Itamar Ronen1, Francesca Branzoli2, Alessandro Colasanti3, Iris Asllani3, Riccardo De Marco3, Neil Harrison4, and Mara Cercignani3
1Department of Radiology, Leiden University Medical Center, Leiden, Netherlands, 2Centre for NeuroImaging Research - CENIR, Brain and Spine Institute - ICM, Paris, France, 3Department of Neuroscience, Brighton and Sussex Medical School, University of Sussex, Brighton, United Kingdom, 4Cardiff University Brain Research Imaging Centre (CUBRIC), School of Medicine, Cardiff University, Cardiff, United Kingdom
Synopsis
Neuroinflammation is a pathomechanism implicated in several neruological, neurodegenerative and psychiatric disorders and is expressed in activation of microglia, the resident brain macrophages. Currently the only neuroimaging method for detecting microglial activation is PET with translocator protein (TSPO) ligands. A well-known model of experimentally inducing systemic inflammation is the administration of lipopolysaccharides (LPS). Here we report a significant increase in the apparent diffusion coefficient of the glial metabolite choline (tCho) in the human thalamus, following injection of LPS. This finding suggests that the ADC(tCho) is a putative marker for glial activation and may be useful in measuring neuroinflammation in disease.
Introduction
Several neurological diseases and mental illnesses, notably schizophrenia,
major depression, and Alzheimer's disease, have been linked to inflammation and altered
immune function, and more specifically actions of ‘activated’ microglia,
the principal immune cell in the brain1. During systemic inflammation,
microglia undergo a temporary change in shape and function. This shift from a
‘resting’ to an ‘activated’ state is associated with release of pro-inflammatory
proteins that can disrupt the function of surrounding neurons2,3.
Translocator Protein (TSPO) PET is the current gold-standard for imaging activated
microglia in-vivo4.
However, it is hampered by poor specificity, expense and limited availability. Unlike PET, Diffusion-weighted (DW)-MRS5 does not rely on radiotracer administration
and has the advantage of being acquired with other routine MR images. DW-MRS measures the apparent
diffusion coefficient (ADC) of metabolites, thus providing cell-specific information
about cellular morphology. As such, DW-MRS is sensitive to cell-specific morphological
changes in disease, particularly in the central nervous system, where metabolites
are differentially expressed across cell types6. Of the most commonly
measured metabolites, N-Acetyl-aspartate (NAA) resides exclusively in neurons, total
creatine (tCr) in both neural and glial cells, while total choline (tCho) is
predominantly found in glia. It is therefore conceivable that ADC(tCho) is
sensitive to changes in glial morphology. In line with this hypothesis, an
increase in ADC(tCho) was recently shown to correlate with disease activity
scores in neuropsychiatric systemic lupus erythematosus, an inflammatory
autoimmune disease7.
Here we used a lipopolysaccharide (LPS) challenge, a widely used paradigm for
glial activation in vitro and in vivo but rarely in humans8,
to cause a peripheral immune response in healthy volunteers. We measured the
changes induced in metabolite ADC due to the LPS challenge to test the hypothesis that ADC(tCho) is
selectively affected as a result of microglial activation.Methods
Data were collected from 7 healthy male volunteers, 26±6 years of age.
All subjects received an intravenous injection of either LPS (1ng/Kg) or saline
(placebo control) in random order on two separate sessions. Serial blood
samples were taken to measure peripheral inflammatory markers, and leucocyte subsets
and the Profile of Mood States (POMS) to monitor mood change hourly until 6
hours after injection. During each session patients underwent a 60 minute DPA-714
TSPO PET scan followed by MRI. DW-MRS was acquired approximately 5-5½ hours
after injection. Data were collected from two 4.5cc volumes of interest (VOI) (Figure
1). One VOI was positioned in the thalamus, a brain structure that shows high
uptake levels of DPA-714 (Figure 2) and is susceptible to peripheral inflammation9,10
and the other in the corona radiata, white matter (low DPA-714 uptake) as a
control region. The DW-MRS sequence used was a bipolar sequence based on
sLASER. TE/TR=100ms/5s. Diffusion weighting conditions: one at b=0 s/mm2
and three spectra taken at b=3823 s/mm2 with gradients in three
orthogonal directions; NSA=32 for each condition. A short (NSA=4) scan without
water suppression was performed for eddy current correction. Individual
spectroscopic data were phase/frequency corrected and averaged per condition. Spectral
analyses were performed with linear prediction singular value decomposition
(LPSVD) analysis using Matlab (Mathworks, Natick MA, USA), and the peak area estimates
were used for ADC calculation. The spectra at b=0 were used to estimate relative
tCho and tNAA concentrations, expressed as the ratio between their peak area to
the tCr peak area. The ADC of the three metabolites as well as [tCho]/[tCr] and
[tNAA]/[tCr] were compared between sessions (LPS vs placebo) using a
paired-sample t-test. The association between changes in ADC(tCho) and changes
in mood (POMS) were assessed using Pearson’s correlation coefficient.Results
Figure 3 shows the mean metabolite ADC for the thalamus and WM VOIs,
for both sessions. Paired t-test revealed a significant increase in ADC(tCho)
in the thalamus for the LPS condition (p=0.008). No significant difference was
detected in ADC(tCho) in the WM
between the two conditions. ADC(tNAA) and ADC(tCr) did not differ significantly
between conditions in both VOIs. No significant differences in [tCho]/[tCr] or
in [tNAA]/[tCr] were observed in either VOI between conditions (Figure
4). LPS-induced changes in POMS total and negative mood were significantly
associated with changes in ADC(tCho) of the thalamus (Adj R2 = 0.78;
p = 0.005 and Adj R2 = 0.64; p = 0.019, respectively) – see Figure 5.Discussion and Conclusions
The significant increase in ADC(tCho) but not ADC(tNAA)
or ADC(tCr) suggests that peripheral LPS challenge selectively affects human microglial
and/or astrocytic cytomorphology. In contrast to effects in mice, LPS does not appears to activate
human astrocytes11, implying that the effect we
measured is rooted in microglial activation. In-vitro NMR measurements on
LPS-activated human microglia do not report a significant increase in tCho
concentration12.
Spectra shown in the same report suggest that [tCho] in microglia is
significantly higher than e.g. [tCr]. This may explain why despite the low total
microglial volume, microglial tCho may be responsible for the effects reported here.
These encouraging findings imply
that ADC(tCho) is a unique marker for microglial activation in humans. Future work will involve validation
through correlations of the DW-MRS data with TSPO-PET data we obtained from the
same subjects.Acknowledgements
We thank Gosia Marjanska and Eddie Auerbach from the Center for Magnetic Resonance Research, University of Minnesota, for generously letting us use their DW-MRS sequences on the Siemens platform.
We would like to thank Dr
Anthony Suffredini and the National Institutes of Health for making freely
available the Clinical Center Reference Endotoxin employed in this study.
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