Molecular drivers and cortical spread of lateral entorhinal cortex dysfunction in preclinical Alzheimer's disease

Abstract

The entorhinal cortex has been implicated in the early stages of Alzheimer's disease, which is characterized by changes in the tau protein and in the cleaved fragments of the amyloid precursor protein (APP). We used a high-resolution functional magnetic resonance imaging (fMRI) variant that can map metabolic defects in patients and mouse models to address basic questions about entorhinal cortex pathophysiology. The entorhinal cortex is divided into functionally distinct regions, the medial entorhinal cortex (MEC) and the lateral entorhinal cortex (LEC), and we exploited the high-resolution capabilities of the fMRI variant to ask whether either of them was affected in patients with preclinical Alzheimer's disease. Next, we imaged three mouse models of disease to clarify how tau and APP relate to entorhinal cortex dysfunction and to determine whether the entorhinal cortex can act as a source of dysfunction observed in other cortical areas. We found that the LEC was affected in preclinical disease, that LEC dysfunction could spread to the parietal cortex during preclinical disease and that APP expression potentiated tau toxicity in driving LEC dysfunction, thereby helping to explain regional vulnerability in the disease.

Figure 1

Whole-brain ROI analysis identified dysfunction in the entorhinal cortex and other cortical regions in preclinical Alzheimer’s disease. (a) An example of automated whole-brain segmentation of a CBV image. Left, pre-contrast image in a coronal view (for illustration purposes the right brain is shown unsegmented). Right, sagittal view. (b) Compared with a control group (light gray bars), the preclinical group (dark gray bars) showed reliable reductions in the percentage of CBV in the entorhinal cortex and parahippocampal gyrus. *P < 0.05. Error bars represent s.e.m. (c) Among brain regions affected in preclinical Alzheimer’s disease, only the parahippocampal gyrus and the precuneus cortex were significantly correlated with entorhinal cortex dysfunction by a correlational analysis (**Pearson correlation coefficient >0.7).

Figure 2

Voxel-based analysis pinpoints dysfunction in preclinical Alzheimer’s disease to the LEC. (a) A voxel-based analysis was performed in the temporal lobes (indicated by highlighted areas at left) and the voxels that showed significantly lower CBV in the preclinical are indicated in color in a coronal view (left) and an axial view (right). (b) A higher magnification of the coronal view showing the anatomy of the medial temporal (left and middle). Right, LEC and PRC were the regions with reliable CBV reductions in the preclinical compared to the control group. TEC, transentorhinal cortex; HC, hippocampus proper. (c) An analysis of the control group pinpointed the right LEC as a region in which CBV was correlated with performance on a delayed retention task. Statistics are represented as heat maps of t values corresponding to P < 0.05, corrected for multiple comparisons using the FWE approach.

Figure 3

The LEC is affected by tau and APP coexpression and leads to cortical dysfunction. (a) A volume rendering of the mouse brain, mapping the anatomical locations of the LEC (yellow), MEC (white) and PRC (red). (b) Voxel-based analyses revealed that, compared with controls, EC-APP mice had CBV reductions in the MEC and the CA1 (left), EC-Tau mice showed little differences in CBV (middle), and EC-APP/Tau mice had reliable CBV reductions [in the left LEC and PRC (right). All images are axial slices whose location is illustrated in the volume rendering shown in a. (c) A higher magnification of the slice (as illustrated in the stippled box in the right panel of b) shows the anatomy of the medial temporal region (left and middle; SUB, subiculum; DG, dentate gyrus), and shows that the LEC and the PRC were the regions with reliable CBV reductions in the EC-APP/Tau group (right). (d) More dorsal slices of the EC-APP/Tau group showing additional areas of reduced CBV in the PRC (left and middle) and reduced CBV in PPC (right).

