Traumatic brain injury



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http://stm.sciencemag.org/content/4/134/134ra60.full

Sci Transl Med 16 May 2012: 


Vol. 4, Issue 134, p. 134ra60 
Sci. Transl. Med. DOI: 10.1126/scitranslmed.3003716
TRAUMATIC BRAIN INJURY

Chronic Traumatic Encephalopathy in Blast-Exposed Military Veterans and a Blast Neurotrauma Mouse Model

  1. Lee E. Goldstein1,2,3,4,*, 

  2. Andrew M. Fisher1,4, 

  3. Chad A. Tagge1,4,

  4. Xiao-Lei Zhang5, 

  5. Libor Velisek5, 

  6. John A. Sullivan5, 

  7. Chirag Upreti5,

  8. Jonathan M. Kracht4, 

  9. Maria Ericsson6, 

  10. Mark W. Wojnarowicz1,

  11. Cezar J. Goletiani5, 

  12. Giorgi M. Maglakelidze5, 

  13. Noel Casey1,3,

  14. Juliet A. Moncaster1,3, 

  15. Olga Minaeva1,3,4, 

  16. Robert D. Moir7,

  17. Christopher J. Nowinski8, 

  18. Robert A. Stern2,8, 

  19. Robert C. Cantu8,9,

  20. James Geiling10, 

  21. Jan K. Blusztajn2, 

  22. Benjamin L. Wolozin2,

  23. Tsuneya Ikezu2, 

  24. Thor D. Stein2,11, 

  25. Andrew E. Budson2,11,

  26. Neil W. Kowall2,11, 

  27. David Chargin12, 

  28. Andre Sharon4,12,

  29. Sudad Saman13, 

  30. Garth F. Hall13, 

  31. William C. Moss14,

  32. Robin O. Cleveland15, 

  33. Rudolph E. Tanzi7, 

  34. Patric K. Stanton5 and

  35. Ann C. McKee2,8,11,*

+Author Affiliations

  1. 1Molecular Aging and Development Laboratory, Boston University School of Medicine, Boston, MA 02118, USA.

  2. 2Boston University Alzheimer’s Disease Center, Boston, MA 02118, USA.

  3. 3Boston University Photonics Center, Boston University, Boston, MA 02215, USA.

  4. 4College of Engineering, Boston University, Boston, MA 02215, USA.

  5. 5Department of Cell Biology and Anatomy, New York Medical College, Valhalla, NY 10595, USA.

  6. 6Electron Microscope Facility, Harvard Medical School, Boston, MA 02115, USA.

  7. 7Genetics and Aging Research Unit, Massachusetts General Hospital, Charlestown, MA 02129, USA.

  8. 8Center for Study of Traumatic Encephalopathy, Boston University School of Medicine, Boston, MA 02118, USA.

  9. 9Department of Neurosurgery, Emerson Hospital, Concord, MA 01742, USA.

  10. 10Department of Medicine, Veterans Affairs Medical Center, White River Junction, VT 05009, USA.

  11. 11Neurology Service, Veterans Affairs Boston Healthcare System, Boston, MA 02130, USA.

  12. 12Fraunhofer Center for Manufacturing Innovation at Boston University, Brookline, MA 02446, USA.

  13. 13Department of Biological Sciences, University of Massachusetts Lowell, Lowell, MA 01854, USA.

  14. 14Lawrence Livermore National Laboratory, Livermore, CA 94551, USA.

  15. 15Institute of Biomedical Engineering, Department of Engineering Science, University of Oxford, Old Road Campus Research Building, Oxford OX3 7DQ, UK.

  1. *To whom correspondence should be addressed. E-mail: lgold@bu.edu(L.E.G.); amckee@bu.edu (A.C.M.)

