The topography of cortical microbleeds in frontotemporal lobar degeneration: a post-mortem 7.0-tesla magnetic resonance study

Introduction

Frontotemporal lobar degeneration (FTLD) is a heterogeneous disorder with various clinical and histological subtypes [22]. FTLD is the second most common cause of presenile dementia with different genetic subtypes [20]. Despite the fact that most cases have a presenile onset, FTLD is not rare amongst elderly patients [3]. A recent neuropathological study showed that age together with vascular and Alzheimer-related co-pathology contributes to the morphological appearance of FTLD-tau [23]. In our recent neuropathological assessment of small cerebrovascular lesions, only a prevalence of white matter changes was observed in the FTLD brains, compared to age-matched controls [10]. These findings are in contrast to the high incidence of macro- and micro-infarcts, and hematomas and small cerebral bleeds observed in brains of patients with Alzheimer’s disease (AD), in particular in those with associated cerebral amyloid angiopathy (CAA) [6,7].
Cerebral microbleeds (CMBs) are frequently de­tected on T2*-weighted gradient-echo magnetic resonance imaging (MRI) of patients with small-vessel diseases [26]. They are also found in asymptomatic patients as well as in those with various degrees of cognitive impairment [5]. The number of CMBs detected during life depends on the MRI characteristics, such as pulse sequence, sequence parameters, spatial resolution, magnetic field strength and image post-processing [17]. 7.0-tesla T2*-weighted gradient-echo MRI is able to detect reliably even the smallest bleeds in the cerebral cortex (CoMBs) on post-mortem brain sections [8]. This technique allows to determine the topographic distribution and to quantify the number of CoMBs on a large number of brain sections [25]. In our previous study we have demonstrated a significant prevalence of CoMBs in the deep cortical layers of the frontal cortex in FTLD [9]. However this frontal predominance of CoMBs is also observed in AD brains with and without CAA and in controls [14].
The aim of the present post-mortem MRI study is to compare the prevalence of CoMBs in twenty-two different cortical gyri of pure FTLD versus controls in order to determine their eventual clinical impact.

Material and methods

Twenty-three patients, followed-up at the Lille University Hospital underwent an autopsy. The cohorts consisted of 11 patients with FTLD and 12 controls who had no clinical history of dementia or stroke. The vascular risk factors were registered from the clinical files. The median age at disease onset of the FTLD patients was 54 (interquartile range of 46-64) years. The clinical phenotype of the FTLD patients was the behavioral variant in 9 and semantic dementia in 2. Two patients had in addition amyo­trophic lateral sclerosis. None of them had on neuropathological examination Alzheimer features or some degree of amyloid angiopathy. The post- mortem diagnosis of FTLD was made according to the neuropathological diagnostic and the nosological criteria of the Consortium for FTLD [4]. The main histological subtypes were FTLD-Tau in 3, FTLD-TDP-A in 1, FTLD-TDP-B in 3, FTLD-TDP-C in 3 brains and FTLD-FUS in 1 brain.
Previously obtained informed consent of the pa­tients or from the nearest family allowed an autopsy for diagnostic and scientific purposes. The brain tissue samples were acquired from the Lille Neuro-Bank of the Lille University, federated to the Centre de Ressources Biologiques that acted as an institutional review board.
One fresh cerebral hemisphere was deeply frozen for biochemical examination. The remaining hemisphere, the brainstem and most of the cerebellum were fixed in formalin for 3 weeks.
The patients with FTLD were compared concerning the incidence of CoMBs to the control group.

Neuropathological examination

The diagnosis of FTLD was made according to a standard procedure examining samples from the primary motor cortex, the associated frontal, temporal and parietal cortex, the primary and secondary visual cortex, the cingulate gyrus, the basal nucleus of Meynert, the amygdaloid body, the hippocampus, basal ganglia, mesencephalon, pons, medulla and cerebellum. Slides from paraffin-embedded sections were immune-stained for protein tau, -amyloid, -synuclein, prion protein and TDP-43. FUS histochemistry was performed in Tau and TDP negative cases.
A quantitative evaluation of the cerebrovascular lesions was performed on a standard coronal section of a cerebral hemisphere, at the level of the mammillary body according to a previously described me­thod [6].

