous, E., Betts, H., Chanson, A.‐L., Reymond, M. K., Mermel, E., Erard, V., van Deuren, M., Read, R. C., Levin, M., Calandra, T. A functional microsatellite of the macrophage migration inhibitory factor gene associated with meningococcal disease. FASEB J. 26, 907–916 (2012). www.fasebj.org

Intestinal Injury and Endotoxemia in Children Undergoing Surgery for Congenital Heart Disease

RATIONALE Children with congenital heart disease are at risk of gut barrier dysfunction and translocation of gut bacterial antigens into the bloodstream. This may contribute to inflammatory activation and organ dysfunction postoperatively. OBJECTIVES To investigate the role of intestinal injury and endotoxemia in the pathogenesis of organ dysfunction after surgery for congenital heart disease. METHODS We analyzed blood levels of intestinal fatty acid binding protein and endotoxin (endotoxin activity assay) alongside global transcriptomic profiling and assays of monocyte endotoxin receptor expression in children undergoing surgery for congenital heart disease. MEASUREMENTS AND MAIN RESULTS Levels of intestinal fatty acid binding protein and endotoxin were greater in children with duct-dependent cardiac lesions. Endotoxemia was associated with severity of vital organ dysfunction and intensive care stay. We identified activation of pathogen-sensing, antigen-processing, and immune-suppressing pathways at the genomic level postoperatively and down-regulation of pathogen-sensing receptors on circulating immune cells. CONCLUSIONS Children undergoing surgery for congenital heart disease are at increased risk of intestinal mucosal injury and endotoxemia. Endotoxin activity correlates with a number of outcome variables in this population, and may be used to guide the use of gut-protective strategies.

Hyperglycemia is associated with morbidity in critically ill children with meningococcal sepsis

Objective: To determine the association between hyperglycemia and outcome in children ventilated for meningococcal sepsis. Design: Retrospective case notes review. Setting: Eight bedded pediatric intensive care unit in London. Patients: Consecutive children ventilated for meningococcal sepsis 2001–2004. Interventions: None. Measurements: Peak glucose for the entire admission was determined and mean glucose was calculated for the following three epochs: 1) first 24 hrs, 2) second 24 hrs, and 3) the entire pediatric intensive care unit admission. Patients were also grouped according to whether their blood glucose rose to >7 mmol/L (126 mg/dL), >10 mmol/L (180 mg/dL), or remained below these levels during the pediatric intensive care unit admission. Outcome measures were predicted mortality (based on pediatric risk of mortality score), ventilator free days at 30 days, nosocomial infection, use of renal replacement therapy, use of inotropes, and skin necrosis. Main Results: Ninety-seven patients were identified with a median age of 2.1 yrs and a median length of stay of 4 days. Four patients died. Peak glucose significantly correlated with predicted mortality and negatively correlated with ventilator free days at 30 days (p < 0.001 and p < 0.001, respectively). Patients who received renal replacement therapy or inotropic support, or developed a nosocomial infection or skin necrosis had significantly higher peak glucose than those who did not (p = 0.006, p < 0.0001, p = 0.022, and p < 0.0001, respectively). Patients who received renal replacement therapy or who developed skin necrosis had significantly higher mean blood glucose in the second 24 hrs of admission (p = 0.017 and p = 0.004, respectively). However, mean blood glucose in the first 24 hrs and over the entire admission did not correlate with outcome. Patients defined as hyperglycemic with blood glucose either >7 mmol/L or >10 mmol/L also had a significantly worse outcome than those who maintained blood glucose below these levels. Conclusions: There was a significant association between hyperglycemia and outcome. Our results support a trial of tight glycemic control in this group of critically ill children.

