Traumatic microbleeds suggest vascular injury and predict disability in traumatic brain injury

Abstract

Traumatic microbleeds are small foci of hypointensity seen on T2*-weighted MRI in patients following head trauma that have previously been considered a marker of axonal injury. The linear appearance and location of some traumatic microbleeds suggests a vascular origin. The aims of this study were to: (i) identify and characterize traumatic microbleeds in patients with acute traumatic brain injury; (ii) determine whether appearance of traumatic microbleeds predict clinical outcome; and (iii) describe the pathology underlying traumatic microbleeds in an index patient. Patients presenting to the emergency department following acute head trauma who received a head CT were enrolled within 48 h of injury and received a research MRI. Disability was defined using Glasgow Outcome Scale-Extended ≤6 at follow-up. All magnetic resonance images were interpreted prospectively and were used for subsequent analysis of traumatic microbleeds. Lesions on T2* MRI were stratified based on 'linear' streak-like or 'punctate' petechial-appearing traumatic microbleeds. The brain of an enrolled subject imaged acutely was procured following death for evaluation of traumatic microbleeds using MRI targeted pathology methods. Of the 439 patients enrolled over 78 months, 31% (134/439) had evidence of punctate and/or linear traumatic microbleeds on MRI. Severity of injury, mechanism of injury, and CT findings were associated with traumatic microbleeds on MRI. The presence of traumatic microbleeds was an independent predictor of disability (P < 0.05; odds ratio = 2.5). No differences were found between patients with punctate versus linear appearing microbleeds. Post-mortem imaging and histology revealed traumatic microbleed co-localization with iron-laden macrophages, predominately seen in perivascular space. Evidence of axonal injury was not observed in co-localized histopathological sections. Traumatic microbleeds were prevalent in the population studied and predictive of worse outcome. The source of traumatic microbleed signal on MRI appeared to be iron-laden macrophages in the perivascular space tracking a network of injured vessels. While axonal injury in association with traumatic microbleeds cannot be excluded, recognizing traumatic microbleeds as a form of traumatic vascular injury may aid in identifying patients who could benefit from new therapies targeting the injured vasculature and secondary injury to parenchyma.

Keywords: MRI biomarkers of traumatic brain injury; mild traumatic brain injury; radiological-pathological analysis; traumatic microbleeds; traumatic vascular injury.

Figure 1

Characterization of TMBs in the clinical population. T₂*-weighted 3 T MRI of individual subject's brain to illustrate presence of TMBs. Coronal (A) and sagittal (B) views of representative punctate TMB shown in axial plane (C). Coronal (E) and sagittal (F) views of linear TMB shown in axial plane (D).

Figure 2

Imaging comparison of mild TBI patient and index patient. (A) Acute CT of index patient reveals subdural haemorrhage on patient’s right side, showing that the index patient was not of mild severity. (B–F) Acute 3 T MRI shows pattern of TMBs similar to that of mild TBI patient. (G) Acute CT of mild TBI patient in THINC study is unremarkable, showing no extra-axial blood or injury to the parenchyma. Hyperintensity viewed on diffusion-weighted imaging and corresponding hypointensity observed on apparent diffusion coefficient (ADC) of both index (B and C) and mild TBI patients (H and I) shows similar patterns of cytotoxic oedema in tissue surrounding TMBs. Pattern of hyperintense signal observed on FLAIR on both the index (D) and mild (J) TBI patients suggest acute vasogenic oedema around the same areas that we see TMBs. Punctate and curvilinear patterns of hypointensities observed on axial gradient recalled echo (GRE) images of index (E) and mild (K) TMBs have a linear shape on coronal view in both patients (F and L). Yellow arrows indicate exact location of each TMB in coronal view. Although not mild, the index patient has similar patterns of linear TMBs as observed in mild TBI patients.