MRI examination

Six coronal sections of a cerebral hemisphere from each brain were submitted to MRI: one at the prefrontal level in front of the frontal horn, one of the frontal lobe at the level of the head of the caudate nucleus, a central one near the mammillary body, a post-central one, a parietal one at the level of the splenium corporis callosi and one at the level of the occipital lobe.
We used a 7.0-tesla MRI Bruker BioSpin SA with an issuer-receiver cylinder coil of 72 mm inner diameter (Ettlingen, Germany), according to a previously described method [8]. The brain sections, previously cleaned from formalin, were placed in a plastic box filled with salt-free water, the size of which did not allow significant tissue movements. Three MRI sequences were used: a positioning sequence, a T2 sequence and a T2* sequence. The positioning sequence allowed determination of the three-direc­tional position of the brain section inside the magnet. The thickness of the T2 images was 1 mm. The field of view was a 9 cm-square slide that was coded by a 256 matrix giving a voxel size of 0.352 × 0.352 × 1 mm. T2 weighted images were obtained by using RARE sequence (Rapid Acquisition with Relaxation Enhancement) with repetition time (TR), echo time (TE) and RARE factor of 2,500 ms, 33 ms and 8, respectively. The acquisition time of this sequence was 80 s The thickness of the T2* images was 0.20 mm. The field of view was also a 9 cm2. It was coded by a 512 matrix, giving a voxel size of 0.176 × 0.176 × 0.2 mm. The slice thickness corresponded to the upper part of the brain section. This sequence was a GRE sequence with a short TR of 60 ms and TE of 22 ms, a flip angle of 30° and number of excitation of 20. The acquisition time of the sequence was 10 minutes.
The total number and the location of CoMBs was determined in 22 gyri of each brain (Table I) by consensus evaluation by three observers (JDR, FA, ND) blinded to the neuropathological diagnosis and based on comparison of brain sections of an anatomical atlas [19]. The inter-rater reliability resulted in an interclass correlation coefficient of 0.79. The mean values of CoMBs in FTLD brains were compared to the controls.

Statistical analyses

Univariate comparisons of unpaired groups were performed with the Fisher’s exact test for categorical data. The non-parametric Mann-Whitney U-test was used to compare continuous variables. The significance level, two-tailed, was set at ≤ 0.01 for significant and ≤ 0.001 for highly significant. Values set at ≤ 0.05 and more than > 0.01 were considered as marginal significant and not included as relevant due to the relative small sample sizes.

Results

The FTLD and control groups did not show any statistical differences according to age, gender distribution and vascular risk factors. However, on the neuropathological examination, white matter changes were significantly increased in the FTLD group compared to the control group (p = 0.01), while only marginally increased for the CoMBs. The other cerebrovascular lesions were rare and not statistical different between both groups (Table II).
When comparing on MRI the mean values of CoMBs in the six coronal sections on T2* GRE MRI an anterior-posterior decreasing gradient was ob­served in the FTLD as well as in the AD and the control brains (Fig. 1).
The mean values of CMBs in FTLD brains were significantly higher in the frontal superior gyrus and in the insular cortex (p ≤ 0.001) (Fig. 2) and significantly in the frontal inferior and the temporal superior gyri (p ≤ 0.01) (Fig. 3), compared to the control brains (Table III).

Discussion

The present study confirms that associated cerebrovascular lesions are rare in FTLD due to tau, TAR DNA-binding protein 43 and FUS [24]. Although in brains of elderly patients suffering from neurodegenerative diseases multiple pathologies are usually present [2], our series consist of pure types of FTLD allowing to determine more exactly the topography of CoMBs related to this disease spectrum. Despite a similar global bleeding load in FTLD and in age-matched control brains [11], a similar anterior-posterior decreasing gradient of CoMBs is observed on the sequential hemispheric sections. This is also common to AD brains with or without CAA [14]. This similarity is probably related to the larger development of the frontal lobe compared to the parietal and occipital lobes [21].
However, the main findings in this study are that the CoMBs predominate in the superior and inferior frontal gyri, the superior temporal gyrus and the insular cortex, regions that are most affected by the neurodegenerative process [1].
The findings in FTLD share similarity with those observed in our post-mortem 7.0-tesla MRI study in progressive supranuclear palsy, in which small bleeds are mainly found around the dentate nucleus of the cerebellum and in the tegmentum pontis, where also the major neurodegenerative changes occur [12].
There is no evidence for increased angiogenesis and microglial activation in the neuropathologically most affected regions of different neurodegenerative diseases such as in Alzheimer, Parkinson, progressive supranuclear palsy, and incidental Lewy body disease. Such neo-angiogenic vessels could contribute to neuroinflammation and lead to disruption of the blood-brain barrier [15,16]. The prevalence of CoMBs and the white matter changes in FTLD brains should not be considered as the hallmark of cerebrovascular diseases. Also the increased iron accumulation observed in the basal ganglia of brains with FTLD should not be related to small-vessel ischemic disease [13] as recently proposed [18].
The fact that the CoMBs prevail in the regions with the most prominent neurodegenerative lesions indicates that blood-brain barrier impairments may interact with the severity of the neurodegeneration in FTLD and are secondary phenomena without impact on the clinical features of the disease.

Disclosure

Authors report no conflict of interest.

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