Control of hyperglycaemia in paediatric intensive care (CHiP): study protocol

BackgroundThere is increasing evidence that tight blood glucose (BG) control improves outcomes in critically ill adults. Children show similar hyperglycaemic responses to surgery or critical illness. However it is not known whether tight control will benefit children given maturational differences and different disease spectrum.Methods/DesignThe study is an randomised open trial with two parallel groups to assess whether, for children undergoing intensive care in the UK aged ≤ 16 years who are ventilated, have an arterial line in-situ and are receiving vasoactive support following injury, major surgery or in association with critical illness in whom it is anticipated such treatment will be required to continue for at least 12 hours, tight control will increase the numbers of days alive and free of mechanical ventilation at 30 days, and lead to improvement in a range of complications associated with intensive care treatment and be cost effective.Children in the tight control group will receive insulin by intravenous infusion titrated to maintain BG between 4 and 7.0 mmol/l. Children in the control group will be treated according to a standard current approach to BG management.Children will be followed up to determine vital status and healthcare resources usage between discharge and 12 months post-randomisation. Information regarding overall health status, global neurological outcome, attention and behavioural status will be sought from a subgroup with traumatic brain injury (TBI).A difference of 2 days in the number of ventilator-free days within the first 30 days post-randomisation is considered clinically important. Conservatively assuming a standard deviation of a week across both trial arms, a type I error of 1% (2-sided test), and allowing for non-compliance, a total sample size of 1000 patients would have 90% power to detect this difference. To detect effect differences between cardiac and non-cardiac patients, a target sample size of 1500 is required. An economic evaluation will assess whether the costs of achieving tight BG control are justified by subsequent reductions in hospitalisation costs.DiscussionThe relevance of tight glycaemic control in this population needs to be assessed formally before being accepted into standard practice.Trial RegistrationCurrent Controlled Trials ISRCTN61735247

Haplotypic diversity in human CEACAM genes: effects on susceptibility to meningococcal disease

Adhesion between the opacity-associated adhesin (Opa) proteins of Neisseria meningitidis and human carcino-embryonic antigen cell adhesion molecule (CEACAM) proteins is an important stage in the pathogenesis of meningococcal disease, a globally important bacterial infection. Most disease is caused by a small number of meningococcal genotypes known as hyperinvasive lineages. As these are also carried asymptomatically, acquisition of them alone cannot explain why only some hosts develop meningococcal disease. Our aim was to determine whether genetic diversity in CEACAM is associated with susceptibility to meningococcal disease. Frequency distributions of alleles, genotypes and haplotypes were compared in four CEACAM genes in 384 case samples and 190 controls. Linkage disequilibrium among polymorphic sites, haplotype structures and relationships were also analysed. A number of polymorphisms were observed in CEACAM genes but the diversity of CEACAM1, to which most Opa proteins bind, was lower, and a small number of high-frequency haplotypes were detected. Dose-dependent associations of three CEACAM haplotypes with meningococcal disease were observed, with the effect of carrying these haplotypes amplified in homozygous individuals. Two haplotypes were protective while one haplotype in CEACAM6 was associated with a twofold increase in disease susceptibility. These data imply that human CEACAM may be one determinant of human susceptibility to meningococcal disease.

A clinical and economic evaluation of Control of Hyperglycaemia in Paediatric intensive care (CHiP): a randomised controlled trial.

