Gehirn-Magnetresonanz-Elastographie zur frühen Krankheitserkennung und Staging korreliert mit Histopathologie und Analyse von Neurogenese und Zellüberleben

Alzheimer's disease (AD), Parkinson's disease (PD) and Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL) show particular neuropathologies prior to cognitive symptoms. Extra- and intracellular amyloid β accumulation (AD), loss of dopaminergic neurons (PD) and vascular and white matter degeneration (CADASIL) are hallmarks disrupting brain homeostasis and observable in hippocampus, substantia nigra and cortex. Methods sensitive enough to detect relevant early histological alterations are needed to enable early interventions. Magnetic resonance elastography (MRE) is a promising method for capturing biomechanical changes of local alterations in tissue viscoelasticity. We investigated how MRE can be used for correlation of viscoelasticity with histology as step towards using MRE for early diagnosis and/or disease staging. Also we studied effects of counteracting mechanisms as varied physical activities and environments. In study 1 we investigated the APP23 mouse model of AD under standard (STD) and enriched (ENR) living conditions at three early disease stages and correlated MRE data to histological changes. The biomechanical response of MRE to brain areas affected in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridin hydrochloride (MPTP) mouse model of PD correlated to histopathology was subject of study 2. In study 3 mice overexpressing wild type Notch3 (TgN3WT ) and mice with a CADASIL mutation were exposed to ENR, a running wheel (RUN) and STD to further elucidate the mutations effect on neurogenesis and cell survival at early disease stages. MRE may also be a candidate for future early diagnosing and staging in CADASIL when early disease features are better understood. In study 1 viscosity (cellular network) and cell numbers in the hippocampus decrease with disease progression in APP23 mice and ENR is insufficient to counteract both processes. Hippocampal elasticity (cell density) is lower in young APP23 mice but increases as intracellular amyloid β deposits transiently rise with age. In study 2 we observed a decrease of viscosity and elasticity in the substantia nigra correlating to neurodegeneration of MPTP treated mice. Study 3 showed that neurogenic stimulation by RUN and ENR is impaired in both TgN3WT and CADASIL mice due to micromilieu changes. In summary with MRE alterations in viscoelasticity in small brain areas are detectable and relatable to early histopathological changes on a cellular level in AD and PD. ENR could not counteract the cell loss and change in viscosity in APP23 mice, nor could neurogenesis be stimulated by RUN or ENR in TgN3WT and CADASIL mice.Morbus Alzheimer (AD), Morbus Parkinson (PD) und Cerebral Autosomal dominante Arteriopathie mit subkortikalen Infarkten und Leukoenzephalopathie (CADASIL) zeigen präsymptomal spezifische Histopathologien. Extra- und intrazelluläre Amyloid-β-Akkumulation (AD), Verlust von dopaminergen Neuronen (PD) und vaskuläre und weiße Substanz Degeneration (CADASIL) stören die Gehirnhomöostase im Hippocampus, in Substantia Nigra und im Kortex. Von besonderem Interesse sind Methoden, die relevante frühhistologische Veränderungen erkennen, um frühzeitig zu intervenieren. Magnetresonanz- Elastographie (MRE) ist eine vielversprechende Methode in der Erfassung lokaler biomechanischer Veränderungen der Gewebe-Viskoelastizität. Viskoelastizitätsparameter wurden mit Histopathologien korreliert, um die Eignung von MRE für die Früherkennung und /oder Stadieneinteilung zu analysieren. Auch wurden Effekte von körperlicher Betätigung und einer reizreichen Umgebung untersucht. In Studie 1 wurde das APP23-Mausmodell für AD unter Standard (STD) und reizvolleren (ENR) Lebensbedingungen in drei frühen Krankheitsstadien untersucht und MRE-Daten zu histologischen Veränderungen korreliert. In Studie 2 wurde die biomechanische Reaktion von MRE auf Hirnareale, die im 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridin-hydrochlorid (MPTP)-Mausmodell von PD betroffen sind untersucht und ebenfalls mit histologischen Veränderungen korreliert. In Studie 3 wurden Mäuse mit Wildtyp Notch3 Überexprimierung (TgN3WT) und Mäuse mit einer CADASIL-Mutation ENR, einem Laufrad (RUN) und STD ausgesetzt, um den Einfluss auf Neurogenese und des Zellüberlebens bei frühen Krankheitsstadien zu ermitteln. MRE kann künftig auch hier nützlich für die Früherkennung/Stadieneinteilung sein, wenn frühe Krankheitsmerkmale besser erfasst sind. In Studie 1 nehmen die Viskosität (zelluläres Netzwerk) und Zellzahlen im Krankheitsverlauf der APP23 Mäuse im Hippocampus ab und ENR ist unzureichend diesen Prozessen entgegenzuwirken. Die hippocampale Elastizität (Zelldichte) ist bei jungen APP23-Mäusen geringer, nimmt aber mit dem vorübergehenden Anstieg der intrazellulären Amyloid-β-Ablagerungen zu. In Studie 2 beobachten wir eine Abnahme der Viskosität und der Elastizität in der Substantia Nigra, die mit der Neurodegeneration von mit MPTP behandelten Mäusen korreliert. Studie 3 zeigte, dass Neurogenese durch RUN und ENR sowohl bei TgN3WT als auch bei CADASIL- Mäusen aufgrund von Veränderungen des Mikromilieus beeinträchtigt wird. Zusammenfassend kann man mit der MRE Veränderungen der Viskoelastizität in kleinen Hirnarealen nachweisen und mit frühen histopathologischen Veränderungen auf zellulärer Ebene korrelieren. ENR konnte dem Zellverlust und der Viskositätsänderung bei APP23-Mäusen nicht entgegenwirken, noch konnte die Neurogenese durch RUN oder ENR in TgN3WT und bei CADASIL-Mäusen stimuliert werden

Dopaminergic Neurodegeneration in the Mouse Is Associated with Decrease of Viscoelasticity of Substantia Nigra Tissue.

