Rapamycin attenuates the progression of tau pathology in P301S tau transgenic mice

Abstract

Altered autophagy contributes to the pathogenesis of Alzheimer's disease and other tauopathies, for which curative treatment options are still lacking. We have recently shown that trehalose reduces tau pathology in a tauopathy mouse model by stimulation of autophagy. Here, we studied the effect of the autophagy inducing drug rapamycin on the progression of tau pathology in P301S mutant tau transgenic mice. Rapamycin treatment resulted in a significant reduction in cortical tau tangles, less tau hyperphosphorylation, and lowered levels of insoluble tau in the forebrain. The favourable effect of rapamycin on tau pathology was paralleled by a qualitative reduction in astrogliosis. These effects were visible with early preventive or late treatment. We further noted an accumulation of the autophagy associated proteins p62 and LC3 in aged tangle bearing P301S mice that was lowered upon rapamycin treatment. Thus, rapamycin treatment defers the progression of tau pathology in a tauopathy animal model and autophagy stimulation may constitute a therapeutic approach for patients suffering from tauopathies.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. The extensive cortical tau tangle pathology present in 5.5 months old vehicle (veh) treated P301S mice (A) was widely attenuated in long-term rapamycin (rapa) treated mice (B).

The lowering in tangle formation was most pronounced in the motor cortex (C: left vehicle treated/right rapamycin) and associated with reduced pathological tau hyperphosphorylation at the AT8 and AT100 epitopes (D, E). In parallel, cortical astrogliosis was diminished following rapamycin treatment (F). While there was a trend towards a reduction of the sparse tangles in the hippocampus, the advanced tau pathology in the brain stem however was not significantly ameliorated by rapamycin (G, H). Short-term treatment at 3 months of age for 6 weeks again resulted in a marked reduction of cortical tangles (I) (A–H: 5MT group; I: 6WT. A–C, G–I: Gallyas silver stain; D: AT8 IHC; E: AT100 IHC; F: GFAP IHC. Bar in A equals 300 µm in A and B, 150 µm in C, G and I, 75 µm in D–F, 600 µm in H).

Figure 2. After long-term rapamycin treatment (n = 6), unbiased stereology confirmed a significant reduction of cortical tau tangles to only 14% of the amount of tangles seen in vehicle treated (n = 5) P301S mice (vehicle = 100%).

The number of AT8 stained cells containing hyperphosphorylated tau was reduced to 30% in aged rapamycin treated mice compared to controls (100%) (5MT group). A significant attenuation of Gallyas-stained (to 39% of controls) and AT8-positive cells (to 46% of controls) was also achieved by late short-term rapamycin treatment (6WT, n = 6/6). The reduction of tangles in the hippocampus (to 33% of controls) and the brain stem (to 72% of controls) did not reach the level of significance adjusted for multiple testing. Pairwise reduction of Gallyas or AT8 positive counts was analysed using one-sample T-tests per brain region. Significant p-values, adjusted for the multiple comparisons of all 6 tested groups by the Holm-Bonferroni method, are outlined as follows: *p<0.05, **p<0.01, and ***p<0.001.

Figure 3. Levels of sarkosyl extracted insoluble tau were significantly reduced in the forebrain of P301S mice after 5 months of long-term rapamycin treatment (R, n = 6) when compared to vehicle treated mice (V, n = 5) (A, 5MT group; Western blot using BR134 antibody).

A comparable lowering of insoluble tau was obtained by late short-term rapamycin administration over 6 weeks (B; 6WT group, n = 6/6). In parallel, the accumulation of tau hyperphosphorylated at the AT8 and AT100 epitopes was significantly lowered (C; 6WT group, n = 6/6). Quantification of tau Western blots was subjected to unpaired T-tests and T-tests adjusted for unequal variances (Welch-Test), yielding both similar results. *p<0.05 and **p<0.01.

Figure 4. Consistent with cerebral mTOR inhibition, phosphorylation of S6 (S6P) was significantly reduced following rapamycin administration (A, 5MT group, n = 4/4).

Compatible with an induced autophagy pathway, LC3II levels were increased upon rapamycin treatment (B, 6WT group, n = 6/6). High levels of the autophagy associated proteins p62 and LC3 were measured in aged vehicle treated P301S transgenic mice (C; 5MT Vehicle, n = 4). This accumulation of p62 and LC3 was prevented by long-term rapamycin administration, pointing towards a restored autophagic flux (C; 5MT Rapa, n = 4). Forebrain tissue was used and data was analyzed by ANOVA. *p<0.05, **p<0.01, and ***p<0.001.