Bilirubin and Brain Injury

Definition

Developmental hyperbilirubinemia refers to the elevation of serum bilirubin values that occur in essentially every newborn infant in the first week of life. In classic earlier studies of formula-fed full-term infants, the serum bilirubin level rose gradually to a mean peak of approximately 6 mg/dL by 48 to 72 hours of life and then declined to a slightly elevated level, approximately 3 mg/dL, at approximately 5 days of life. Little change occurred thereafter until a second, gradual decline resulted in normal levels by approximately 2 weeks. With the current increase in breast-feeding in the United States and elsewhere, peak values for bilirubin clearly are greater than in the earlier studies. Thus in predominantly breast-fed term infants, the normal peak value is 8 to 11 mg/dL, and the decline is slower. In newborns of East Asian ancestry (predominantly breast-fed), the peak values, 10 to 14 mg/dL, are still higher. In the premature infant, the peak serum bilirubin concentration occurs later and is higher than in the full-term infant. More importantly, the term developmental hyperbilirubinemia in the premature infant is misleading because these higher values are potentially dangerous (see later) and phototherapy is recommended well before such levels are reached (see later).

The late preterm infant (i.e., 34 ^0/7 to 36 ^6/7 weeks of gestation) presents a situation intermediate between the clearly premature infant and the full-term infant (37 ^0/7 to 42 weeks of gestation). Thus several studies showed that such infants have a several-fold higher risk of significant hyperbilirubinemia in the first week of life. In a careful study through the first 7 days of life, late preterm infants exhibited a later and higher peak value of bilirubin than did term infants. Moreover, the higher values declined more slowly in the late-preterm infants. Overall, 10% of the full-term infants had hyperbilirubinemia requiring phototherapy, versus 25% in the late-preterm group. These data highlight the importance of particularly close follow-up of late-preterm infants after hospital discharge (see later).

Mechanisms

The mechanisms considered important in the genesis of developmental hyperbilirubinemia are shown in Table 30.2 . Although evidence has been mustered for all these factors, studies of the newborn monkey provided considerable insight into their relative importance. Thus the first phase of hyperbilirubinemia (see previous section) has been shown to relate to the combined effects of (1) increased bilirubin load to the liver and (2) decreased bilirubin-conjugating capacity. The source of the increased bilirubin load to the liver includes both hemoglobin and nonhemoglobin sources, as well as the enterohepatic circulation of bilirubin; the latter is increased in the newborn because of deficient bacterial reduction of bilirubin and increased activity of intestinal beta-glucuronidase. The defective bilirubin conjugating capacity is related to a negligible level of hepatic UGT1A1 activity at birth, which undergoes a rapid increase during the first week of postnatal life but does not approach adult levels for several months. Prematurity results in more severe neonatal hyperbilirubinemia, principally because of a delayed maturation of the hepatic UGT1A1. More recent studies in human infants refined the earlier observations. Thus the possibility of impaired uptake of bilirubin into the hepatocyte because of genetic variants of organic anion transporters (see earlier) is likely important, perhaps especially in infants of East Asian ancestry. The diminished levels of ligandin in hepatic cytosol are likely less important. Genetic variants of the UGT1A1 gene, as well as the long-recognized developmental deficiency of the glucuronyl transferase, are also important. Indeed, the prolonged jaundice associated with breast milk feedings is now considered a prevalent Gilbert syndrome phenotype. Breast milk contains several substances (beta-glucuronidase, lipoprotein lipase, pregnanediol) that augment the enterohepatic circulation bilirubin, which combined with Gilbert syndrome leads to prolonged jaundice .

Unconjugated Bilirubin

Although it is generally recognized that serum levels of unconjugated bilirubin must be elevated to cause neurotoxicity, the relationship between such elevations and brain injury is not simple. In the full-term infant with marked hyperbilirubinemia secondary to hemolytic disease , a correlation can be discerned between the occurrence of kernicterus and the maximal recorded level of serum bilirubin ( Table 30.4 ). In a review of 52 infants with hemolytic disease (33 Rh incompatibility, 19 ABO incompatibility) and comorbid factors, approximately 95% of whom developed kernicterus, the mean peak total serum bilirubin (TSB) in both groups was approximately 32 mg/dL , with an approximate range of 18 to 51 mg/dL .

