Irradiance levels of phototherapy devices: a national study in Dutch neonatal intensive care units.

OBJECTIVE: The objective of this study is to determine whether irradiance levels of phototherapy (PT) devices in Dutch neonatal intensive care units (NICUs) increased between 2008 and 2013. STUDY DESIGN: Irradiance of all types of PT devices, used in combination with incubators, was measured with a Dale 40 Radiometer (Fluke Biomedical, Everett, WA, USA) in all 10 Dutch NICUs. RESULTS: Irradiance increased in seven NICUs. Median (range) irradiance increased from 9.7 (4.3-32.6) to 16.4 (6.8-41) µW cm-2 nm-1 for 24 overhead devices (P=0.004) and from 6.8 (0.8-15.6) to 22.3 (1.1-36.3) µW cm-2 nm-1 for 12 underneath devices (P=0.014). Five light-emitting diode (LED)-based devices were used in 2013 and one in 2008. The mean distance between overhead PT device and infant decreased by ~9 cm (P<0.001). Significantly more devices delivered minimal (10 µW cm-2 nm-1) recommended irradiance levels (80 vs ~45%; P=0.002). CONCLUSION: Irradiance of PT devices still varies, but has markedly improved since 2008 due to shorter distances between PT device and infant, and introduction of better performing LED-based devices.

Bilirubin production and hour-specific bilirubin levels.

OBJECTIVE: We assessed the relative contributions of increased bilirubin production (indexed by end-tidal carbon monoxide (CO) concentrations, corrected for ambient CO (ETCOc)) to hour-specific total bilirubin (TB) levels in healthy late preterm and term newborns. STUDY DESIGN: Post hoc analyses of concurrent ETCOc and TB (at 30±6 h of age) and follow-up TB levels at age 96±12 h and up to 168 h after birth were performed in a cohort of 641 term and late preterm infants. RESULTS: Increased bilirubin production (hour-specific ETCOc ⩾1.7 p.p.m. at age 30±6 h) was noted in ~80%, 42% and 32% of infants in the high-, intermediate- and low-risk TB zones, respectively. One infant with TB <40th percentile and ETCOc <1.7 p.p.m. developed TB ⩾95th percentile at age 168 h, probably due to decreased bilirubin elimination. CONCLUSIONS: Infants in the high-risk quartile of the hour-specific bilirubin nomogram have a higher mean bilirubin production. Infants with TB levels ⩾95th percentile without increased bilirubin production have impaired bilirubin elimination.

Protective effects of inhaled carbon monoxide in endotoxin-induced cholestasis is dependent on its kinetics.

Carbon monoxide (CO), a product of heme oxygenase (HMOX), has many beneficial biological functions and is a promising therapeutic agent for many pathological conditions. However, the kinetics of inhaled CO and its protective role in endotoxin-induced cholestasis is not fully known. Thus, our objective was to characterize the kinetics of inhaled CO and then investigate its use in early phase experimental endotoxin-induced cholestasis. Female Wistar rats were randomly divided into 4 groups: CON (control), LPS (lipopolysaccharide, 6 mg/kg), CO (250 ppm COx1h), and CO + LPS. Rats were sacrificed at 0-12 h after LPS administration. Tissues and blood were collected for liver injury markers and tissue CO distribution measurements. Livers were harvested for measurements of Hmox activity, Hmox1 mRNA expression, cytokines (IL10, IL6, TNF), and bile lipid and pigment transporters. Half-lives of CO in spleen, blood, heart, brain, kidney, liver, and lungs were 2.4 ± 1.5, 2.3 ± 0.8, 1.8 ± 1.6, 1.5 ± 1.2, 1.1 ± 1.1, 0.6 ± 0.3, 0.6 ± 0.2 h, respectively. CO treatment increased liver IL10 mRNA and decreased TNF expression 1 h after LPS treatment and prevented the down-regulation of bile acid and bilirubin hepatic transporters (Slc10a1, Abcb11, and Abcc2, p < 0.05), an effect closely related to the kinetics. The protective effect of CO against cholestatic liver injury persisted even 12 h after CO exposure, as shown by attenuation of serum cholestatic markers in CO-treated animals. CO exposure substantially attenuated endotoxin-induced cholestatic liver injury and was directly related to the kinetics of inhaled CO. This data underscores the importance of the kinetics of inhaled CO for the proper design of experimental and clinical studies of using CO as a treatment strategy.

