Serum Neuron Specific Enolase And S100B Protein In Early Prediction Of Neonatal Bilirubin Brain Damage

Background and objectiveHyperbilirubinemia in neonates with high incidence rate, most prognosis is good, but a few severe hyperbilirubinemia can cause varying degrees of damage to the nervous system.Bilirubin encephalopathy refers to neonatal unconjugated hyperbilirubinemia occurs, due to increased serum bilirubin concentration or blood brain barrier damage, free bilirubin through blood brain barrier deposition in the basal nucleus, thalamus, subthalamic nucleus, cerebral cortex and other parts, brain injury induced by cerebral cell oxidative phosphorylation process.ystem, causing great harm to the family and the society.The diagnosis of bilirubin encephalopathy is not recognized and reliable gold standard. Clinical diagnosis depends on the clinical manifestation, brainstem auditory evoked potentials, visual evoked potential, neonatal behavioral neurological assessment, MRI etc. But these examinations, whose abnormal results will lag in diagnosis, have some limitations in clinical use. Because the early bilirubin encephalopathy often lack specific performance, most cases are mild symptoms or even no significant abnormal performance, so it is difficult to forecast bilirubin brain injury early and timely. The key to preventing bilirubin encephalopathy is early discovery and intervention.Therapeutic regimen of hyperbilirubinemia was formulated on the basis of the levels of serum total bilirubin mainly at home and abroad. Clinical observation has been suggested bilirubin brain damage can also occur in a "safe" level of TSB.According to the TSB level as the intervention is not reliable.In fact, there is a certain correlation between in high risk factors and bilirubin encephalopathy, such as neonatal blood brain barrier function, brain bilirubin level, metabolic acidosis, infection, serum albumin, bilirubin binding state, free bilirubin level.To reduced the incidence of neonatal bilirubin encephalopathy is a pediatrician’s target in the treatment of neonatal hyperbilirubinemia.Therefore, the clinical doctors need more objective and sensitive index to judge the toxic effect of bilirubin on the nervous system so that they can early detecte and prevent bilirubin encephalopathy.In recent years, research on neural biochemical markers of central nervous system injury has made great progress,such as S100B protein, neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP), fatty acid binding protein, creatine kinase brain isoenzyme (CKBB), β-amyloid precursor protein (β-APP), Tau protein etc. More commonly used in neonatal field is the S100B protein and NSE.S100B protein is a calcium binding protein and growth factor of axons, but also has many other important physiological functions which mainly distributes in the central nervous system glial cells and Schwann cells of the peripheral nervous system. It is the specific proteins of the nervous system, increasing level in serum is closely related with the degree of brain injury and prognosis.S100B protein as a marker for brain damage hae time variation after brain injury.S100B protein’s biological half-life is2h, discharged by the kidneys after metabolizing in the body, These physiological characteristics are conducive to dynamic monitoring.S100B protein can promote mitosis, effect glial cell growth and differentiation, maintain calcium homeostasis, promote the development of the brain. After cerebral injury synthesis and secretion of S100B protein will increase in order to repair nerve injury. After brain ischemia injury, Ca2+in the neurons will increase associated with S100B protein, which reduces neuronal mitochondrial dysfunction and protects the neurons.The level of S100B protein in predicting brain damage value is Very useful. It is more stability than other neural biochemical markers and not easy to be influenced by other factors.Research has shown that the protein level can reflect the severity of neonatal hypoxic-ischemic brain injury, can be used as a biochemical marker of prognosis of brain injury.NSE is the key glycolytic enzyme,.with molecular weight of78KD, stable physical and chemical properties. Normally NSE present in neurons and neuroendocrine tissues, in addition the surrounding neurons, endocrine glands, lymphocytes, erythrocytes, platelets also contains NSE. Under normal circumstances its level in the body fluid is extremely low. After nerve cell disintegration due to the brain damage, NSE can be tested in the cerebrospinal fluid and blood which reflect the degree of the central nervous system damage. Many studies have been proved that NSE concentration in blood and cerebrospinal fluid increased when the neonatal hypoxic ischemic brain injury and intracranial hemorrhage had happened.The two neural markers in many reports were used to monitor neonatal hypoxic ischemic brain injury, but there was little research and application in bilirubin encephalopathy. For now, there isn’t an accepted reference range in diagnosis of neonatal brain injury. The purpose of this study was to investigate that if the serum levels of NSE and S100B protein can predict early neonatal bilirubin encephalopathy,help