A Dedicated 21-Plex Proximity Extension Assay Panel for High-Sensitivity Protein Biomarker Detection Using Microdialysis in Severe Traumatic Brain Injury: The Next Step in Precision Medicine?

Cerebral protein profiling in traumatic brain injury (TBI) is needed to better comprehend secondary injury pathways. Cerebral microdialysis (CMD), in combination with the proximity extension assay (PEA) technique, has great potential in this field. By using PEA, we have previously screened >500 proteins from CMD samples collected from TBI patients. In this study, we customized a PEA panel prototype of 21 selected candidate protein biomarkers, involved in inflammation (13), neuroplasticity/-repair (six), and axonal injury (two). The aim was to study their temporal dynamics and relation to age, structural injury, and clinical outcome. Ten patients with severe TBI and CMD monitoring, who were treated in the Neurointensive Care Unit, Uppsala University Hospital, Sweden, were included. Hourly CMD samples were collected for up to 7 days after trauma and analyzed with the 21-plex PEA panel. Seventeen of the 21 proteins from the CMD sample analyses showed significantly different mean levels between days. Early peaks (within 48 h) were noted with interleukin (IL)-1ß, IL-6, IL-8, granulocyte colony-stimulating factor, transforming growth factor alpha, brevican, junctional adhesion molecule B, and neurocan. C-X-C motif chemokine ligand 10 peaked after 3 days. Late peaks (>5 days) were noted with interleukin-1 receptor antagonist (IL-1ra), monocyte chemoattractant protein (MCP)-2, MCP-3, urokinase-type plasminogen activator, Dickkopf-related protein 1, and DRAXIN. IL-8, neurofilament heavy chain, and TAU were biphasic. Age (above/below 22 years) interacted with the temporal dynamics of IL-6, IL-1ra, vascular endothelial growth factor, MCP-3, and TAU. There was no association between radiological injury (Marshall grade) or clinical outcome (Extended Glasgow Outcome Scale) with the protein expression pattern. The PEA method is a highly sensitive molecular tool for protein profiling from cerebral tissue in TBI. The novel TBI dedicated 21-plex panel showed marked regulation of proteins belonging to the inflammation, plasticity/repair, and axonal injury families. The method may enable important insights into complex injury processes on a molecular level that may be of value in future efforts to tailor pharmacological TBI trials to better address specific disease processes and optimize timing of treatments.

Monitoring of Protein Biomarkers of Inflammation in Human Traumatic Brain Injury Using Microdialysis and Proximity Extension Assay Technology in Neurointensive Care.

Traumatic brain injury (TBI) is followed by secondary injury mechanisms strongly involving neuroinflammation. To monitor the complex inflammatory cascade in human TBI, we used cerebral microdialysis (MD) and multiplex proximity extension assay (PEA) technology and simultaneously measured levels of 92 protein biomarkers of inflammation in MD samples every three hours for five days in 10 patients with severe TBI under neurointensive care. One µL MD samples were incubated with paired oligonucleotide-conjugated antibodies binding to each protein, allowing quantification by real-time quantitative polymerase chain reaction. Sixty-nine proteins were suitable for statistical analysis. We found five different patterns with either early (<48 h; e.g., CCL20, IL6, LIF, CCL3), mid (48-96 h; e.g., CCL19, CXCL5, CXCL10, MMP1), late (>96 h; e.g., CD40, MCP2, MCP3), biphasic peaks (e.g., CXCL1, CXCL5, IL8) or stable (e.g., CCL4, DNER, VEGFA)/low trends. High protein levels were observed for e.g., CXCL1, CXCL10, MCP1, MCP2, IL8, while e.g., CCL28 and MCP4 were detected at low levels. Several proteins (CCL8, -19, -20, -23, CXCL1, -5, -6, -9, -11, CST5, DNER, Flt3L, and SIRT2) have not been studied previously in human TBI. Cross-correlation analysis revealed that LIF and CXCL5 may play a central role in the inflammatory cascade. This study provides a unique data set with individual temporal trends for potential inflammatory biomarkers in patients with TBI. We conclude that the combination of MD and PEA is a powerful tool to map the complex inflammatory cascade in the injured human brain. The technique offers new possibilities of protein profiling of complex secondary injury pathways.

Scandinavian Multicenter Acute Subdural Hematoma (SMASH) Study: Study Protocol for a Multinational Population-Based Consecutive Cohort.

BACKGROUND: Traumatic acute subdural hematomas (ASDHs) are associated with high rate of morbidity and mortality, especially in elderly individuals. However, recent reports indicate that the morbidity and mortality rates might have improved. OBJECTIVE: To evaluate postoperative (30-d) mortality in younger vs elderly (≥70 yr) patients with ASDH. Comparing younger and elderly patients, the secondary objectives are morbidity patterns of care and 6 mo outcome according to Glasgow outcome scale (GOS). Finally, in patients with traumatic ASDH, we aim to provide prognostic variables. METHODS: This is a large-scale population-based Scandinavian study including all neurosurgical departments in Denmark and Sweden. All adult (≥18 yr) patients surgically treated between 2010 and 2014 for a traumatic ASDH in Denmark and Sweden will be included. Identification at clinicaltrials.gov is NCT03284190. EXPECTED OUTCOMES: We expect to provide data on potential differences between younger vs elderly patients in terms of mortality and morbidity. We hypothesize that elderly patients selected for surgery have a similar pattern of care as compared with younger patients. We will provide functional outcome in terms of GOS at 6 mo in younger vs elderly patients undergoing ASDH evacuation. Finally, clinical useful prognostic factors for favorable (GOS 4-5) vs unfavorable (GOS 1-3) will be identified. DISCUSSION: An improved understanding of the clinical outcome, treatment and resource allocation, clinical course, and the prognostic factors of traumatic ASDH will allow neurosurgeons to make better treatment decisions.