Blast exposure is associated with traumatic brain injury (TBI), neuropsychiatric symptoms, and long-term cognitive disability. We examined a case series of postmortem brains from U.S. military veterans exposed to blast and/or concussive injury. We found evidence of chronic traumatic encephalopathy (CTE), a tau protein–linked neurodegenerative disease, that was similar to the CTE neuropathology observed in young amateur American football players and a professional wrestler with histories of concussive injuries. We developed a blast neurotrauma mouse model that recapitulated CTE-linked neuropathology in wild-type C57BL/6 mice 2 weeks after exposure to a single blast. Blast-exposed mice demonstrated phosphorylated tauopathy, myelinated axonopathy, microvasculopathy, chronic neuroinflammation, and neurodegeneration in the absence of macroscopic tissue damage or hemorrhage. Blast exposure induced persistent hippocampal-dependent learning and memory deficits that persisted for at least 1 month and correlated with impaired axonal conduction and defective activity-dependent long-term potentiation of synaptic transmission. Intracerebral pressure recordings demonstrated that shock waves traversed the mouse brain with minimal change and without thoracic contributions. Kinematic analysis revealed blast-induced head oscillation at accelerations sufficient to cause brain injury. Head immobilization during blast exposure prevented blast-induced learning and memory deficits. The contribution of blast wind to injurious head acceleration may be a primary injury mechanism leading to blast-related TBI and CTE. These results identify common pathogenic determinants leading to CTE in blast-exposed military veterans and head-injured athletes and additionally provide mechanistic evidence linking blast exposure to persistent impairments in neurophysiological function, learning, and memory.

IntroductionBack to Top

Blast exposure from conventional and improvised explosive devices (IEDs) affects combatants and civilians in conflict regions around the world (14). Individuals exposed to explosive blast are at increased risk for traumatic brain injury (TBI) (2515) that is often reported as mild (16 cf., 17). Blast-related TBI represents a neuropsychiatric spectrum disorder that clinically overlaps with chronic traumatic encephalopathy (CTE), a progressive tau protein–linked neurodegenerative disease associated with repetitive concussive injury in athletes (1821). Neuropathological hallmarks of CTE include widespread cortical foci of perivascular tau pathology, disseminated microgliosis and astrocytosis, myelinated axonopathy, and progressive neurodegeneration. Clinical symptoms of CTE include progressive affective lability, irritability, distractability, executive dysfunction, memory disturbances, suicidal ideation, and in advanced cases, cognitive deficits and dementia.



Blast exposure is a known precipitant of brain injury in animals (2237) and humans (3842) and has been linked to CTE neuropathology in a single case report by Omalu et al. (43). Despite growing awareness of blast-related TBI, the mechanisms of injury and biological basis underpinning blast neurotrauma and sequelae remain largely unknown and a matter of significant controversy. Given the overlap of clinical signs and symptoms in military personnel with blast-related TBI and athletes with concussion-related CTE, we hypothesized that common biomechanical and pathophysiological determinants may trigger development of CTE neuropathology and sequelae in both trauma settings. Here, we combine clinicopathological correlation analysis and controlled animal modeling studies to test this hypothesis.

ResultsBack to Top

CTE neuropathology in blast-exposed military veterans and athletes with repetitive concussive injury