the doctor to treat hyperbilirubinemia determine prognosis. Materials and methods1Material1.1The research objectFrom January to December2010,51normal full-term newborns aged from2～7days hospitalized in the affiliated Nanfang Hospital of Southern Medical University obstetrics and gynecology department were randomly selected as control group without any complications and complicating disease during pregnancy(male34cases, female17cases,15cases of cesarean section,36cases of vaginal delivery).115neonates with hyperbilirubinemia (unconjugated bilirubin increased) aged from2-28days hospitalized in Neonatal department were chosed to be the observed group(male66cases, female49cases,12cases of cesarean section,103cases of vaginal delivery). There were no obvious clinical manifestations of nervous system abnormalities(lethargy, poor response, dystonia, convulsion, apnea, opisthotonus, excessive crying, faint cry, trunk torsion spasm, light coma etc) in the observation group cases.1.2The diagnostic criteria of neonatal hyperbilirubinemia was refered to the third edition practice Neonatology.1.3Exclusion criteriaFamily history of hearing impairment, perinatal ototoxic drug use hi-story, birth asphyxia history, hypoxic ischemic encephalopathy, intracranial hemorrha ge or intracranial infection,TORCH intrauterine infection (Toxoplasma, rubella v irus, cytomegalovirus,herpes simplex virus and other), congenital malformatio n and inherited metabolic diseases,serious heart and lung disease。2Methods2.1Sample pretreatmentBlood samples of infants were collected on admission during venipuncture for bi- lirubin level or other biochemical tests. Venous blood samples were static in no antic-oagulation drying tube for30min and centrifuged at3,000rpm for10minutes. Serum samples were stored at-70℃until the assays were performed,except hemolysis.2.2Total serum bilirubin levels were measured by automatic biochemical analyzer (Japanese Olympus AU400).2.3NSE and S100B protein levels were measured by enzyme linked immunosorbent assay (ELISA), using the United States RB company production of ELISA kit and ELX800enzyme American Pet Company production scale instrument.2.4Brainstem auditory-evoked responseBAEP recordings were performed using a Nicolet Viking Onset IV system (Am-erican HIS company). All patients were informed and agreed by the family and clean-ed the external auditory canal before the test.In all testing cases the examination was completed during sleep, induced by feeding100g/L chloral hydrate at a dosage of0.5ml/kg. The skin sebum was wiped before pasted electrode. Tests were conducted in an electrically shielded soundproof room.2.5Neonatal Behavioral Neurological AssessmentTests were conducted on the28th day after birth, required in light-dark and quiet environment. After putting the baby alone in this environment about30minutes,the tests were begun in the state of sleep and In the middle of the two breast-feeding interval stage. Room temperature was requied24～28℃.All tests were completed in10minutes.Using the bow’s20NBNA method to tested the babys, each with0,1,2index score, out of40points,37points for the pass.2.6Statistical AnalysisAll statistical calculations were performed using the SPSS software program (ve-rsion release11.5, SPSS). Statistical significance was set at P<0.05. Results1. The levels of NSE,S100B protein and TSB were significantly higher in the study group than in the control group (P=0.000). The levels of NSE and S100B were positively correlated with the level of TSB in study group.2. The results showed that the levels of NSE and S100B were higher significantly when TSB level exceeded278.1umol/L(P<0.05).3. The levels of NSE, S100B and TSB were negatively correlation with NBNA. The levels of NSE and S100B in infants whose NBNA were abnormal were significa-ntly higer than those of the infants with normal NBNA (P<0,05)4. The levels of NSE、S100B and TSB in infants whose BAEP were abnormal were significantly higer than those of the infants with normal BAEP (P<0.05).The sensitivities of measuring NSE and S100B were higher than BAEP as a procedure for predicting bilirubin brain damage.5. With the analysis of receiver operating characteristic curves(ROCC), it was determined that NSE level of52.7ng/ml and S100B level of4108.8ng/L provided the sensitivity of70%and the specificity of82.2%respectively.Conclusions1. NSE and S100B protein increased with the concentration of TSB which were the sensitive index of bilirubin brain damage. The levels of NSE and S100B were positively correlated with the level of TSB in study group.The levels of NSE, S100B were negatively correlation with NBNA.2. The levels of NSE and S100B were higher significantly when TSB level exceeded278.1umol/L, suggesting that brain damage may happen even though TSB does not reach340umol/L. 3. The sensitivity and specificity of NSE and S100B protein to prompt brain damage were70%and82.2%. The sensitivity of NSE and S100B protein to predicte brain damage were hgher than BAEP.4. The serum NSE and S100B protein were valuable neural markers in early predicting bilirubin brain damage in term newborns.They can help to determine the condition and interfere with the treatment of hyperbilirubinemia.