Stefania Mondello

Elevated Levels of Serum Glial Fibrillary Acidic Protein Breakdown Products in Mild and Moderate Traumatic Brain Injury Are Associated With Intracranial Lesions and Neurosurgical Intervention

STUDY OBJECTIVE This study examines whether serum levels of glial fibrillary acidic protein breakdown products (GFAP-BDP) are elevated in patients with mild and moderate traumatic brain injury compared with controls and whether they are associated with traumatic intracranial lesions on computed tomography (CT) scan (positive CT result) and with having a neurosurgical intervention. METHODS This prospective cohort study enrolled adult patients presenting to 3 Level I trauma centers after blunt head trauma with loss of consciousness, amnesia, or disorientation and a Glasgow Coma Scale (GCS) score of 9 to 15. Control groups included normal uninjured controls and trauma controls presenting to the emergency department with orthopedic injuries or a motor vehicle crash without traumatic brain injury. Blood samples were obtained in all patients within 4 hours of injury and measured by enzyme-linked immunosorbent assay for GFAP-BDP (nanograms/milliliter). RESULTS Of the 307 patients enrolled, 108 were patients with traumatic brain injury (97 with GCS score 13 to 15 and 11 with GCS score 9 to 12) and 199 were controls (176 normal controls and 16 motor vehicle crash controls and 7 orthopedic controls). Receiver operating characteristic curves demonstrated that early GFAP-BDP levels were able to distinguish patients with traumatic brain injury from uninjured controls with an area under the curve of 0.90 (95% confidence interval [CI] 0.86 to 0.94) and differentiated traumatic brain injury with a GCS score of 15 with an area under the curve of 0.88 (95% CI 0.82 to 0.93). Thirty-two patients with traumatic brain injury (30%) had lesions on CT. The area under these curves for discriminating patients with CT lesions versus those without CT lesions was 0.79 (95% CI 0.69 to 0.89). Moreover, the receiver operating characteristic curve for distinguishing neurosurgical intervention from no neurosurgical intervention yielded an area under the curve of 0.87 (95% CI 0.77 to 0.96). CONCLUSION GFAP-BDP is detectable in serum within an hour of injury and is associated with measures of injury severity, including the GCS score, CT lesions, and neurosurgical intervention. Further study is required to validate these findings before clinical application.

Occult hepatitis B virus in liver tissue of individuals without hepatic disease

BACKGROUND/AIMS While many data are available concerning occult hepatitis B virus (HBV) infection in patients with hepatic disorders, there is little information about this cryptic infection in individuals without liver disease. The aim of this study was to investigate the prevalence of occult HBV in the general population by examining liver specimens from a large series of HBV-surface-antigen negative individuals with no clinical and biochemical evidence of liver disease. METHODS The presence of HBV DNA was evaluated by testing, through polymerase chain reaction techniques, DNA extracts from 98 liver-disease-free individuals who underwent liver resection or needle biopsy during abdominal surgery. Sixteen of them were anti-HBV-core antigen (anti-HBc) positive and 82 were HBV serum-marker negative. All patients were negative for antibody to hepatitis C virus. RESULTS Occult HBV infection was revealed in 16 of the 98 cases (16.3%). In particular, 10/16 anti-HBc positive (62.5%) versus 6/82 (7.3%) HBV-seronegative individuals were occult carriers (p<0.0001). CONCLUSIONS This study revealed that about 1/6 of the Italian general population might be carriers of occult HBV infection, and this condition is significantly associated with the anti-HBc positive status.

αII-Spectrin Breakdown Products (SBDPs): Diagnosis and Outcome in Severe Traumatic Brain Injury Patients