We performed comprehensive neuropathological analyses (table S1) of postmortem brains obtained from a case series of military veterans with known blast exposure and/or concussive injury (n = 4 males; ages 22 to 45 years; mean, 32.3 years). We compared these neuropathological analyses to those of brains from young amateur American football players and a professional wrestler with histories of repetitive concussive injury (n = 4 males; ages 17 to 27 years; mean, 20.8 years) and brains from normal controls of comparable ages without a history of blast exposure, concussive injury, or neurological disease (n = 4 males; ages 18 to 24 years; mean, 20.5 years). Case 1, a 45-year-old male U.S. military veteran with a single close-range IED blast exposure, experienced a state of disorientation without loss of consciousness that persisted for ~30 min after blast exposure. He subsequently developed headaches, irritability, difficulty sleeping and concentrating, and depression that continued until his death 2 years later from a ruptured basilar aneurysm. His medical history is notable for a remote history of concussion associated with a motor vehicle accident at age 8 years. Case 2, a 34-year-old male U.S. military veteran without a history of previous concussive injury, sustained two separate IED blast exposures 1 and 6 years before death. Both episodes resulted in loss of consciousness of indeterminate duration. He subsequently developed depression, short-term memory loss, word-finding difficulties, decreased concentration and attention, sleep disturbances, and executive function impairments. His neuropsychiatric symptoms persisted until death from aspiration pneumonia after ingestion of prescription analgesics. Case 3, a 22-year-old male U.S. military veteran with a single close-range IED blast exposure 2 years before death. He did not lose consciousness, but reported headache, dizziness, and fatigue that persisted for 24 hours after the blast. He subsequently developed daily headaches, memory loss, depression, and decreased attention and concentration. In the year before his death, he became increasingly violent and verbally abusive with frequent outbursts of anger and aggression. He was diagnosed with posttraumatic stress disorder (PTSD) 3 months before death from an intracerebral hemorrhage. His past history included 2 years of high school football and multiple concussions from fist fights. Case 4, a 28-year-old male U.S. military veteran with two combat deployments, was diagnosed with PTSD after his first deployment 3 years before death. His history was notable for multiple concussions as a civilian and in combat, but he was never exposed to blast. His first concussion occurred at age 12 after a bicycle accident with temporary loss of consciousness and pre/posttraumatic amnesia. At age 17, he experienced a concussion without loss of consciousness from helmet-to-helmet impact injury during football practice. At age 25, he sustained a third concussion during military deployment with temporary alteration in mental status without loss of consciousness. Four months later at age 26, he sustained a fourth concussion with temporary loss of consciousness and posttraumatic amnesia resulting from a motor vehicle–bicycle collision. Afterward, he experienced persistent anxiety, difficulty concentrating, word-finding difficulties, learning and memory impairment, reduced psychomotor speed, and exacerbation of PTSD symptoms. He died from a self-inflicted gunshot wound 2 years after his last concussion. The athlete group included Case 5, a 17-year-old male high school American football player who died from second impact syndrome 2 weeks after sustaining a concussion; Case 6, an 18-year-old high school American football and rugby player with a history of three to four previous concussions, one requiring hospitalization, who died 10 days after his last concussion; Case 7, a 21-year-old male college American football player, who played as a lineman and linebacker but had never been diagnosed with a concussion during his 13 seasons of play beginning at age 9, and who died from suicide; and Case 8, a 27-year-old male professional wrestler who experienced more than 9 concussions during his 10-year professional wrestling career who died from an overdose of OxyContin. The normal control group included Case 9, an 18-year-old male who died suddenly from a ruptured basilar aneurysm; Case 10, a 19-year-old male who died from a cardiac arrhythmia; Case 11, a 21-year-old male who died from suicide; and Case 12, a 24-year-old male who died from suicide.

Neuropathological analysis of postmortem brains from military veterans with blast exposure and/or concussive injury revealed CTE-linked neuropathology characterized by perivascular foci of tau-immunoreactive neurofibrillary tangles (NFTs) and glial tangles in the inferior frontal, dorsolateral frontal, parietal, and temporal cortices with predilection for sulcal depths (Fig. 1, A, B, E, F, and I to X). NFTs and dystrophic axons immunoreactive for monoclonal antibody CP-13 (Fig. 1, A to I, L, Q, R, and U, and fig. S1) directed against phosphorylated tau protein at Ser202 (pS202) and Thr205 (pT205), monoclonal antibody AT8 (Fig. 1S) directed against phosphorylated tau protein at Ser202 (pS202) and Thr205 (pT205), and monoclonal antibody Tau-46 (Fig. 1T) directed against phosphorylation-independent tau protein were detected in superficial layers of frontal and parietal cortex and anterior hippocampus. Evidence of axon degeneration, axon retraction bulbs, and axonal dystrophy were observed in the subcortical white matter subjacent to cortical tau pathology (Fig. 1, M and U to X, and fig. S1). Distorted axons and axon retraction bulbs were prominent in perivascular areas. Large clusters of LN3-immunoreactive activated microglia clusters (Fig. 1, K and P) were observed in subcortical white matter underlying focal tau pathology, but not in unaffected brain regions distant from tau lesions. Neuropathological comparison to brains from young-adult amateur American football players (Fig. 1, C, D, G, and H) with histories of repetitive concussive and subconcussive injury exhibited similar CTE neuropathology marked by perivascular NFTs and glial tangles with sulcal depth prominence in the dorsolateral and inferior frontal cortices. The young-adult athlete brains also revealed evidence of robust astrocytosis and multifocal axonopathy in subcortical white matter. Clusters of activated perivascular microglia were noted in the subcortical U-fibers. Neuropathological findings in the military veterans with blast exposure and/or concussive injury and young-adult athletes with repetitive concussive injury were consistent with our previous CTE case studies (2021) and could be readily differentiated from neuropathology associated with Alzheimer’s disease, frontotemporal dementia, and other age-related neurodegenerative disorders. Control sections omitting primary antibody demonstrated no immunoreactivity. By contrast, none of the brains from the four young-adult normal control subjects demonstrated phosphorylated tau pathology, axonal injury, subcortical astrocytosis, or microglial nodules indicative of CTE or other neurodegenerative disease (fig. S2).