In vitro inhibition of heme oxygenase isoenzymes by metalloporphyrins.

OBJECTIVE: Neonatal jaundice results from an increased bilirubin production and decreased hepatic bilirubin conjugation and excretion. Severe hyperbilirubinemia is currently treated with phototherapy or exchange transfusion; however, its prevention by inhibiting bilirubin formation is a more logical strategy. Heme oxygenase (HO), with inducible (HO-1) and constitutive (HO-2) isoenzymes, is the rate-limiting enzyme in heme catabolism, producing equimolar amounts of bilirubin and carbon monoxide (CO). Metalloporphyrins (Mps) are heme derivatives that competitively inhibit HO and thereby suppress hyperbilirubinemia. No systematic studies have been reported evaluating whether the HO isoenzymes are inhibited differentially by various Mps. Identification of Mps that selectively inhibit the inducible HO-1 without affecting the 'housekeeping' HO-2 isoenzyme might be desirable in the clinical setting of hemolytic disease, in which the Hmox1 gene is greatly induced. Although bilirubin production is due to the activity of both HO-1 and HO-2, the inhibition of HO-1 with a relative sparing of HO-2 activity might provide the most selective approach for the treatment of hemolytic disease. STUDY DESIGN: We determined for the deutero-, proto-, meso- and bis-glycol porphyrins with zinc, tin and chromium as central atoms, respectively, the concentration needed for 50% inhibition (I(50)) of HO-1 and HO-2 activities in rat spleen and brain tissue. RESULT: For a given Mp, HO-1 activity was less inhibited than that of HO-2. The order of inhibitor potency of each Mp was nearly identical for both isoenzymes. Tin mesoporphyrin was the most potent inhibitor for both isoenzymes. HO-2 selectivity was greatest for tin protoporphyrin. Conversely, the Zn compounds were least inhibitory toward HO-2. No Mp preferentially inhibited HO-1. CONCLUSION: Mps that produce a less inhibitory effect on HO-2, while limiting the response of the inducible HO-1, such as ZnPP, may be a useful clinical tool.

Influence of clinical status on the association between plasma total and unbound bilirubin and death or adverse neurodevelopmental outcomes in extremely low birth weight infants.

OBJECTIVES: To assess the influence of clinical status on the association between total plasma bilirubin and unbound bilirubin on death or adverse neurodevelopmental outcomes at 18-22 months corrected age in extremely low birth weight infants. METHOD: Total plasma bilirubin and unbound bilirubin were measured in 1101 extremely low birth weight infants at 5 +/- 1 days of age. Clinical criteria were used to classify infants as clinically stable or unstable. Survivors were examined at 18-22 months corrected age by certified examiners. Outcome variables were death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death prior to follow-up. For all outcomes, the interaction between bilirubin variables and clinical status was assessed in logistic regression analyses adjusted for multiple risk factors. RESULTS: Regardless of clinical status, an increasing level of unbound bilirubin was associated with higher rates of death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss and death before follow-up. Total plasma bilirubin values were directly associated with death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death before follow-up in unstable infants, but not in stable infants. An inverse association between total plasma bilirubin and death or cerebral palsy was found in stable infants. CONCLUSIONS: In extremely low birth weight infants, clinical status at 5 days of age affects the association between total plasma bilirubin and death or adverse neurodevelopmental outcomes at 18-22 months of corrected age. An increasing level of UB is associated a higher risk of death or adverse neurodevelopmental outcomes regardless of clinical status. Increasing levels of total plasma bilirubin are directly associated with increasing risk of death or adverse neurodevelopmental outcomes in unstable, but not in stable infants.

Severe hemolysis with normal blood count in a glucose-6-phosphate dehydrogenase deficient neonate.