BACKGROUND Early research in adults admitted to intensive care suggested that tight control of blood glucose during acute illness can be associated with reductions in mortality, length of hospital stay and complications such as infection and renal failure. Prior to our study, it was unclear whether or not children could also benefit from tight control of blood glucose during critical illness. OBJECTIVES This study aimed to determine if controlling blood glucose using insulin in paediatric intensive care units (PICUs) reduces mortality and morbidity and is cost-effective, whether or not admission follows cardiac surgery. DESIGN Randomised open two-arm parallel group superiority design with central randomisation with minimisation. Analysis was on an intention-to-treat basis. Following random allocation, care givers and outcome assessors were no longer blind to allocation. SETTING The setting was 13 English PICUs. PARTICIPANTS Patients who met the following criteria were eligible for inclusion: ≥ 36 weeks corrected gestational age; ≤ 16 years; in the PICU following injury, following major surgery or with critical illness; anticipated treatment > 12 hours; arterial line; mechanical ventilation; and vasoactive drugs. Exclusion criteria were as follows: diabetes mellitus; inborn error of metabolism; treatment withdrawal considered; in the PICU > 5 consecutive days; and already in CHiP (Control of Hyperglycaemia in Paediatric intensive care). INTERVENTION The intervention was tight glycaemic control (TGC): insulin by intravenous infusion titrated to maintain blood glucose between 4.0 and 7.0 mmol/l. CONVENTIONAL MANAGEMENT (CM) This consisted of insulin by intravenous infusion only if blood glucose exceeded 12.0 mmol/l on two samples at least 30 minutes apart; insulin was stopped when blood glucose fell below 10.0 mmol/l. MAIN OUTCOME MEASURES The primary outcome was the number of days alive and free from mechanical ventilation within 30 days of trial entry (VFD-30). The secondary outcomes comprised clinical and economic outcomes at 30 days and 12 months and lifetime cost-effectiveness, which included costs per quality-adjusted life-year. RESULTS CHiP recruited from May 2008 to September 2011. In total, 19,924 children were screened and 1369 eligible patients were randomised (TGC, 694; CM, 675), 60% of whom were in the cardiac surgery stratum. The randomised groups were comparable at trial entry. More children in the TGC than in the CM arm received insulin (66% vs. 16%). The mean VFD-30 was 23 [mean difference 0.36; 95% confidence interval (CI) -0.42 to 1.14]. The effect did not differ among prespecified subgroups. Hypoglycaemia occurred significantly more often in the TGC than in the CM arm (moderate, 12.5% vs. 3.1%; severe, 7.3% vs. 1.5%). Mean 30-day costs were similar between arms, but mean 12-month costs were lower in the TGC than in CM arm (incremental costs -£3620, 95% CI -£7743 to £502). For the non-cardiac surgery stratum, mean costs were lower in the TGC than in the CM arm (incremental cost -£9865, 95% CI -£18,558 to -£1172), but, in the cardiac surgery stratum, the costs were similar between the arms (incremental cost £133, 95% CI -£3568 to £3833). Lifetime incremental net benefits were positive overall (£3346, 95% CI -£11,203 to £17,894), but close to zero for the cardiac surgery stratum (-£919, 95% CI -£16,661 to £14,823). For the non-cardiac surgery stratum, the incremental net benefits were high (£11,322, 95% CI -£15,791 to £38,615). The probability that TGC is cost-effective is relatively high for the non-cardiac surgery stratum, but, for the cardiac surgery subgroup, the probability that TGC is cost-effective is around 0.5. Sensitivity analyses showed that the results were robust to a range of alternative assumptions. CONCLUSIONS CHiP found no differences in the clinical or cost-effectiveness of TGC compared with CM overall, or for prespecified subgroups. A higher proportion of the TGC arm had hypoglycaemia. This study did not provide any evidence to suggest that PICUs should stop providing CM for children admitted to PICUs following cardiac surgery. For the subgroup not admitted for cardiac surgery, TGC reduced average costs at 12 months and is likely to be cost-effective. Further research is required to refine the TGC protocol to minimise the risk of hypoglycaemic episodes and assess the long-term health benefits of TGC. TRIAL REGISTRATION Current Controlled Trials ISRCTN61735247. FUNDING This project was funded by the NIHR Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 18, No. 26. See the NIHR Journals Library website for further project information.

Polymorphisms in PARP, IL1B, IL4, IL10, C1INH, DEFB1, and DEFA4 in meningococcal disease in three populations.

The pathogenesis of meningococcal infections involves activation of the complement system, proinflammatory and anti-inflammatory mediators, antimicrobial peptides, and apoptosis. We hypothesized that variations in genes encoding these products are involved in the susceptibility to and severity of pediatric meningococcal infections. Polymorphisms in poly (adenosine diphosphate-ribose) polymerase (PARP), serine protease C1 inhibitor (C1INH), IL4, IL10 and IL1B, &agr;-defensin 4, and &bgr;-defensin 1 (DEFB1) were analyzed in two independent Caucasian case control cohorts from the United Kingdom and the Netherlands and in a family-based transmission disequilibrium test cohort from the UK. In the UK case control cohort, the DEFB1 -44 G/G homozygous genotype was overrepresented in patients with meningococcal disease compared with the G/C and C/C genotypes when combined (odds ratio, 1.57; 95% confidence interval, 1.12-2.20). The transmission disequilibrium test analysis did not confirm this, but did find an association and linkage of the IL4 -524 and the C1INH 480 polymorphisms with susceptibility to meningococcal infection. Hematological failure was present more often in UK patients with the DEFB1 -44 G/G genotype compared with the C allele carriers (odds ratio, 2.17; 95% confidence interval, 1.22-3.85). Additional studies are necessary to elucidate the conflicting results obtained for the DEFB1, IL4, and C1INH polymorphisms and their role in susceptibility to and severity of meningococcal disease.ABBREVIATIONS-C1INH-serine protease C1 inhibitor; DEFA4-&agr; defensin 4; DEFB1-&bgr;-defensin 1; EMC-Erasmus MC Sophia; IC-Imperial College; MRF-Meningitis Research Foundation; PARP-poly (ADP-ribose) polymerase; PICU-pediatric intensive care unit; SNP-single nucleotide polymorphism; TDT-transmission disequilibrium test