fig. 1

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Fig. 1

CTE neuropathology in postmortem brains from military veterans with blast exposure and/or concussive injury and young athletes with repetitive concussive injury. (A and E) Case 1, phosphorylated tau (CP-13) neuropathology with perivascular neurofibrillary degeneration in the frontal cortex of a 45-year-old male military veteran with a history of single close-range blast exposure 2 years before death and a remote history of concussion. Whole-mount section. Scale bar (E), 100 μm. (B and F) Case 2, phosphorylated tau (CP-13) neuropathology with perivascular neurofibrillary degeneration in the frontal cortex of a 34-year-old male military veteran with history of two blast exposures 1 and 6 years before death and without a history of concussion. Whole-mount section. Scale bar (F), 100 μm. (C and G) Case 6, phosphorylated tau (CP-13) neuropathology with perivascular neurofibrillary degeneration in the frontal cortex of an 18-year-old male amateur American football player with a history of repetitive concussive injury. Whole-mount section. Scale bar (G), 100 μm. (D and H) Case 7, phosphorylated tau (CP-13) neuropathology with perivascular neurofibrillary degeneration in the frontal cortex of a 21-year-old male amateur American football player with a history of repetitive subconcussive injury. Whole-mount section. Scale bar (H), 100 μm. (I) Case 1, phosphorylated tau (CP-13) immunostaining in the parietal cortex revealed a string of perivascular foci demonstrating intense immunoreactivity (areas enclosed by hash lines). Whole-mount section. (J) Case 1, phosphorylated neurofilament (SMI-34) immunostaining in adjacent parietal cortex section demonstrating colocalization of multifocal axonal swellings and axonal retraction bulbs surrounding small blood vessels (black circles) relative to perivascular tau foci (areas enclosed by hash lines). Whole-mount section. (K) Case 1, human leukocyte antigen–DR (HLA-DR) (LN3) immunostaining in adjacent parietal cortex section demonstrating colocalization of microglial clusters (black circles) relative to perivascular tau foci (areas enclosed by hash lines). Whole-mount section. (L) Case 1, high-magnification micrograph of phosphorylated tau (CP-13) immunostaining in the parietal cortex demonstrating string of perivascular phosphorylated tau foci. Whole-mount section. (M) Case 1, phosphorylated tau (PHF-1, brown) and phosphorylated neurofilament (SMI-34, red) double immunostaining in parietal cortex demonstrating axonal swellings and a retraction bulb (arrow) in continuity with phosphorylated tau neuritic abnormalities. Whole-mount section. Scale bar, 100 μm. (N) Case 1, phosphorylated neurofilament (SMI-34) immunostaining showing diffuse axonal degeneration and multifocal irregular axonal swellings in subcortical white matter subjacent to cortical tau pathology. Whole-mount section. (O) Case 1, phosphorylated neurofilament (SMI-34) immunostaining demonstrating perivascular axonal pathology and axonal retraction bulbs near a small cortical blood vessel. Whole-mount section. (P) Case 1, activated microglia (LN3) immunostaining showing a large microglial nodule in the subcortical white matter subjacent to cortical tau pathology. LN3 immunostaining was not observed in brain areas devoid of tau pathology. Whole-mount section. Scale bar, 100 μm. (Q) Case 2, phosphorylated tau (CP-13) immunostaining showing diffuse neuronal tau pathology (pre-tangles) in the hippocampal CA1 field. Whole-mount section. (R) Case 2, phosphorylated tau (CP-13) pathology in temporal cortex. Whole-mount section. (S) Case 1, phosphorylated tau (AT8) immunostaining showing diffuse neuronal tau pathology (pre-tangles) in the hippocampal CA1 field. Whole-mount section. (T) Case 1, phosphorylation-independent total tau (Tau-46) immunostaining in the frontal cortex. Whole-mount section. (U) Case 3, phosphorylated tau (CP-13) immunostained axonal varicosities in the external capsule of a 22-year-old male military veteran with a history of a single close-range IED blast exposure and remote history of concussions. Whole-mount section. (Vto X) Case 3, SMI-34 immunostained axonal varicosities and retraction bulbs in the thalamic fasiculus and external capsule. Whole-mount sections.