A premature glucose-6-phosphate dehydrogenase (G-6-PD) deficient neonate was readmitted for exponential rise in the plasma bilirubin concentration to 33.0 mg dl(-1). Blood carboxyhemoglobin (2.8% of total hemoglobin, >threefold normal value) confirmed the presence of hemolysis; however, hematological indices were unchanged from the birth hospitalization. Serum unbound bilirubin, although present, was probably at a concentration insufficient to cause bilirubin encephalopathy. In G-6-PD deficient neonates, severe hemolysis may occur in the absence of hematological changes typical of a hemolytic process.

Standardized bench method for evaluating the efficacy of phototherapy devices.

AIM: As phototherapy (PT) devices employ a variety of broadband light sources, we developed and tested a standardized bench method for evaluating the efficacy of some devices. METHODS: To evaluate efficacy, we quantified the in vitro photodegradation rate (expressed as t1/2) of unconjugated bilirubin in solution at 37 degrees C during exposure to a given light source at its mean delivered irradiance to the 2D body surface area (BSA) of newborn models. Reproducibility (between-day variation) of the method was determined at irradiance levels from 10 to 70 microW/cm2/nm on three different days. RESULTS: Between-day t1/2 measurements had coefficients of variation from 3% to 10%. When t1/2 values were normalized to the exposable 2D horizontal BSA, halogen lamp devices, without and with fiberoptics, were least effective (t1/2=60-108 min and 100-126 min for preterm and term models, respectively). Fluorescent tube devices had t1/2=19-78 min and 25-78 min, for preterm and term models, respectively. Light-emitting diode (LED)-based devices yielded the shortest t1/2 values (16-24 min) for preterm and term newborn models. CONCLUSIONS: We demonstrated the applicability of the method through the determination of the efficacy of several commercially available PT devices. This standardized method is reproducible and effectively evaluates the relative in vitro efficacy of various devices and may guide further in vitro and in vivo evaluations of devices.

Neonatal bilirubin production-conjugation imbalance: effect of glucose-6-phosphate dehydrogenase deficiency and borderline prematurity.

OBJECTIVE: To evaluate relations between production and conjugation of bilirubin in the pathophysiology of jaundice in glucose-6-phosophate dehydrogenase (G6PD) deficient neonates. METHODS: Term and borderline premature (35-37 weeks gestational age), healthy, male, G6PD deficient neonates were studied close to the beginning of the 3rd day. Blood carboxyhaemogobin corrected for inspired CO (COHbc; an index of bilirubin production) and serum total conjugated bilirubin (TCB; a reflection of bilirubin conjugation) were measured in simultaneously drawn blood samples by gas chromatography and reverse phase high performance liquid chromatography respectively. A bilirubin production-conjugation index comprising COHbc/TCB was determined; a high index reflects imbalance between the bilirubin production and conjugation processes. COHbc and TCB individually and the production-conjugation index were studied in relation to serum total bilirubin (STB) concentration. RESULTS: Fifty one G6PD deficient neonates were sampled at 51 (8) hours. COHbc values did not correlate with STB (r=0.22, p=0.15). TCB did correlate inversely with STB (r=-0.42, p=0.004), and there was a positive correlation between the production-conjugation index and STB (r=0.45, p=0.002). The production-conjugation index (median (interquartile range)) was higher in the premature (n=8) than term neonates (2.31 (2.12-3.08) v 1.05 (0.53-1.81), p=0.003). This difference was the result of changes in TCB. CONCLUSIONS: The data show that jaundice in G6PD deficient neonates is the result of an imbalance between production and conjugation of bilirubin with a tendency for inefficient bilirubin conjugation over increased haemolysis in its pathogenesis. Borderline premature infants are at special risk of bilirubin production-conjugation imbalance.

The jaundiced newborn. Understanding and managing transitional hyperbilirubinemia.

Neonatal jaundice is one of the most common conditions diagnosed by the pediatrician. This normally benign transitional phenomenon is a dynamic balance between the production and elimination of bilirubin. These processes can be exacerbated by a number of pathophysiologic conditions, which cause either an increase in bilirubin production rates, such as hemolysis, or a decrease in bilirubin elimination rates, such as bilirubin conjugation defects. The most dangerous circumstance for an infant is the combination of increased bilirubin production with impaired elimination. These infants are at considerable risk for developing excessive and potentially dangerous hyperbilirubinemia and subsequent kernicterus. Therefore, the importance of early recognition of the imbalance is paramount. In this review, we will discuss the various risk factors associated with hyperbilirubinemia and describe strategies for the diagnosis and management of transitional hyperbilirubinemia.