In this study we assessed the clinical utility of quantitative assessments of alphaII-spectrin breakdown products (SBDP145 produced by calpain, and SBDP120 produced by caspase-3) in cerebrospinal fluid (CSF) as markers of brain damage and outcome after severe traumatic brain injury (TBI). We analyzed 40 adult patients with severe TBI (Glasgow Coma Scale [GCS] score <or=8) who underwent ventriculostomy. Patients requiring CSF drainage for other medical reasons served as controls. CSF samples were taken at admission and every 6 h thereafter for a maximum of 7 days and assessed using novel quantitative fragment-specific ELISAs for SBDPs. Outcome was assessed using the 3-month Glasgow Outcome Scale. Mean CSF levels of SBDPs were significantly higher in TBI patients than in controls at all time points examined. Different temporal release patterns of CSF SBDP145 and SBDP120 were observed. SBDP145 provided accurate diagnoses at all time points examined, while SBDP120 release was more accurate 24 h after injury. Within 24 h after injury, SBDP145 CSF concentrations significantly correlated with GCS scores, while SBDP120 levels correlated with age. SBDP levels were significantly higher in patients who died than in those who survived. SBDP145 levels (>6 ng/mL) and SBDP120 levels (>17.55 ng/mL) strongly predicted death (odds ratio 5.9 for SBDP145, and 18.34 for SBDP120). The time course of SBDPs in nonsurvivors also differed from that of survivors. These results suggest that CSF SBDP levels can predict injury severity and mortality after severe TBI, and can be useful complements to clinical assessment.

Neuronal and glial markers are differently associated with computed tomography findings and outcome in patients with severe traumatic brain injury: A case control study

IntroductionAuthors of several studies have studied biomarkers and computed tomography (CT) findings in the acute phase after severe traumatic brain injury (TBI). However, the correlation between structural damage as assessed by neuroimaging and biomarkers has not been elucidated. The aim of this study was to investigate the relationships among neuronal (Ubiquitin carboxy-terminal hydrolase L1 [UCH-L1]) and glial (glial fibrillary acidic protein [GFAP]) biomarker levels in serum, neuroradiological findings and outcomes after severe TBI.MethodsThe study recruited patients from four neurotrauma centers. Serum samples for UCH-L1 and GFAP were obtained at the time of hospital admission and every 6 hours thereafter. CT scans of the brain were obtained within 24hrs of injury. Outcome was assessed by Glasgow Outcome Scale (GOS) at discharge and at 6 months.Results81 severe TBI patients and 167 controls were enrolled. The mean serum levels of UCH-L1 and GFAP were higher (p < 0.001) in TBI patients compared to controls. UCH-L1 and GFAP serum levels correlated significantly with Glasgow Coma Scale (GCS) and CT findings. GFAP levels were higher in patients with mass lesions than in those with diffuse injury (2.95 ± 0.48 ng/ml versus 0.74 ± 0.11 ng/ml) while UCH-L1 levels were higher in patients with diffuse injury (1.55 ± 0.18 ng/ml versus 1.21 ± 0.15 ng/ml, p = 0.0031 and 0.0103, respectively). A multivariate logistic regression showed that UCH-L1 was the only independent predictor of death at discharge [adjusted odds ratios 2.95; 95% confidence interval, 1.46-5.97], but both UCH-L1 and GFAP levels strongly predicted death 6 months post-injury.ConclusionsRelationships between structural changes detected by neuroimaging and biomarkers indicate each biomarker may reflect a different injury pathway. These results suggest that protein biomarkers could provide better characterization of subjects at risk for specific types of cellular damage than that obtained with neuroimaging alone, as well as provide valuable information about injury severity and outcome after severe TBI.

Serum levels of ubiquitin C-terminal hydrolase distinguish mild traumatic brain injury from trauma controls and are elevated in mild and moderate traumatic brain injury patients with intracranial lesions and neurosurgical intervention.

BACKGROUND: This study compared early serum levels of ubiquitin C-terminal hydrolase (UCH-L1) from patients with mild and moderate traumatic brain injury (TBI) with uninjured and injured controls and examined their association with traumatic intracranial lesions on computed tomography (CT) scan (CT positive) and the need for neurosurgical intervention (NSI). METHODS: This prospective cohort study enrolled adult patients presenting to three tertiary care Level I trauma centers after blunt head trauma with loss of consciousness, amnesia, or disorientation and a Glasgow Coma Scale (GCS) score 9 to 15. Control groups included normal uninjured controls and nonhead injured trauma controls presenting to the emergency department with orthopedic injuries or motor vehicle crash without TBI. Blood samples were obtained in all trauma patients within 4 hours of injury and measured by enzyme-linked immunosorbent assay for UCH-L1 (ng/mL ± standard error of the mean). RESULTS: There were 295 patients enrolled, 96 TBI patients (86 with GCS score 13–15 and 10 with GCS score 9–12), and 199 controls (176 uninjured, 16 motor vehicle crash controls, and 7 orthopedic controls). The AUC for distinguishing TBI from uninjured controls was 0.87 (95% confidence interval [CI], 0.82–0.92) and for distinguishing those TBIs with GCS score 15 from controls was AUC 0.87 (95% CI, 0.81–0.93). Mean UCH-L1 levels in patients with CT negative versus CT positive were 0.620 (±0.254) and 1.618 (±0.474), respectively (p < 0.001), and the AUC was 0.73 (95% CI, 0.62–0.84). For patients without and with NSI, levels were 0.627 (0.218) versus 2.568 (0.854; p < 0.001), and the AUC was 0.85 (95% CI, 0.76–0.94). CONCLUSION: UCH-L1 is detectable in serum within an hour of injury and is associated with measures of injury severity including the GCS score, CT lesions, and NSI. Further study is required to validate these findings before clinical application. LEVEL OF EVIDENCE: II, prognostic study.