The biomechanical properties of brain tissue are altered by histopathological changes due to neurodegenerative diseases like Parkinson's disease (PD). Such alterations can be measured by magnetic resonance elastography (MRE) as a non-invasive technique to determine viscoelastic parameters of the brain. Until now, the correlation between histopathological mechanisms and observed alterations in tissue viscoelasticity in neurodegenerative diseases is still not completely understood. Thus, the objective of this study was to evaluate (1) the validity of MRE to detect viscoelastic changes in small and specific brain regions: the substantia nigra (SN), midbrain and hippocampus in a mouse model of PD, and (2) if the induced dopaminergic neurodegeneration and inflammation in the SN is reflected by local changes in viscoelasticity. Therefore, MRE measurements of the SN, midbrain and hippocampus were performed in adult female mice before and at five time points after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridin hydrochloride (MPTP) treatment specifically lesioning dopaminergic neurons in the SN. At each time point, additional mice were utilized for histological analysis of the SN. After treatment cessation, we observed opposed viscoelastic changes in the midbrain, hippocampus and SN with the midbrain showing a gradual rise and the hippocampus a distinct transient increase of viscous and elastic parameters, while viscosity and-to a lesser extent-elasticity in the SN decreased over time. The decrease in viscosity and elasticity in the SN was paralleled by a reduced number of neurons due to the MPTP-induced neurodegeneration. In conclusion, MRE is highly sensitive to detect local viscoelastic changes in specific and even small brain regions. Moreover, we confirmed that neuronal cells likely constitute the backbone of the adult brain mainly accounting for its viscoelasticity. Therefore, MRE could be established as a new potential instrument for clinical evaluation and diagnostics of neurodegenerative diseases

Representative images of DAB–stained brain slices showing the substantia nigra.

<p>TH+ cells at baseline at -3dpi (a) and directly after MPTP treatment at 3dpi (b) at 50x magnification, indicating a severe loss of dopaminergic neurons induced by MPTP. Iba-1+ at -3dpi (c) and 3dpi (d) at 50x magnification with detail in 200x magnification (scale bar 100 μm), showing a reactive increase in the number of microglia and macrophages in the substantia nigra immediately after MPTP treatment.</p

Representative images of MRI signal and complex modulus map of G' and G''.

<p>Regions of interest: substantia nigra (red line), midbrain (blue line) and hippocampus (yellow line) were marked in T1w-MRI.</p

Results of MRE measurements and histological cell counts in the substantia nigra.

<p>MPTP induced a significant reduction in MRE elasticity (a) and viscosity (b) in the substantia nigra (mean±SEM, n(-3,3,6,10,14,18dpi) = 5). DAB-stained brain sections showed an immediate significant drop in TH+ dopaminergic neurons in the substantia nigra after MPTP treatment (c) (mean±SEM, n(-3dpi) = 4, n(3,6,10,14,18dpi) = 5). DAPI-stained cell amount was decreased by MPTP (d) (mean±SEM, n(-3,3,6,10,14,18dpi) = 2, no statistical analysis). Initially, the amount of Iba1+ microglia and macrophages was significantly raised after MPTP treatment, but ceased over time (e) (mean±SEM, n(-3dpi) = 4, n(3,6,10,14,18dpi) = 5). * vs. -3dpi, *p≤0.05, **p≤0.01, ***p≤0.001. ^# vs. 3dpi, ^# p≤0.05.</p

Results of MRE measurement and histological cell count in the midbrain.

<p>MPTP induced a significant increase of MRE elasticity (a) and viscosity (b) in the midbrain (mean±SEM, n(-3,3,6,10,14,18dpi = 5). DAPI-stained brain sections showed a reduction following MPTP-treatment (c) (mean±SEM, n(-3,3,6,10,14,18dpi) = 2, no statistical analysis). * vs. -3dpi, *p≤0.05, **p≤0.01.</p

Schematic of the mouse MRE apparatus.

<p>Schematic of the mouse MRE apparatus.</p

Results of MRE measurement and histological cell count in the hippocampus.

<p>MPTP induced a transient increase of elasticity (a) and viscosity (b) in the hippocampus at 6dpi (mean±SEM, n(-3,3,6,10,14,18dpi = 5). Quantification of DAPI-stained cells showed an elevated amount at 6dpi (mean±SEM, n(-3,3,6,10,14,18dpi) = 2, no statistical analysis). * vs. -3dpi, **p≤0.01, ***p≤0.001.</p