Antileukotriene therapy by reducing tau phosphorylation improves synaptic integrity and cognition of P301S transgenic mice

Abstract

The 5-lipoxygenase (5LO) is a source of inflammatory leukotrienes and is upregulated in Alzheimer's disease and related tauopathies. However, whether it directly modulates tau phosphorylation and the development of its typical neuropathology in the absence of Aβ or is a secondary event during the course of the disease pathogenesis remains to be fully elucidated. The goal of this study was to evaluate the effect that pharmacologic blockade of this inflammatory pathway has on the phenotype of a transgenic mouse model of tauopathy, the P301S mice. Starting at 3 months of age, P301S mice were randomized to receive zileuton, a specific 5LO blocker, for 7 months; then, its effect on their behavioral deficits and neuropathology was assessed. Inhibition of leukotrienes formation was associated with a reduction in tau phosphorylation and an amelioration of memory and learning as well as synaptic integrity, which were secondary to a downregulation of the cdk5 kinase pathway. Our results demonstrate that the 5LO enzyme is a key player in modulating tau phosphorylation and pathology and that blockade of its enzymatic activity represents a desirable disease-modifying therapeutic approach for tauopathy.

Keywords: leukotrienes; neuroinflammation; rodent behavior; tauopathy; transgenic mice.

Figure 1

Antileukotriene therapy ameliorates behavioral deficits in P301S transgenic mice. Starting at 3 months of age, wild‐type (WT) and P301S mice were randomized to receive zileuton or vehicle until 10 months of age and then assessed in the Y‐maze and the Morris water maze. (a) Total number of arm entries for WT, WT‐zileuton, P301S, and P301S‐zileuton mice. (b) Percentage of alterations for the same four groups of mice (*p < .001). (c) Morris water maze training phase. Latency to initial platform crossing for WT, WT‐zileuton, P301S, and P301S‐zileuton. (d) Average swim speed for the four groups of mice. (e) Probe phase. Total number of entries to the target platform zone for each of the four groups of mice (WT, WT‐zileuton, P301S, and P301S‐zileuton mice) (*p < .001). (f) Latency to initial platform crossing for the same four groups of mice (*p < .001). (n = 10 per group). Results are mean ± SEM

Figure 2

Antileukotriene therapy modulates tau phosphorylation in the brains of P301S tau transgenic mice. Starting at 3 months of age, P301S mice were randomized to receive zileuton or vehicle until 10 months of age and then euthanized, and brain was extracted for biochemistry. (a) Representative Western blot analyses for 5‐lipoxygenase (5LO) in brain cortex homogenates from P301S and P301S‐zileuton mice. (b) Densitometric analyses of the immunoreactivity to the antibodies in panel A. (c) Measurement of LTB ₄ levels in brain cortex homogenates from P301S and P301S‐zileuton mice (*p < .0001) (n = 10 per group). (d) Representative Western blot analyses for total tau (HT7) and phosphorylated tau at residues Ser202/Thr205 (AT8), Thr231/Thr235 (AT180), Thr181 (AT270), Ser396/Ser404 (PHF1), and Ser396 (PHF13) in brain cortex homogenates from P301S and P301S‐zileuton mice. (e) Densitometric analyses of the immunoreactivities to the antibodies shown in panel D (*p < .05). Results are mean ± SEM (n = 7 per group)

Figure 3

Involvement of the cdk5 kinase pathway in the leukotriene ‐dependent effect on tau phosphorylation. Starting at 3 months of age, P301S mice were randomized to receive zileuton or vehicle until 10 months of age and then euthanized, and brain was extracted for biochemistry. (a) Representative Western blot analyses for insoluble total tau fraction (HT7) in brain cortex homogenates from P301S and P301S‐zileuton mice. (b) Densitometric analysis of the immunoreactivity to the antibody presented in panel A (*p < .01). (c) Representative Western blot analyses for cdk5, p35, p25, GSK3α, GSK3β, p‐GSK3α, and p‐GSK3β, in brain cortex homogenates from P301S and P301S‐zileuton. (d) Densitometric analysis of the immunoreactivity to the antibody presented in the previous panel (*p < .01). Values represent mean ± SEM (n = 10 per group)

Figure 4

Antileukotriene therapy ameliorates synaptic integrity and neuroinflammation in P301S transgenic mice. Starting at 3 months of age, P301S mice were randomized to receive zileuton or vehicle until 10 months of age and then euthanized, and brain was extracted for biochemistry. (a) Representative Western blot analyses for synaptophysin (SYP), postsynaptic density protein 95 (PSD95), and microtubule‐associated protein‐2 (MAP2) in brain cortex homogenates from P301S and P301S‐zileuton mice. (b) Densitometric analyses of the immunoreactivities shown in the previous panel (*p < .001) (n = 6 per group). (c) Representative Western blot analyses for glial fibrillary acidic protein (GFAP) and cluster domain (CD) 45 in brain cortex homogenates from P301S and P301S‐zileuton mice. (d) Densitometric analyses of the immunoreactivities to the antibodies from the previous panel (*p < .01) (n = 6 per group). Values represent mean ± SEM