Endogenous carbon monoxide formation by chorionic villi of term human placenta.

Carbon monoxide (CO) is a novel messenger that is proposed to play a complementary role with nitric oxide in the regulation of placental haemodynamics. In a previous study, CO formation from exogenous haem has been measured in the microsomal fraction of chorionic villi as an index of haem oxygenase activity. The objective of the present study was to determine whether endogenous CO is formed by dissected chorionic villi of term human placenta, to which no exogenous substrate or co-factor had been added. Each sample of freshly isolated chorionic villi (approximately 0.4 g) of term human placenta from caesarean delivery was incubated in a sealed vial containing 1 ml of Krebs' solution (pH 7.4) at 37 degrees C. CO formation was determined by quantitating, using a gas-chromatographic method, the amount of CO released into the headspace gas of the incubation vial. There was time-dependent formation of endogenous CO in chorionic villi incubated at 37 degrees C during a 60-min time course. CO formation was found to be minimal in chorionic villi samples incubated at 4 degrees C and was increased relative to tissue weight. The data demonstrate that there is endogenous CO formation by chorionic villi of term human placenta.

Differing pathogenesis of perinatal bilirubinemia in glucose-6-phosphate dehydrogenase-deficient versus-normal neonates.

The objective was to compare the contribution to perinatal bilirubinemia of hemolysis and UDP-glucuronosyltransferase (UGT) gene promoter polymorphism, seen in Gilbert's syndrome, between glucose-6-phosphate dehydrogenase (G-6-PD)-deficient and -normal neonates. Serum total bilirubin (STB) values from 52 G-6-PD-deficient and 166 G-6-PD-normal term, male neonates, sampled within 3 h of delivery (first sample) and on d 3 (second sample), were analyzed in relation to blood carboxyhemoglobin corrected for inspired CO (COHbc), an accurate index of hemolysis, and UGT promoter genotype. COHbc values (% total Hb) were greater in G-6-PD-deficient neonates than controls: first sample 1.00 +/- 0.25% versus 0.84 +/- 0.24%, p < 0.0001; second sample 0.83 +/- 0.20% versus 0.76 +/- 0.19%, p = 0.002. First sample COHbc and STB values did not correlate in either the G-6-PD-deficient or control groups, whereas second sample COHbc values correlated significantly with corresponding STB values in the control population only (r = 0.28, p = 0.0007). At second sampling, there was a higher allele frequency of the variant UGT promoter among those with STB values > or =75th percentile than those <75th among the G-6-PD-deficient neonates (0.60 versus 0.33, respectively, p = 0.025), but not controls (0.31 versus 0.40, respectively, p = 0.24). Among those infants with at least one variant UGT promoter allele, STB values were higher in the G-6-PD-deficient neonates than controls at second sampling only (181 +/- 56 microM versus 149 +/- 46 microM, respectively, p = 0.03). Both within and between the G-6-PD-deficient and control groups, our data demonstrate changing and differing contributions of hemolysis and UGT promoter polymorphism to bilirubinemia during the first 3 d of life.

Acute hemolysis and severe neonatal hyperbilirubinemia in glucose-6-phosphate dehydrogenase-deficient heterozygotes.

Two premature female infants had severe hyperbilirubinemia caused by hemolysis. Both neonates were heterozygotes for the glucose-6-phosphate dehydrogenase Mediterranean mutation as determined by DNA analysis. Glucose-6-phosphate dehydrogenase-deficient heterozygotes may be susceptible to the complications of this enzyme deficiency.

Prediction of hyperbilirubinemia in near-term and term infants.