Biokinetic Analysis of Ubiquitin C-Terminal Hydrolase-L1 (UCH-L1) in Severe Traumatic Brain Injury Patient Biofluids

Ubiquitin C-terminal hydrolase-L1 (UCH-L1) is a neuron-specific enzyme that has been identified as a potential biomarker of traumatic brain injury (TBI). The study objectives were to determine UCH-L1 exposure and kinetic metrics, determine correlations between biofluids, and assess outcome correlations in severe TBI patients. Data were analyzed from a prospective, multicenter study of severe TBI (Glasgow Coma Scale [GCS] score ≤ 8). Cerebrospinal fluid (CSF) and serum data from samples taken every 6 h after injury were analyzed by enzyme-linked immunosorbent assay (ELISA). UCH-L1 CSF and serum data from 59 patients were used to determine biofluid correlations. Serum samples from 86 patients and CSF from 59 patients were used to determine outcome correlations. Exposure and kinetic metrics were evaluated acutely and up to 7 days post-injury and compared to mortality at 3 months. There were significant correlations between UCH-L1 CSF and serum median concentrations (r(s)=0.59, p<0.001), AUC (r(s)=0.3, p=0.027), Tmax (r(s)=0.68, p<0.001), and MRT (r(s)=0.65, p<0.001). Outcome analysis showed significant increases in median serum AUC (2016 versus 265 ng/mL*min, p=0.006), and Cmax (2 versus 0.4 ng/mL, p=0.003), and a shorter Tmax (8 versus 19 h, p=0.04) in those who died versus those who survived, respectively. In the first 24 h after injury, there was a statistically significant acute increase in CSF and serum median Cmax((0-24h)) in those who died. This study shows a significant correlation between UCH-L1 CSF and serum median concentrations and biokinetics in severe TBI patients, and relationships with clinical outcome were detected.

Blood-based diagnostics of traumatic brain injuries

Traumatic brain injury is a major health and socioeconomic problem that affects all societies. However, traditional approaches to the classification of clinical severity are the subject of debate and are being supplemented with structural and functional neuroimaging, as the need for biomarkers that reflect elements of the pathogenetic process is widely recognized. Basic science research and developments in the field of proteomics have greatly advanced our knowledge of the mechanisms involved in damage and have led to the discovery and rapid detection of new biomarkers that were not available previously. However, translating this research for patients’ benefits remains a challenge. In this article, we summarize new developments, current knowledge and controversies, focusing on the potential role of these biomarkers as diagnostic, prognostic and monitoring tools of brain-injured patients.

Brain Injury Biomarkers May Improve the Predictive Power of the IMPACT Outcome Calculator

