Fluorescent protein-based detection of unconjugated bilirubin in newborn serum.

Increased serum levels of unconjugated bilirubin are associated with the development of brain damage in newborns. In current clinical settings, there are no methods for directly determining serum levels of unconjugated bilirubin. UnaG, a fluorescent protein from Japanese eel muscle that specifically binds to unconjugated bilirubin was used in this study. Linear regression analysis was carried out to compare unconjugated bilirubin levels measured by UnaG and conventional bilirubin oxidase methods. Unconjugated bilirubin levels in the serum of newborns who were untreated or treated with phototherapy were compared. Effects of interfering factors in the serum (conjugated bilirubin, hemoglobin, and lipid) on unconjugated bilirubin concentration measured by the UnaG method were also evaluated. Unconjugated bilirubin levels measured by the UnaG method were highly correlated with those determined by the bilirubin oxidase assay. Unconjugated bilirubin levels determined by bilirubin oxidase and UnaG assays were similar in serum samples containing conjugated bilirubin. The performance of the UnaG assay was unaffected by phototherapy and the presence of serum hemoglobin and lipid emulsion. These results demonstrate the clinical applicability of the UnaG method for direct measurement of unconjugated bilirubin levels in newborn serum.

Transcutaneous bilirubin monitoring predicts unexplained late-onset hemolysis in a very low birthweight infant.

BACKGROUND: In term infants, transcutaneous bilirubin (TcB) monitoring can be used to predict hemolytic hyperbilirubinemia. However, it is not clear whether the technique can also be used to predict unexplained late-onset hemolysis in very low birthweight (VLBW) infants. CASE PRESENTATION: The case was an infant with a birthweight of 1154 g who developed unexplained late-onset hemolysis at 8 days of age. The hyperbilirubinemia rapidly worsened, and therefore both phototherapy and exchange transfusion were performed. TcB levels were measured using the JM-105 jaundice meter and found to have increased by >3 mg/dL since before the onset, demonstrating for the first time that the device clearly detects changes in hemolytic rate. CONCLUSIONS: Although TcB levels did not correspond directly with total serum bilirubin levels in VLBW infants, the two values exhibited parallel changes in this case. Therefore, serial TcB monitoring may be useful in the early prediction of unexplained late-onset hemolysis in VLBW infants.

Screening for Hyperbilirubinemia in Japanese Very Low Birthweight Infants Using Transcutaneous Bilirubinometry.

OBJECTIVES: To assess the accuracy of transcutaneous bilirubin (TcB) measurements at 5 different body sites in Japanese very low birthweight (VLBW) infants and to determine a cut-off value of TcB to detect total serum/plasma bilirubin (TB) levels ≥10 mg/dL (171 µM). STUDY DESIGN: In a prospective multicenter study, 85 Japanese VLBW infants were enrolled from 5 neonatal intensive care units during the study period. A total of 383 blood samples from infants not receiving phototherapy or ≥24 hours postphototherapy were analyzed. TcB was measured at the forehead, sternum, upper back, lower abdomen, and waist within 1 hour of blood collection. Linear regression analysis and Bland-Altman plots were used to compare TcB values at each site with TB levels. The TcB cut-off value for detecting TB ≥10 mg/dL was determined by receiver operating characteristics curve analysis. RESULTS: TcB significantly correlated with TB, but the coefficient of determination varied among the sites (forehead: 0.5294, sternum: 0.6488, upper back: 0.6321, lower abdomen: 0.5430, waist: 0.7396). At a TcB value ≥8, the sensitivity was 100% at the sternum and upper back, 85% at the waist, 84% at the forehead, and 64% at the lower abdomen to detect TB ≥10 mg/dL. CONCLUSIONS: In Japanese VLBW infants, the accuracy of TcB measurements varies according to body site. TcB ≥8 on the sternum or upper back is more reliable than that on the forehead, lower abdomen, or waist to detect TB levels ≥10 mg/dL.

Postnatal serum concentrations of endogenous free fatty acids in newborns admitted to the neonatal intensive care unit: effects on unbound bilirubin.