OBJECTIVE: The purpose of this study was to determine whether end-tidal carbon monoxide (CO) corrected for ambient CO (ETCOc), as a single measurement or in combination with serum total bilirubin (STB) measurements, can predict the development of hyperbilirubinemia during the first 7 days of life. METHODS: From 9 multinational clinical sites, 1370 neonates completed this cohort study from February 20, 1998, through February 22, 1999. Measurements of both ETCOc and STB were performed at 30 +/- 6 hours of life; STB also was measured at 96 +/- 12 hours and subsequently following a flow diagram based on a table of hours of age-specific STB. An infant was defined as hyperbilirubinemic if the hours of age-specific STB was greater than or equal to the 95th percentile as defined by the table at any time during the study. RESULTS: A total of 120 (8.8%) of the enrolled infants became hyperbilirubinemic. Mean STB in breastfed infants was 8.92 +/- 4.37 mg/dL at 96 hours versus 7.63 +/- 3.58 mg/dL in those fed formula only. The mean ETCOc at 30 +/- 6 hours for the total population was 1.48 +/- 0.49 ppm, whereas those of nonhyperbilirubinemic and hyperbilirubinemic infants were 1.45 +/- 0.47 ppm and 1.81 +/- 0.59 ppm, respectively. Seventy-six percent (92 of 120) of hyperbilirubinemic infants had ETCOc greater than the population mean. An ETCOc greater than the population mean at 30 +/- 6 hours yielded a 13.0% positive predictive value (PPV) and a 95.8% negative predictive value (NPV) for STB >/=95th percentile. When infants with STB >95th percentile at <36 hours of age were excluded, the STB at 30 +/- 6 hours yielded a 16.7% PPV and a 98.1% NPV for STB >75th percentile. The combination of these 2 measurements at 30 +/- 6 hours (either ETCOc more than the population mean or STB >75th percentile) had a 6.4% PPV with a 99.0% NPV. Conclusions. This prospective cohort study supports previous observations that measuring STB before discharge may provide some assistance in predicting an infant's risk for developing hyperbilirubinemia. The addition of an ETCOc measurement provides insight into the processes that contribute to the condition but does not materially improve the predictive ability of an hours of age-specific STB in this study population. The combination of STB and ETCOc as early as 30 +/- 6 hours may identify infants with increased bilirubin production (eg, hemolysis) or decreased elimination (conjugation defects) as well as infants who require early follow-up after discharge for jaundice or other clinical problems such as late anemia. Depending on the incidence of hyperbilirubinemia within an institution, the criteria for decision making should vary according to its unique population.

Carbon monoxide and bilirubin production in neonates.

Neonatal hyperbilirubinemia is a normal postnatal phenomenon resulting from a transitional imbalance between the production and elimination of bilirubin in the neonate. Bilirubin has been shown to be not only a potent antioxidant, but also toxic at excessive concentrations. As a result, the biology of bilirubin, its production, regulation, and measurements have been the focus of extensive studies. Bilirubin, carbon monoxide, and iron are derived from the degradation of heme, a ubiquitous two-step pathway catalyzed by the enzyme, heme oxygenase. It has been shown that these metabolically active products from the heme catabolic pathway may, in turn, influence many other biologic processes. This report provides a brief overview of these interrelationships in the hope that it may provide insight into the central role this pathway plays in the existence of most organisms.

Prediction of hyperbilirubinemia in near-term and term infants.

