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Specific Enolase for Detection of Auditory Neuropathy in Neonatal

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Specific Enolase for Detection of Auditory Neuropathy in Neonatal Evaluation of Auditory Brainstem Evoked Responses and Neuron- Specific Enolase For Detection of Auditory Neuropathy in Neonatal Hyperbilirubinemia Samir Tamer Abdullah,1 Abdalraof Omar Abd Albaki,2 and Khaled Salah Osman3 From the Departments of Pediatrics,1 Neurology2 and Clinical Pathology,3 Al -Minya University Hospital, Egypt Abstract: This study was designed to evaluate the correlation between neuron-specific enolase and neonatal hyperbilirubinemia in children diagnosed as having auditory neuropathy by auditory brainstem evoked responses. Thirty infants admitted in neonatology unit of AL-Minya university hospital in the period from July 2002 to January 2004 for treatment from hyperbilirubinemia and 20 neonates as a control group. All infants had bilirubin levels above 20mg/dl and had a full work-up for hyperbilirubinemia. All study group infants were treated with phototherapy, and 16 infants required blood exchange transfusion as well. Hyperbilirubinemic infants were placed into two groups according to serum bilirubin values group (A): total bilirubin 20-25mg/dl, group (B): total bilirubin > 25 mg/dl. The control group consisted of 20 healthy full-term neonates with bilirubin levels within physiologic ranges (< 13 mg/dl). Serum samples for neuron specific enolase (NSE) determination were taken on the day of admission and stored at -20° C until the time of assayed by a commercial enzyme immunoassay (EIA) kit. All hyperbilirubinemic infants in the study were evaluated with auditory brain stem evoked responses (ABERs) and transient evoked otoacoustic emission (TEOAE) tests. ABERs were recorded using Nihon Kohden 4 channels equipment, while TEOAEs were obtained by using the quickscreen option of the ILO 92 OAE System. The results showed no significant differences between serum NSE value in hyperbilirubinemic groups (50.19 ±34.37) when compared with control infants (44.50 ± 27.68 ng/ml) (p = 0. 253). A significant difference was detected between serum NSE values of group A (33.29 ± 16. 98 ng/ml) and group B (67.09± 39.33 ng/ml) (p = 0.02). NSE and total bilirubin levels of patients with absent ABRs but present TEOAEs (36.43 ± 16.47 ng/ml and 25.06 ± 4.25 mg/dl, reciprocal ) were significantly higher than those of the patients with normal ABRs (70.83 ± 43.82 ng/ml and 31.29± 7.34 mg/dl , reciprocal), (p = 0.001). No correlation was found between serum NSE and bilirubin values (r-= 0.15, p = 0.33). There was no relationship between NSE concentration and the duration of the hyperbilirubinemia (r = 0.29, p=0.23). Conclusion: In this preliminary study, although we could not demonstrate any correlation between serum NSE and bilirubin levels but NSE levels were significantly higher in infants with auditory neuropathy which diagnosed by ABER. Thus, this finding indicated the importance of a close follow-up with dual screening of hearing by ABER and TEOAEs in hyperbilirubinemic newborns to avoid the auditory neuropathy. Biochemical index of neuronal damage (e.g. NSE) and ABERs can be used to evaluate the neurological sequels of neonatal hyperbilirubinemia like auditory neuropathy. For appropriate results to demonstrate the role of NSE and ABER for diagnosis of auditory neuropathy in neonatal hyperbilirubinemia, we must test a much larger sample of subjects in the future. Introduction: information on the sustained effects of bilirubin in the body.2 Hyperbilirubinemia is a common phenomenon in the Enolase is a glycolytic enzyme found in the cytoplasm neonatal period and attracts a lot of attention due to of neurons and cells with neuroendocrine its potential for neurotoxicity. The most serious differentiation. Enolase is a dimer isoenzyme with complication of neonatal jaundice is kernicterus.1 three possible subunits α, β and γ, the α α and γ γ Several studies have shown that bilirubin may exert isoenzymes are predominantly in neurons of the effects on neurons via multiple mechanisms including central nervous system and peripheral nervous inhibition of protein phosphorylation and release of system and are called neuron- specific enolases neurotransmitters. However, these studies were (NSEs).3 NSEs are released into both cerebrospinal carried out in vitro, and the results provide limited fluid (CSF) and serum after damage to the CNS. Alexandria Journal of Pediatrics, Volume 19, Number 2, July 2005 423 When brain tissue suffers an injury , the increased biochemical index of neuronal damage) and auditory membrane permeability of neurons causes leakage of neuropathy diagnosed by ABER and TEOAEs. proteins such as NSEs into CSF and / or blood, so NSEs assays in CSF or serum are not specific for a Subjects and Methods: single neurologic disease.4 Hansen et al.,5 in 1992, The study was carried out on 50 full term infants. have shown that assays for NSEs in CSF and plasma Thirty infants admitted for treatment from are helpful in detecting neuronal damage in bilirubin hyperbilirubinemia in the