BACKGROUND: Few studies have reported the characterization of postnatal serum concentrations of endogenous free fatty acids (FFAs) in high-risk newborns and their effects on unbound bilirubin (UB). METHODS: Serum concentrations of FFA, albumin (Alb), UB and total bilirubin (TB) were measured in 713 samples obtained within 5 days after birth from 439 newborns without intravenous lipid supplementation admitted to the neonatal intensive care unit (NICU). Serum FFA was reported as the day-specific percentile-based curve. Serum FFA and FFA/Alb ratios were compared in term and preterm patients. To assess the impact of FFA on UB, daily changes in FFA/Alb and UB/TB ratios were compared in term patients without receiving phototherapy or any drugs, and linear regression analysis was performed between FFA/Alb ratio and serum UB concentration or UB/TB ratio using 140 sera with hyperbilirubinemia of term and preterm patients. RESULTS: A percentile-based curve showed that serum FFA peaked at 1 day of age and progressively decreased. Serum FFA and the FFA/Alb ratio were significantly higher in term than in preterm patients at birth and 1 and 3 days of age. FFA/Alb ratio significantly changed over 5 days after birth, but UB/TB ratio remained constant. FFA/Alb ratio did not correlate with serum UB concentration or UB/TB ratio in sera with hyperbilirubinemia. CONCLUSIONS: We assessed postnatal concentrations of serum FFA in a large number of high-risk newborns admitted to the NICU. The concentration of endogenous FFAs in newborns admitted to the NICU was not rising until it influenced UB.

Nervous system disruption and concomitant behavioral abnormality in early hatched pufferfish larvae exposed to heavy oil.

BACKGROUND AND OBJECTIVE: Spills of heavy oil (HO) over the oceans have been proven to have an adverse effect on marine life. It has been hypothesized that exposure of early larvae of sinking eggs to HO leads largely to normal morphology, whereas abnormal organization of the developing neural scaffold is likely to be found. HO-induced disruption of the nervous system, which controls animal behavior, may in turn cause abnormalities in the swimming behavior of hatched larvae. To clarify the toxicological effects of HO, we performed exposure experiments and morphological and behavioral analyses in pufferfish (Takifugu rubripes) larvae. EXPERIMENTAL PROCEDURE: Fertilized eggs of pufferfish were exposed to 50 mg/L of HO for 8 days and transferred to fresh seawater before hatching. The hatched larvae were observed for their swimming behavior, morphological appearance, and construction of muscles and nervous system. RESULTS AND DISCUSSION: In HO-exposed larvae, we did not detect any anomaly of body morphology. However, they showed an abnormal swimming pattern and disorganized midbrain, a higher center controlling movement. Our results suggest that HO-exposed fishes suffer developmental disorder of the brain that triggers an abnormal swimming behavior and that HO may be selectively toxic to the brain and cause physical disability throughout the life span of these fishes.

Acetylated YY1 regulates Otx2 expression in anterior neuroectoderm at two cis-sites 90 kb apart.

The mouse homeobox gene Otx2 plays essential roles at each step and in every tissue during head development. We have previously identified a series of enhancers that are responsible for driving the Otx2 expression in these contexts. Among them the AN enhancer, existing 92 kb 5' upstream, directs Otx2 expression in anterior neuroectoderm (AN) at the headfold stage. Analysis of the enhancer mutant Otx2(DeltaAN/-) indicated that Otx2 expression under the control of this enhancer is essential to the development of AN. This study demonstrates that the AN enhancer is promoter-dependent and regulated by acetylated YY1. YY1 binds to both the AN enhancer and promoter region. YY1 is acetylated in the anterior head, and only acetylated YY1 can bind to the sequence in the enhancer. Moreover, YY1 binding to both of these two sites is essential to Otx2 expression in AN. These YY1 binding sites are highly conserved in AN enhancers in tetrapods, coelacanth and skate, suggesting that establishment of the YY1 regulation coincides with that of OTX2 function in AN development in an ancestral gnathostome.