OBJECTIVE: The purpose of this study was to determine whether end-tidal carbon monoxide (CO) corrected for ambient CO (ETCOc), as a single measurement or in combination with serum total bilirubin (STB) measurements, can predict the development of hyperbilirubinemia during the first 7 days of life. METHODS: From nine multinational clinical sites, 1370 neonates completed this cohort study from February 20, 1998 through February 22, 1999. Measurements of both ETCOc and STB were performed at 30+/-6 hours of life; STB also was measured at 96+/-12 hours and subsequently following a flow diagram based on a table of hours of age-specific STB. An infant was defined as hyperbilirubinemic if the hours of age-specific STB was greater than or equal to the 95th percentile as defined by the table at any time during the study. RESULTS: A total of 120 (8.8%) of the enrolled infants became hyperbilirubinemic. Mean STB in breast-fed infants was 8.92+/-4.37 mg/dl at 96 hours versus 7.63+/-3.58 mg/dl in those fed formula only. The mean ETCOc at 30+/-6 hours for the total population was 1.48+/-0.49 ppm, whereas those of nonhyperbilirubinemic and hyperbilirubinemic infants were 1.45+/-0.47 and 1.81+/-0.59 ppm, respectively. Seventy-six percent (92 of 120) of hyperbilirubinemic infants had ETCOc greater than the population mean. An ETCOc greater than the population mean at 30+/-6 hours yielded a 13.0% positive predictive value (PPV) and a 95.8% negative predictive value (NPV) for STB > or =95th percentile. When infants with STB > or =95th percentile at <36 hours of age were excluded, the STB at 30+/-6 hours yielded a 16.7% PPV and a 98.1% NPV for STB >75th percentile. The combination of these two measurements at 30+/-6 hours (either ETCOc more than the population mean or STB >75th percentile) had a 6.4% PPV with a 99.0% NPV. CONCLUSIONS: This prospective cohort study supports previous observations that measuring STB before discharge may provide some assistance in predicting an infant's risk for developing hyperbilirubinemia. The addition of an ETCOc measurement provides insight into the processes that contribute to the condition but does not materially improve the predictive ability of an hours of age-specific STB in this study population. The combination of STB and ETCOc as early as 30+/-6 hours may identify infants with increased bilirubin production (eg, hemolysis) or decreased elimination (conjugation defects) as well as infants who require early follow-up after discharge for jaundice or other clinical problems such as late anemia. Depending on the incidence of hyperbilirubinemia within an institution, the criteria for decision making should vary according to its unique population.

Detection of heme oxygenase activity by measurement of CO.

Heme oxygenase (HO) is the first and rate-limiting step in degradation of heme, and in the presence of NADPH-cytochrome P-450 reductase it produces equimolar amounts of biliverdin and CO. CO produced in a closed system can be quantified as described in this unit by gas chromatography as a measure of HO activity in tissue slices, tissue homogenates, and tissue fractions.

Association of fetal and maternal carboxyhemoglobin levels in normal and Rh-alloimmune pregnancies.

OBJECTIVE: To compare paired antepartum fetal/maternal COHb ratios in whole blood from control and alloimmunized pregnancies and to examine the relationships between fetal and maternal COHb. METHODS: COHb levels were measured in paired fetal and maternal blood samples obtained at cordocentesis in 47 control and 16 Rh-alloimmunized pregnancies. COHb was determined by gas chromatography. Results were analyzed by t-test, regression and analysis of covariance. RESULTS: Although fetal/maternal COHb ratios for control and alloimmunized pregnancies were not statistically significantly different, i.e. 1. 11+/-0.04 and 1.26+/-0.09, respectively (P=0.09), fetal COHb levels were higher in Rh-alloimmunized fetuses (P=0.0002). Fetal COHb levels were also higher than paired maternal levels among the alloimmunized group (P=0.011), but not among the control group (1. 04+/-0.04, P=ns). In univariate regression analysis, fetal and maternal COHb levels were significantly correlated with one another in both control (r=0.52, P=0.0002) and alloimmunized pregnancy groups (r=0.52, P=0.05). Comparison of the slopes of the fetal versus maternal COHb plots for the two groups showed a significant difference (P=0.02), with the alloimmunized group having the steeper slope. CONCLUSION: Differences in the antepartum fetal-maternal COHb relationships in control and alloimmunized groups likely reflect increased endogenous CO production among alloimmunized fetuses as a result of pathologic hemolysis.

Carbon monoxide detection and biological investigations.

Even though the heme degradation pathway consists of only two reactions, it and its major enzyme (i.e. HO), nonetheless, impact other processes not only through the removal of excess heme, but also through the production of several metabolically active compounds. Thus CO and biliverdin along with reactive iron, Fe2, are the primordial products of this ancient, highly conserved reaction. That every component of the heme catabolic pathway is directly or indirectly related to other reactions involving oxygen or light is, perhaps, no accident of nature. That a fundamentally destructive event can be linked with a multiplicity of synthetic events and various biological effects, depending on the timing and location of the HO activity, is testament to the economy and the ultimate beauty of nature. Furthermore, the interaction of the heme catabolic pathway with that of the NOS system may lead to even more exciting avenues of research. It may be shown that the integrity of the heme catabolic pathway, which is ever present and plays a role in every tissue, is central to the existence of most complex organisms.