Figure 4

Patterns of cortical spread in mouse models overlap with patterns observed in preclinical Alzheimer’s disease. (a) As illustrated by volume rendering, the brain regions affected in the EC-APP/Tau mice are the LEC (yellow), PRC (red) and the PPC (green). (b) As illustrated by volume rendering, among the regions affected in the human preclinical group, dysfunction in the entorhinal cortex (yellow) was reliably correlated with CBV in the parahippocampal gyrus (red) and precuneus (green).

Figure 5

APP expression acts to potentiate and accelerate tau toxicity in the LEC. (a) Mapping group × age interactions revealed that age-dependent worsening of relative CBV (rCBV) was increased in the MEC of EC-APP mice compared with their controls (left) and increased in the LEC of EC-Tau mice compared with their controls (right). (b) Mapping a group × age interaction revealed age-dependent worsening of CBV in the LEC and PRC (left), and the PPC (right), of EC-APP/Tau mice compared with EC-APP and EC-Tau mice. Line graph illustrations of the group × age interactions in the LEC (upper graph) and the PPC (lower graph) of EC-APP/Tau (red) mice compared with EC-APP and EC-Tau mice (blue). *P < 0.05, **P < 0.001; ^#F_1,83 = 14.346, P = 0.0003; ^##F_1,83 = 4.829, P = 0.031. Data are presented as mean ± s.e.m.

Figure 6

Mapping histological markers of tau and APP in mouse models. (a) Histology analysis in the MTL of APP/Aβ with 6E10 antibody in young EC-APP and EC-APP/Tau mice (left), abnormally conformed human tau with MC1 antibody in young EC-Tau and EC-APP/Tau mice (middle), and total human tau with CP27 antibody in young EC-Tau and EC-APP/Tau (right). Scale bar represents 500 µm. (b) Histology analysis in the MTL of APP/Aβ with 6E10 antibody in old EC-APP and EC-APP/Tau mice (left), conformation changes in tau with MC1 antibody in old EC-Tau and EC-APP/Tau mice (middle), and total human tau with CP27 antibody in old EC-Tau and ECAPP/Tau mice (right). Scale bar represents 500 µm. (c) Histology analysis in the MTL of phospho-tau with AT8 antibody in old EC-Tau and EC-APP/Tau mice (left). In older EC-APP/Tau mice, MC1 immunohistochemistry (20×) revealed the occurrence of redistribution of tau from axons into the somatodendritic compartments compared with EC-Tau mice (middle). A semiquantitative analysis revealed a significant increase in MC1-positive neurons in the entorhinal cortex in the EC-APP/Tau mice (440.1 ± 79.60, n = 7), mainly in the MEC, compared with the EC-Tau mice (99.80 ± 22.92, n = 5; t-test, P = 0.006; right). Data are presented as mean ± s.e.m. Scale bars represent 500 µm (left) and 100 µm (middle). (d) Histology analysis in the parietal lobe of APP/Aβ with 6E10 antibody in old EC-APP and EC-APP/Tau mice (left), conformation changes in tau with MC1 antibody in old EC-Tau and EC-APP/Tau mice (middle), and phospho-tau with AT8 antibody in old EC-Tau and EC-APP/Tau mice (right). Scale bar represents 500 µm.

Figure 7

The LEC shows evidence of high metabolism in young unaffected individuals. (a) The entorhinal cortex was segmented into the MEC (pink) and the LEC (blue) in template brains of young wild-type mice (left) and young healthy human subjects (right). (b) In young wild-type mice (left), the LEC (blue) was found to have higher CBV values than the MEC (pink) (**F_1,34 = 475.176, P < 0.001), and the left LEC was found to have higher CBV values than the right LEC (^#F_1,34 = 6.680, P = 0.01). In young healthy human subjects (right), the LEC (blue) was found to have higher CBV values than the MEC (pink) (*F_1,34 = 706.199, P < 0.001). Data are presented as mean ± s.e.m. (c) Thresholded mean CBV maps in young wild-type mice (left) revealed that the CBV was higher in the LEC than the MEC, with the left LEC showing the highest CBV. Mean CBV maps in young healthy human subjects (right) revealed that the LEC had a higher CBV than the MEC.