Blast exposure induces traumatic head acceleration in a blast neurotrauma mouse model

We developed a murine blast neurotrauma model to investigate mechanistic linkage between blast exposure, CTE neuropathology, and neurobehavioral sequelae. Our compressed gas blast tube was designed to accommodate mice (fig. S3) and allowed free movement of the head and cervical spine to model typical conditions associated with military blast exposure (tables S2 and S3 and figs. S3 to S7). Wild-type C57BL/6 male mice (2.5 months) were anesthetized and exposed to a single blast with a static (incident) pressure profile comparable in amplitude, waveform shape, and impulse to detonation of 5.8 kg of trinitrotoluene (TNT) at a standoff distance of 5.5 m and in close agreement with ConWep (Conventional Weapons Effects Program) (Fig. 2A and table S3) (44). The model blast is comparable to a common IED fabricated from a 120-mm artillery round and is within the reported range of typical explosives, blast conditions, and standoff distances associated with military blast injury (45).



fig. 2

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Fig. 2

Free-field pressure (FFP) and intracranial pressure (ICP) dynamics and head kinematics during single-blast exposure in a blast neurotrauma mouse model. (A) Measured incident static blast pressure (blue line) and blast impulse (red line) are compared to equivalent explosive blast waveform expected from 5.8 kg of TNT at a standoff distance of 5.5 m (black line) calculated according to software analysis using ConWep (44). The positive phase terminates at 4.8 ms (t+ = 4.8 ms; black hash line). Blast characteristics and waveform structure are comparable to a typical IED fabricated from a 120-mm artillery round (4.53 kg of TNT equivalent charge weight). The measured blast waveform and equivalent TNT blast waveform are in close agreement with a leading shock wavefront followed by a smooth decay. Note that ConWep presents an idealized blast resulting from an above-ground spherical charge and does not model negative-phase pressure transients or modulating factors commonly encountered in military blast scenarios. Reflecting surfaces, bounding structures (for example, crew compartments in armored vehicles, rooms within buildings, walled streets, and alleyways), local geometry, device and deployment characteristics (for example, encapsulation, internal reflectors, and open versus buried deployment), ambient environmental conditions, and other factors strongly influence blast pressure amplitude (positive and negative), phase duration, impulse history, waveform structure, and target interactions (308486). (B and C) ICP waveform and impulse profile in the brain of an intact living mouse (B) and isolated mouse head severed at the cervical spine (C) subjected to the same blast conditions as in (A). Blast waveforms recorded in the brains of living mice (B) and isolated heads (C) were similar in amplitude to each other and to the measured free-field static pressure. Small differences in the ICP signal waveforms were within the expected range given differences in frequency-dependent transducer response characteristics and experimental preparations. (D) Kinetographic representation of projected Cartesian motion of a representative mouse head during blast exposure as determined by high-speed videography acquired at 100,000 frames per second. Cartesian motion of the head was calculated by tracking a reflective paint mark on the snout. Labeled time points identify corresponding time points in (A) and (E) to (G). (E to G) Relative position (E), angular velocity (F), and angular acceleration (G) of the mouse head referenced to the horizontal (blue) and sagittal (red) planes of motion as determined by analysis of high-speed videographic records obtained during blast exposure. Head acceleration was most significant during the positive phase of the blast shock wave.