Outcome prediction following severe traumatic brain injury (sTBI) is a widely investigated field of research. A major breakthrough is represented by the IMPACT prognostic calculator based on admission data of more than 8500 patients. A growing body of scientific evidence has shown that clinically meaningful biomarkers, including glial fibrillary acidic protein (GFAP), ubiquitin C-terminal hydrolase-L1 (UCH-L1), and αII-spectrin breakdown product (SBDP145), could also contribute to outcome prediction. The present study was initiated to assess whether the addition of biomarkers to the IMPACT prognostic calculator could improve its predictive power. Forty-five sTBI patients (GCS score≤8) from four different sites were investigated. We utilized the core model of the IMPACT calculator (age, GCS motor score, and reaction of pupils), and measured the level of GFAP, UCH-L1, and SBDP145 in serum and cerebrospinal fluid (CSF). The forecast and actual 6-month outcomes were compared by logistic regression analysis. The results of the core model itself, as well as serum values of GFAP and CSF levels of SBDP145, showed a significant correlation with the 6-month mortality using a univariate analysis. In the core model, the Nagelkerke R(2) value was 0.214. With multivariate analysis we were able to increase this predictive power with one additional biomarker (GFAP in CSF) to R(2)=0.476, while the application of three biomarker levels (GFAP in CSF, GFAP in serum, and SBDP145 in CSF) increased the Nagelkerke R(2) to 0.700. Our preliminary results underline the importance of biomarkers in outcome prediction, and encourage further investigation to expand the predictive power of contemporary outcome calculators and prognostic models in TBI.

Glial Neuronal Ratio: A Novel Index for Differentiating Injury Type in Patients with Severe Traumatic Brain Injury

Neurobiochemical marker levels in blood after traumatic brain injury (TBI) may reflect structural changes detected by neuroimaging. This study evaluates whether correlations between neuronal (ubiquitin carboxy-terminal hydrolase-L1 [UCH-L1]) and glial (glial fibrillary acidic protein [GFAP]) biomarkers may be used as an indicator for differing intracranial pathologies after brain trauma. In 59 patients with severe TBI (Glasgow Coma Scale [GCS] score≤8) serum samples were obtained at the time of hospital admission and analyzed for UCH-L1 and GFAP. Glial neuronal ratio (GNR) was evaluated as the ratio between GFAP and UCH-L1 concentrations. A logistic regression analysis was used to identify variables associated with type of injury. GNR had a median of 0.85 and was positively correlated with age (R=0.45, p=0.003). Twenty-nine patients presented with diffuse injury and 30 with focal mass lesions as assessed by CT scan at admission and classified according to the Marshall Classification. GNR was significantly higher in the focal mass lesion group compared with the diffuse injury group (1.77 versus 0.48, respectively; p=0.003). Receiver operating characteristic curve analysis showed that GNR discriminated between types of injury (area under the curve [AUC]=0.72; p=0.003). GNR was more accurate earlier (≤12 h after injury) than later (AUC=0.80; p=0.002). Increased GNR was independently associated with type of injury, but not age, gender, GCS score, or mechanism of injury. GNR was significantly higher in patients who died, but was not an independent predictor of death. The data from the present study indicate that GNR provides valuable information about different injury pathways, which may be of diagnostic significance. In addition, GNR may help to identify different pathophysiological mechanisms following different types of brain trauma, with implications for therapeutic interventions.

Human Traumatic Brain Injury Induces Autoantibody Response against Glial Fibrillary Acidic Protein and Its Breakdown Products

The role of systemic autoimmunity in human traumatic brain injury (TBI) and other forms of brain injuries is recognized but not well understood. In this study, a systematic investigation was performed to identify serum autoantibody responses to brain-specific proteins after TBI in humans. TBI autoantibodies showed predominant immunoreactivity against a cluster of bands from 38–50 kDa on human brain immunoblots, which were identified as GFAP and GFAP breakdown products. GFAP autoantibody levels increased by 7 days after injury, and were of the IgG subtype predominantly. Results from in vitro tests and rat TBI experiments also indicated that calpain was responsible for removing the amino and carboxyl termini of GFAP to yield a 38 kDa fragment. Additionally, TBI autoantibody staining co-localized with GFAP in injured rat brain and in primary rat astrocytes. These results suggest that GFAP breakdown products persist within degenerating astrocytes in the brain. Anti-GFAP autoantibody also can enter living astroglia cells in culture and its presence appears to compromise glial cell health. TBI patients showed an average 3.77 fold increase in anti-GFAP autoantibody levels from early (0–1 days) to late (7–10 days) times post injury. Changes in autoantibody levels were negatively correlated with outcome as measured by GOS-E score at 6 months, suggesting that TBI patients with greater anti-GFAP immune-responses had worse outcomes. Due to the long lasting nature of IgG, a test to detect anti-GFAP autoantibodies is likely to prolong the temporal window for assessment of brain damage in human patients.