Diffusion magnetic resonance imaging-derived free water detects neurodegenerative pattern induced by interferon-γ

Abstract

Imaging biomarkers for immune activation may be valuable for early-stage detection, therapeutic testing, and research on neurodegenerative conditions. In the present study, we determined whether diffusion magnetic resonance imaging-derived free water signal is a sensitive marker for neuroinflammatory effects of interferon-gamma (Ifn-γ). Neonatal wild-type mice were injected in the cerebral ventricles with recombinant adeno-associated viruses expressing the inflammatory cytokine Ifn-γ. Groups of mice expressing Ifn-γ and age-matched controls were imaged at 1, 5 and 8 months. Mice deficient in Ifngr1^-/- and Stat1^-/- were scanned at 5 months as controls for the signaling cascades activated by Ifn-γ. The results indicate that Ifn-γ affected fractional anisotropy (FA), mean diffusivity (MD), and free water (FW) in white matter structures, midline cortical areas, and medial thalamic areas. In these structures, FA and MD decreased progressively from 1 to 8 months of age, while FW increased significantly. The observed reductions in FA and MD and increased FW with elevated brain Ifn-γ was not observed in Ifngr1^-/- or Stat1^-/- mice. These results suggest that the observed microstructure changes involve the Ifn-gr1 and Stat1 signaling. Interestingly, increases in FW were observed in midbrain of Ifngr1^-/- mice, which suggests alternative Ifn-γ signaling in midbrain. Although initial evidence is offered in relation to the sensitivity of the FW signal to neurodegenerative and/or inflammatory patterns specific to Ifn-γ, further research is needed to determine applicability and specificity across animal models of neuroinflammatory and degenerative disorders.

Keywords: Aging; Diffusion MRI; Free water; Inflammation; Interferon gamma; White matter.

Figure 1.

Steps used in image preprocessing. A) Alignment of anatomical scans to a structural reference scan and segmented atlas. B) Comparison of fractional anisotropy images, before and after free water elimination. C) Shows FA maps with color encoded directionality (sphere with red green blue indicates direction). D) FA and E) mean diffusivity values (mm²/s) for grey matter (frontal cortex) and white matter (corpus callosum) of 8 control WT mice. Data presented as Tukey box and whisker plots (median, 25–75^th percentile and 1.5 interquartile range). **Significantly different from uncorrected values (Tukey’s post hoc test).

Figure 2.

Elevated Ifn-γ expression produces age-progressive increases in free water signal in brain regions with coincident reductions in fractional anisotropy and inflammation-induced lesions. A) Representative structural, fractional anisotropy and free water maps of control mice and mice overexpressing Ifn-γ, shown at different ages. B) Reduced nigral DARPP32 and Th immunostaining demonstrate age-progressive degeneration in the midbrain of Ifn-γ mice compared to control mice. Arrow in immunohistochemical panels denotes dystrophic calcinosis.

Figure 3.

Composite statistical maps indicate that Ifn-γ produced significant changes in fractional anisotropy (FA), free water (FW) and mean diffusivity (MD) in white matter and surrounding regions of the mouse brain. Shown are significant voxels in left (L) and right (R) hemispheres (arrows note significantly different voxels). All images are false discovery rate (FDR) corrected with a p value corresponding to a FDRq threshold ≤ 1–5%. Ifn-γ produced reductions in FA and MD and increases in FW relative to WT control mice.

Figure 4.

Ifn-γ reduced fractional anisotropy and mean diffusivity and increased free water in white matter, midline cortical and thalamic areas. Two factor ANOVA and Bonferroni post hoc multiple comparison test. *main effect of Ifn-γ treatment, ** main effect of age, and *** treatment × age interaction (Bonferroni corrected p<0.003; n=4–5 mice/group). Error bars are standard deviation. Symbols above bars are significantly different from: a =1 month control, b = 1 month Ifn-γ, c = 5month control, d = 5 month Ifn-γ, e = 8month control, f=8 month Ifn-γ. Abbreviations: AH, anterior hypothalamus; ATA, anterior thalamic area; CA1/3, cornus Ammonis 1/3; cc, corpus callosum; CPu, caudate-putamen; fi, fimbria; ic, internal capsule; MD, mediodorsal thalamus; MPA, medial preoptic area; MS, medial septum; PH, posterior hypothalamus; Po, posterior thalamus; PrL, prelimbic cortex; RSC/R, retrosplenial cortex – caudal/rostral; SN, substantia nigra; ventral tegmental area, VTA.

Figure 5.

Ifn-γ induced increases in free water, reduced fractional anisotropy, and inflammation induced lesions are not observed in Ifngr1 and Stat1 null mice.

Figure 6.

Ifn-γ induced changes in fractional anisotropy, mean diffusivity and free water were largely absent in Ifngr1^−/− and Stat1^−/− mice. Two factor ANOVA and Bonferroni post hoc multiple comparison test. *main effect of Ifn-γ treatment, ** main effect of age, and *** treatment × age interaction (Bonferroni corrected p<0.003; n=4–10 mice/group). Error bars are standard deviation. Symbols above bars are as in Figure 4. Abbreviations are as in Figure 4.