To investigate intracranial pressure (ICP) dynamics during blast exposure, we inserted a needle hydrophone into the hippocampus of living mice and monitored pressure dynamics during blast exposure. We detected blast wavefront arrival times in the brain that were indistinguishable from corresponding free-field pressure (FFP) measurements in air (Fig. 2B) and in close agreement with ConWep analysis of an equivalent TNT blast (Fig. 2Aand table S3). To investigate possible thoracic contributions to blast-induced ICP transients, we evaluated pressure tracings in the hippocampus of intact living mice (Fig. 2B) and compared results to the same measurements obtained in isolated mouse heads severed at the cervical spine (Fig. 2C). Blast-induced pressure amplitudes in the two experimental preparations were comparable to each other and to the corresponding FFP measurements in air, after accounting for the addition of the dynamic pressure on the head. Small differences in the pressure waveforms were within the expected range given frequency-dependent response characteristics of the transducers and differences in the two experimental preparations. We did not detect delayed blast-induced ICP transients in either preparation over recording times up to 100 ms. These observations indicate that blast wavefront transmission in the mouse brain is mediated without significant contributions from thoracovascular or hydrodynamic mechanisms.

In our experimental system, the blast shock wave traveling at ~450 m/s encountered the left lateral surface of the mouse head first, then traversed the ~11-mm skull width (46) in ~24 μs. The pressure differential associated with this traversal has an insignificant effect on skull displacement due to the short time interval. For the remainder of the waveform duration, the static pressures at the lateral surfaces of the skull are virtually identical and the corresponding transient effects are negligible. The air-skull impedance mismatch creates a back-reflected air shock as well as a rapidly moving (≥1500 m/s) transmitted shock wave, the latter taking a maximum of ~7 μs to traverse the cranium and cranial contents. Although the reflected and transmitted shock waves are large (~2.5 times greater than the 77-kPa incident overpressure), the ~7-μs traversal time of the skull-brain transmitted wave is short enough to allow rapid equilibration across the skull. Thus, the head acts acoustically as a “lumped element” (4748). The only significant pressure term remaining is the ~19-kPa peak dynamic pressure generated by blast wind. We concluded that an ICP transducer in the brain parenchyma should measure pressure differentials that do not differ by more than 19 kPa from FFP values, at least beyond the initial 30 μs after blast arrival. This analysis was confirmed by experimental measurements (Fig. 2B). Only the initial rise of the blast wave has a short enough time scale to be affected by propagation effects in the head, a prediction confirmed by the longer rise time of the ICP compared to the static FFP waveforms (Fig. 2, B and C). The remaining waveform components evenly distribute through the brain with amplitude and shape that approximate the static FFP (Fig. 2A).

The blast wave had a measured Mach number of 1.26 ± 0.04 (fig. S6), from which the calculated blast wind velocity was 150 m/s (336 miles/hour). Kinematic analysis of high-speed videographic records of head movement during blast exposure confirmed rapid oscillating acceleration-deceleration of the head in the horizontal and sagittal planes of motion (Fig. 2, D to G, and video S1). We calculated peak average radial head acceleration of 954 ± 215 krad/s2 (Fig. 2G), corresponding to 100.2 N exerted on the head during blast exposure. Peak angular and centripetal acceleration were most significant during the positive phase of the blast shock wave. No appreciable head acceleration was detected after ~8 ms.

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