neonatology unit of AL- encephalopathy, but there have been no reports Minya University Hospital in the period from July 2002 regarding the sensitivity of NSEs in predicting the to January 2004 were included in the study and 20 high –risk group for brain damage among neonates as a control group. They were 12 males (40 hyperbilirubinemic newborn infants. %) and 18 females (60%) with gestational age ±37 Neonatal Hyperbilirubinemia is a common cause of weeks and postnatal age 7 days (2-18). The control early onset sensorineural hearing loss. Rance et al.,6 group were 20 healthy full term neonates, they were 8 in 2002, reported that this occurred in 10 of 20 of their males (40%) and 12 females (60%). None of the auditory neuropathy cases with bilirubin infants in the study group had a fifth-minute Apgar concentrations >350 μ mol/l. There is no exact score of less than 7. All study group infants were method to detect the extent of the neurotoxicity of treated with phototherapy, and 16 infants required bilirubin, on the other hand the auditory pathway is blood exchange transfusion according to the known to be one of the most sensitive parts of the recommendations of the American Academy of central nervous system (CNS) to this toxic agent, so Pediatrics. All infants had bilirubin levels above auditory brainstem evoked responses (ABERs) are a 20mg/dl and had a full work-up for hyperbilirubinemia, sensitive indicator for bilirubin neurotoxicity in which included blood groups, complete blood counts, humans.7 Abnormal ABERs have been found in reticulocyte count, Coomb's test, peripheral smear, human neonates with hyperbilirubinemia, where urinalysis for urinary reducing substances and urine ABER changes induced by bilirubin may be progress culture where indicated. Infants who showed from reversible prolongation of the absolute latencies evidence of hemolytic anemia (hematocrit value of waves III and V followed by loss of wave amplitude <45% with increased reticulocytic count and/or until the inability to detect an identifiable wave. These evidence of hemolysis in peripheral smear) were reversible changes may persist for up to 24 hours excluded from the study. after the decrease in serum total bilirubin (TB) In the study group, the bilirubin levels of all the concentrations. Furthermore, prolonged bilirubin patients returned to normal after therapy, which toxicity may cause irreversible sensorineural hearing eliminated Crigler-Najjar syndrome as a possible loss.8 Transient evoked otoacoustic emissions cause. Hyperbilirubinemic infants were placed into (TEOAEs) are sensitive to outer hair cell dysfunction, two groups according to serum bilirubin values; group whereas auditory brainstem evoked responses (A): total bilirubin 20-25 mg/dl and group (B): total (ABERs) are sensitive to the eight nerve and auditory bilirubin > 25 mg/dl. The control group consisted of 20 brain stem pathway. The diagnosis of auditory healthy full-term neonates with bilirubin levels within neuropathy is made when TEOAEs are present and physiologic ranges (< 13 mg/dl). ABERs are absent.9 Serum samples for NSE determination were taken on The pathogenesis of bilirubin encephalopathy is the day of admission, when the infant had the highest complex, clinical factors, bilirubin-albumin binding and measured bilirubin concentration, before any therapy the integrity of the blood-brain barrier all are thought had been initiated. All samples were immediately to play significant roles in bilirubin toxicity.10 The frozen and stored at -20° C until the time of analysis. specific sites and mechanisms of auditory Serum samples with any visible hemolysis were neuropathy are not yet known the possible sites of discarded. NSEs were assayed by a commercial auditory neuropathy include the inner hair cells, the enzyme immunoassay (EIA) kit. tectorial membrane, the synaptic junction between All hyperbilirubinemic infants in the study were the inner hair cells, auditory neurons in the spiral evaluated with ABER and transient-evoked ganglion, the eighth nerve fibers, or any combination otoacoustic emission (TEOAE) tests before discharge of these. Neural problems may be axonal or after a mean duration of hospitalization of 4 ± 1days. demyelinating, afferent as well as efferent pathways The patients were tested either during natural sleep may be also involved.11 or after sedation with chloral hydrate. ABERs were The current study was designed to evaluate whether recorded using Nihon Kohden 4 channels equipment. a correlation exists between increased serum bilirubin Electrodes were placed on the vertex (Cz) as a non- and neuron-specific enolase (NSE) assays (a inverting electrode, on both mastoid- (Ai and Ac) as Alex J Pediatr, 19(2),July 2005 424 inverting electrodes, and on the forehead as a μ s in duration) with an 80/s repetition rate, at 80 ± 6 ground. The responses were filtered with a band pass dBSPL. of 100-2500 Hz. The analysis time was 15ms. Statistical Methods: Alternating-polarity clicks of 100 ms duration were Data are indicated as mean ± standard error of presented monaurally with a repetition rate of 11/s.
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