Figure 3

Evolution of TMBs in index patient. (A) Acute T₂*-weighted 3 T MRI obtained within 48 h of injury shows pattern of TMBs (within yellow oval) consistent with T₂*-weighted 3 T MRI obtained 3 months post injury (B), the last MRI obtained. The patient died ∼7 months post-injury. (C) Post-mortem T₂*-weighted 7 T MRI showed TMBs remained stable from acute in vivo MRI to post-mortem MRI, with some morphology changes from brain extraction. Pattern of linear TMBs observed on post-mortem coronal (D) and axial (E) images of the whole brain. Region targeted for histopathological analysis outlined in yellow (E). (F) High-resolution small-bore 7 T MRI of targeted region to confirm TMB imaging within tissue prior to embedding region of interest for histological co-localization of TMB.

Figure 4

MRI guided pathology shows evidence of injury to the vasculature. (A) 7 T small-bore 3D T₂*-weighted MRI scan of region with visible TMBs within the index case. Red line indicates outline of the region that was included within the tissue block for 20-μm thick histology sections. (B–D) Perls Prussian Blue stain shows iron deposits in haemosiderin-laden macrophages. Solid line box (B) includes the grey-white matter junction and indicates the enlarged region shown in C. Dotted line box (C) indicates enlarged region in D showing cellular detail. (E–G) Nissl cytological stain. Solid line box (E) indicates enlarged region shown in F. Dotted line box (F) indicates enlarged region in G showing a linear pattern of high cell density. (H–J) Gallyas silver stain for myelin. Solid line box (H) indicates enlarged section shown in I. Grey dotted line box (H) shows grey-white matter border and indicates enlarged section in I. Within the white matter, myelin is absent from a linear region (I). In the adjacent area, myelin is slightly reduced (J) and cells with iron deposits are present (D). Scale bars in B, E and H = 3 mm; C and F = 500 μm; D and I = 100 μm; G = 50 μm; J = 30 μm.

Figure 5

Pathology underlying TMBs: injury to the vasculature. (A) High-resolution 7 T MRI of tissue block containing region of interest co-localized to TMBs observed in vivo and ex vivo whole-brain MRI. (B) Standard 2D histopathology section (20-μm thick) from the imaged tissue block (A) stained with Perls Prussian Blue reveals iron in macrophages surrounding a large vessel. (C) Serially cut tissue sections digitized as a 3D volume, shown as side views in the adjacent panels. A 3D minimum intensity projection (MIP) image of the Perls stained histology reconstructed to match high-resolution 7 T MRI (A). (C) 3D reconstruction of interleaved tissue sections stained for iron deposits demonstrates iron laden macrophages that are connected and branching over centimetres of tissue. (D) Enlarged section of (C) showing iron-laden macrophages that co-localized with a vessel. These interleaved serial sections of the TMB in tri-planar view reveal a territory of injured vasculature in areas visible on MRI as punctate or linear hypointensities. Injury to the large vessel is indicated by confluent regions of iron-laden macrophages that create sufficient signal for detection on MRI. Injury to some of the other smaller vessels is only detected on post-mortem histology in a 3D reconstruction. The tri-planar view and high resolution identifies connectivity of the injured vessels ranging from large to small diameters.

Figure 6

Traumatic vascular injury. Traditionally, TMBs have been described as markers for diffuse axonal injury. However, TMBs have not been characterized as a form of traumatic vascular injury. MRI and 3D histology identified an underlying pathology of TMBs to be an injury to not just a single vessel, but injury that extends to a vascular network. Injury to a large blood vessel causes iron deposition that is detectable on in vivo and ex vivo MRI (at 3 T and 7 T). Smaller vessels that form a broader vascular network with the larger vessels are missed in these magnetic resonance images. High-resolution 3D histology of iron deposits in macrophages identified more extensive vascular injury involving smaller vessels that connect to the larger injured vessel detected on MRI. The schematic above illustrates a model of this vascular injury pattern and potential levels of detection with MRI and histological approaches.