94 Chapter 4 Figure 4. Microglia in the AppNL-G-F and TAUP301S model increase TSPO expression. a,b Representative images of TSPO expression in microglia and astrocytes in AppNL-G-F hippocampus. c An increase was observed in IBA1+ microglia at 28 weeks (P = 0.0078, t=3.522, df=8) but not 10 weeks (P = 0.8788, t=0.1565, df=10) in AppNL-G-F hippocampus compared to control. d No increase in astrocytes was observed (10 weeks: P = 0.6266, t=0.5019, df=10; 28 weeks: P = 0.4425, t=0.8080, df=8). e TSPO+ cells were increased at 28 weeks (P = 0.0079, U=0, ranks=15, 40) but not at 10 weeks (P = 0.2375, t=1.257, df=10) in the AppNL-G-F mice. f,g Both TSPO+ microglia (P = 0.0005, t=5.658, df=8) and astrocytes (P = 0.0030, t=4.207, df=8) were increased at 28 weeks in the hippocampus of AppNL-G-F mice but not at 10 weeks (microglia: P = 0.7213, t=0.3670, df=10; astrocytes: P = 0.9561, t=0.056, df=10) . h Activated microglia (P < 0.0001, t=7.925, df=8), but not astrocytes (P = 0.3095, U=7, ranks=33, 22), in the AppNL-G-F model have increased TSPO expression. i,j Representative images of TSPO expression in microglia and astrocytes in TAUP301S hippocampus. k-m No increases in microglia (8 weeks: P = 0.3687, t=0.9608, df=7; 20 weeks; P = 0.9647, t=0.04580, df=7), astrocytes (8 weeks: P = 0.7353, t=0.3519, df=7; 20 weeks; P = 0.0870, t=1.989, df=7) or TSPO+ cells (8 weeks: P = 0.8492, U=9, ranks=19, 26; 20 weeks; P = 0.0876, t=1.985, df=7) were observed in the hippocampus of TAUP301S mice. n,o No increase was observed in the number of TSPO+ microglia (8 weeks: P = 0.2787, t=1.174, df=7; 20 weeks; P = 0.0907, t=1.961, df=7) or astrocytes (8 weeks: P = 0.8684, t=0.1718, df=7; 20 weeks; P = 0.1984, U=4.5, ranks=14.5, 30.5). p Microglia in the TAUP301S increase TSPO expression (P = 0.0133, t=3.471, df=6) whereas astrocytes do not (P = 0.5800, t=0.5849, df=6). Statistical significance in c-h and k-p was determined by a two-tailed unpaired t-test or Mann-Whitney U-test when not normally distributed. Box and whiskers mark the 25th to 75th percentiles and min to max values, respectively, with the median indicated. Scale bar = 50µm, inserts are digitally zoomed in (200%). observed in microglia (Fig. 4k) or astrocyte (Fig. 4l) densities, in TSPO+ cell density (Fig. 4m), or in the density of TSPO+ microglia (Fig. 4n) or of TSPO+ astrocytes (Fig. 4o) in the hippocampus at either 8 or 20 weeks (Fig. 4) However, as with the AppNL-G-F mouse (and in contrast to the human), a 2-fold increase in individual cellular TSPO expression was observed within microglia in TAUP301S mice (Fig 4p). Again, as with the AppNL-G-F mouse, individual cellular TSPO expression within astrocytes was unchanged. In summary, we showed that TSPO cellular expression is increased within microglia from AppNL-G-F and TAUP301S mice, but not in microglia from AD tissue. TSPO was also unchanged in astrocytes from both mouse models and the human disease. Microglial TSPO is upregulated in SOD1G93A mice but not in ALS Spinal cord and brain microglia differ with respect to development, phenotype and function70. We therefore next investigated ALS (Table S2), that primarily affects the spinal cord rather than the brain. We compared this data to that from the commonly used SOD1G93A mouse model of ALS. TSPO expression was investigated in the ventral horn and lateral columns of the spinal cord in cervical, thoracic, and lumbar regions (Fig. 5a-c). An increase in microglia (Fig. 5d), HLA-DR+ microglia (Fig. 5e) and astrocytes (Fig. 5f) was observed in human ALS spinal cord. The density of TSPO+ cells was increased by 2.5-fold in ALS spinal cords across all regions when compared to controls (Fig. 5g). No additional changes were found when stratifying the cohort based on disease duration or spinal cord regions, white or grey matter, or spinal cord levels. In comparison to the controls, ALS samples exhibited a 3-fold increase in the density of TSPO+ microglia (TSPO+IBA1+ cells, Fig. 5h) and a 3-fold increase in TSPO+ activated microglia/macrophages (TSPO+HLA-DR+ cells, Fig. 5i). A 2.5-fold increase in the density of TSPO+ astrocytes (TSPO+GFAP+ cells) was observed in ALS compared to control (Fig. 5j). We then quantified TSPO+ area in microglia and astrocytes as an index of individual cellular TSPO expression (Fig. 5k). No increase in TSPO+ area (µm2) was found in microglia or astrocytes in ALS when compared to control (Fig. 5k), implying that TSPO expression does not increase in microglia or astrocytes with ALS.
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