The neural risk of marked hyperbilirubinemia in full-term infants without hemolysis is less clear. Indeed, an earlier analysis of available studies by Newman and Maisels suggested that the risk for neurological sequelae is distinctly less for hyperbilirubinemic infants without hemolytic disease compared with the risk for infants with hemolytic disease. Subsequent observations supported this contention, including a study by Newman and associates of 140 treated infants (phototherapy, n = 136) and exchange transfusion ( n = 5) with peak serum bilirubin levels largely between 25 and 29.9 mg/dL. There were no cases of kernicterus. Similarly, Gamaleldin and colleagues reported that bilirubin encephalopathy was only observed at a TSB greater than 31.5 mg/dL in infants ( n = 111) without evidence of hemolytic disease, sepsis, and low gestational age. However, multiple reports describe the occurrence of kernicterus, identified by neuropathological, neuroradiological, or clinical criteria, after apparent nonhemolytic hyperbilirubinemia. Furthermore in a review of 35 selected infants with nonhemolytic, “idiopathic” hyperbilirubinemia and documented acute or chronic bilirubin encephalopathy, all infants had peak TSB levels greater than 20 mg/dL. Indeed, more than 90% of the infants with kernicterus had peak levels higher than 25 mg/dL. Notably, however, fully 25% had peak levels lower than 30 mg/dL. Nevertheless, current observations suggest that hemolytic conditions are often overlooked in the absence of detailed search for immune-mediated mechanisms by advanced techniques. It seems reasonable to conclude that hyperbilirubinemia, in most cases, is a necessary but usually not a sufficient condition to explain kernicterus. Factors acting in concert with bilirubin, including duration of exposure to bilirubin or albumin binding of bilirubin, must be evaluated to seek a satisfactory explanation for the risk of developing kernicterus (see later). These issues are discussed in detail later (see ‘Relationship of Neurological Sequelae in Term Infants to Degree of Hyperbilirubinemia’ section). The relationships between neonatal bilirubin values and neurological outcome in premature infants are probably different in magnitude but not in principle from those in full-term infants. Indeed, kernicterus has been demonstrated repeatedly in the premature infant without marked hyperbilirubinemia (see later discussion). These infants usually have exhibited a variety of complicating conditions (e.g., acidosis, hyperbilirubinemia, sepsis, asphyxia, hypoalbuminemia, hypothermia, intraventricular hemorrhage) that likely augment bilirubin neurotoxicity. In one often-cited, relatively large collection of premature infants with kernicterus ( n = 6), peak TSB levels were in excess of 20 mg/dL (range, 22 to 26 mg/dL). However, in this report, the gestational age of the six infants ranged from 34 to 36 weeks. Studies of bilirubin-induced auditory disturbances in smaller premature infants (28 to 32 weeks of gestational age) document neurological dysfunction at much lower bilirubin levels. The critical issue of the premature infant is discussed later (see ‘Clinical Features’ section).

Free Bilirubin

Because of the recognition that the fraction of bilirubin not bound to albumin is the critical component involved in bilirubin entry into the brain and in neurotoxicity (see later) and because of the demonstration of kernicterus in premature infants without markedly elevated levels of unconjugated bilirubin, intensive investigation has been directed toward measurement of the quantity of that fraction of unconjugated serum bilirubin, namely, unbound (i.e., “free”) bilirubin. In general, premature infants with higher levels of free bilirubin exhibit kernicterus at postmortem examination more often than those with lower levels. In a recent study of 1100 extremely low-birth weight infants in whom total and unbound bilirubin levels were measured at 5 days, irrespective of the clinical status, an increasing level of unbound bilirubin was associated with higher rates of death or neurodevelopmental impairment, or hearing loss at 18 to 22 month follow-up. Disturbances of the brainstem auditory-evoked response (BSAER) are observed at lower levels of free bilirubin in premature versus full-term infants (see later). Experimental studies also support a relationship between free or unbound bilirubin and neuronal injury.

However, there is currently little agreement about what constitutes the neurotoxic threshold for unbound bilirubin and what values of unbound circulating bilirubin should be used for initiating treatment. Indeed, other determinants of risk (e.g., concerning albumin binding of bilirubin, status of the blood-brain barrier, and neuronal susceptibility; see Table 30.3 ) are often present in the sick premature infant and contribute to the likelihood of bilirubin injury (see subsequent sections). Notably, the measurement of circulating unbound bilirubin is generally not clinically available.

In contrast, measuring the total bilirubin/albumin molar ratio (BAMR) is clinically obtainable and has been shown to be highly correlated with unbound bilirubin. Although the BAMR is an imperfect surrogate of free bilirubin and CNS bilirubin exposure, during extreme hypoalbuminemia the BAMR may become a meaningful proxy of bilirubin neurotoxicity risk as exemplified by the phenomenon of low-bilirubin kernicterus. In the low-bilirubin kernicterus case series of Govaert and colleagues, serum albumin levels ranged from 1.3 to 1.9 g/dL and every neonate demonstrated an elevated BAMR that met or exceeded recommended BAMR exchange transfusion treatment thresholds.

Affinity of Newborn Albumin for Bilirubin

The affinity of albumin for bilirubin is less in the newborn than in the older infant. Adult levels of binding affinity are not reached until as late as 5 months of age. Moreover, binding affinity is lower in the premature infant than in the term infant and is lower in sick infants than in well infants. The explanation for the lower binding affinity of neonatal albumin is not entirely clear. The search for competing anions or for compositional differences in the protein as the unifying explanation has not been fruitful. The leading possibility is that a difference in conformation of the albumin is responsible. Moreover, it is likely that the conformational difference relates to the humoral environment of the neonatal albumin because adult serum albumin infused into newborns loses its superior binding affinity over 24 hours.

Endogenous Anions

Endogenous anions that may compete with bilirubin for albumin binding sites include nonesterified fatty acids and other organic anions. Nonesterified fatty acids are anions at physiological pH and are present in high concentrations with hypothermia, hypoxemia, hypoglycemia, sepsis, starvation, the administration of heparinized blood (through heparin’s activation of triglyceride lipase), and intravenous alimentation with lipid. A recent report describes the uncoupling of unbound from total bilirubin as a result of unbound free fatty acids in premature infants treated with intralipids. Studies of asphyxiated infants with metabolic acidosis demonstrated impaired bilirubin binding to albumin that could not be attributed to the low pH per se. Rather, the data indicated the presence of organic anions in the plasma of asphyxiated, acidotic infants that compete with bilirubin for albumin binding sites. This finding may represent one of several mechanisms of potential importance in enhancing bilirubin neurotoxicity with asphyxia ( Table 30.5 ).