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Brain-enriched protein biomarkers of tissue fate are being introduced clinically to aid in traumatic brain injury (TBI) management. The aim of this study was to determine how concentrations of six different protein biomarkers, measured in samples collected during the first weeks after TBI, relate to injury severity and outcome. We included neurocritical care TBI patients that were prospectively enrolled from 2007 to 2013, all having one to three blood samples drawn during the first 2 weeks. The biomarkers analyzed were S100 calcium-binding protein B (S100B), neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP), ubiquitin carboxy-terminal hydrolase-L1 (UCH-L1), tau, and neurofilament-light (NF-L). Glasgow Outcome Score (GOS) was assessed at 12 months. In total, 172 patients were included. All serum markers were associated with injury severity as classified on computed tomography scans at admission. Almost all biomarkers outperformed other known outcome predictors with higher levels the first 5 days, correlating with unfavorable outcomes, and UCH-L1 (0.260, pseduo-R2) displaying the best discrimination in univariate analyses. After adjusting for acknowledged TBI outcome predictors, GFAP and NF-L added most independent information to predict favorable/unfavorable GOS, improving the model from 0.38 to 0.51 pseudo-R2. A correlation matrix indicated substantial covariance, with the strongest correlation between UCH-L1, GFAP, and tau (r = 0.827–0.880). Additionally, the principal component analysis exhibited clustering of UCH-L1 and tau, as well as GFAP, S100B, and NSE, which was separate from NF-L. In summary, a panel of several different protein biomarkers, all associated with injury severity, with different cellular origin and temporal trajectories, improve outcome prediction models.

The complement cascade has been suggested to be involved in the development of secondary brain injuries following brain contusions, based on animal experiments. The aim of the present study was to examine the possible involvement of the complement cascade following traumatic head injury in the human brain. Sixteen patients were included in this study, 12-77 years of age, treated at the neurointensive care unit for traumatic brain contusions. All of these patients were operated with frontal or temporal lobe resection due to intractable intracranial hypertension. The resected tissue was analyzed with regard to components related to complement activation. The time interval between accident and operation was 2-82 h. Brain tissue from three patients operated with hippocampectomy due to epilepsy, including temporal lobe resection, were used as controls. We found increased immunoreactivity for complement components C1q, C3b, and C3d and the membrane attack complex (MAC), C5b-9, in the immediate vicinity of neurons in the penumbra area of the contusion. These findings constitute histological evidence for activation of the complement cascade in the penumbra of cortical contusions in the human brain. Using in situ hybridization, we also found C3-mRNA in the penumbra, suggesting a local synthesis of complement. Furthermore, upregulation of the endogenous complement regulator clusterin was found in some neurons in the same area. We suggest that unknown compounds in the debris from injured neurons or myelin breakdown products trigger complement activation, including formation of C5b-9. Activated complement components may stimulate accumulation of inflammatory cells and formation of brain edema, as well as having membrane destructive effects by the end product MAC, thereby being mediators in the development of secondary brain damage.

Neuronal survival is important to functional restitution following axotomy. Proximal lesions of the facial nerve, due to head trauma or tumor growth, for example, may cause long-standing or even permanent facial nerve palsy. Betamethasone has been used by several neurosurgical clinics for the treatment of postoperative facial nerve palsy; however, this practice is based only on clinical experience. The aim of the present study was to explore the putative effect on facial motor neuron survival of a novel lazaroid (pyrrolopyrimidine, PNU-101033-E) and furthermore to compare the effects with those of betamethasone, following intracranial transection of the facial nerve in adult rats. Both agents are known to inhibit lipid peroxidation by free radical scavenging. The lesion model used has recently been reported to induce massive neuronal cell death with a relative survival of 26.8 +/- 11.3% 1 month after lesion. Oral administration of lazaroids or daily injections of betamethasone followed surgery for 1 month, after which quantification of motor neuronal profiles was performed in the facial nucleus. Lazaroid-treated animals showed a significantly enhanced neuronal survival (68.0 +/- 9.8%), whereas no significant difference was found in betamethasone-treated animals (33.1 +/- 11.7%). The microglial and astrocytic responses in the facial nucleus were intense on the operated sides in betamethasone-treated as well as lazaroid-treated animals, and no differences in comparison with untreated animals were found. In conclusion, we found that the novel pyrrolopyrimidine PNU-101033-E, but not betamethasone, significantly enhanced nerve cell survival. This agent may therefore serve as a useful neuroprotective agent following intracranial trauma to the facial nerve and should be further evaluated for clinical use.

The complement cascade has been suggested to be involved in development of secondary brain damage following traumatic brain injury (TBI). Previous studies have shown that reactive microglia are involved in activation of the complement cascade following various injuries to the nervous system. Macrophages seem to have a significant role in this process, but it is still unclear whether these cells, as well as the complement components, are derived from reactive microglia or if these biological events only can occur as a result from the influx of plasma and monocytes via a disrupted blood-brain barrier (BBB). The aim of this study was to investigate the response of microglial cells and the complement system in the absence of plasma/blood components following a standardized crush injury in an entorhinal-hippocampal slice culture. There was a clear increase in complement component C1q and C5b-9-IR (Membrane Attack Complex, MAC) in the area near the crush injury. MAC-IR appeared as numerous dots in clusters which co-localized with anti-NeuN labelled neurons in the injury border zone. Complement C1q-IR co-localized with reactive microglia, co-labelled with OX42 antisera. These findings show activation of the complement cascade near the injury zone and in particular, formation of MAC at the surface of neurons in this area. There was a distinct activation of microglial cells (OX42-IR) near the site of injury, as well as an increase in ED-1 expressing macrophages. In the absence of blood and plasma components it is likely that ED-1-labelled cells represent reactive microglia transformed into macrophages. In addition, Neurons (Neun-IR) near the injury were found to co-localize with clusterin-IR indicating upregulation of a defense system to the endogenous complement attack. The present study provides evidence that microglia and complement is activated in the injury border zone of the tissue slice in a similar fashion as in vivo following TBI, despite the absence of plasma/blood products and cells. These findings support the hypothesis that reactive microglia have a key role in complement activation following TBI by local synthesis of complement with a potential impact on development of secondary neuronal insults.

Activation of the complement cascade following peripheral nerve axotomy and following traumatic brain injury has been demonstrated in previous studies. This study investigates the temporal pattern of microglia/macrophages and complement activation following axotomy of sensory CNS neurons, using a standardized experimental crush injury of the optic nerve in adult rats. Numerous ED1-labeled macrophages were found at the lesion site and distal to the injury at 7 days post injury (dpi). Complement C3-mRNA was upregulated 2-28 days post lesion, indicating local synthesis of complement in the optic nerve. Furthermore, increased immunoreactivity (IR) for the end product of the complement cascade, the membrane attack complex (MAC), was detected along disintegrating axons co-labeled with anti-neurofilament distal to the injury. Double-labeling for microglia show MAC-immunoreactivity expressed in their immediate vicinity, indicating a key role of microglia/macrophages in complement activation. The complement regulator Clusterin was upregulated in astrocytes at the lesion site as well as in the distal portion of the injured optic nerve, suggesting activation of a defense response to endogenous complement attack. A crush injury of the optic nerve leads to complement activation at the site of lesion and along the distal portion of the nerve, as well as upregulation of the complement inhibitor Clusterin at least in astrocytes. Reactive microglial cells seem to have a key role in complement activation as a local source of C3. We suggest that the balance between complement activation and their regulators may have impact on axonal degeneration following optic nerve injury.