Neuroprotective effects of Cerebrolysin in triple repeat Tau transgenic model of Pick's disease and fronto-temporal tauopathies

Abstract

Background: Tauopathies are a group of neurodegenerative disorders with accumulation of three-repeat (3R) or four-repeat (4R) Tau. While 3R tau is found in Pick's disease and Alzheimer's disease (AD), 4R tau is more abundant in corticobasal degeneration, progressive supranuclear palsy, and AD. We have previously shown that Cerebrolysin™ (CBL), a neuropeptide mixture with neurotrophic effects, ameliorates the pathology in amyloid precursor protein transgenic (tg) mouse model of AD and 4R tau, however it is unclear if CBL ameliorates the deficits and neuropathology in the mouse model of Pick's disease over expressing 3R tau.

Results: Mice expressing 3R tau (L266V and G272V mutations) under the mThy-1 promoter were treated with CBL in two separate groups, the first was 3 months old (treated for 3 months, IP) and the second was 6 months old (treated for 3 months, IP) at the start of the treatment. We found that although the levels of total 3R tau were unchanged, CBL reduced the levels of hyper-phosphorylated tau in both groups of mice. This was accompanied by reduced neurodegenerative pathology in the neocortex and hippocampus in both groups and by improvements in the behavioral deficits in the nest-building test and water maze in the 3-6 month group.

Conclusion: Taken together these results support the notion that CBL may be beneficial in other taupathy models by reducing the levels of aberrantly phosphorylated tau.

Fig. 1

Immunocytochemical analysis of effects of CBL on levels of 3R Tau transgenic mice. a Immunocytochemistry with an antibody against 3R Tau in the 3–6 month group. b Image analysis of levels of 3R Tau immunoreactivity in the neocortex and hippocampus. c Immunocytochemistry with an antibody against 3R Tau in the 6–9 month group. d Image analysis of levels of 3R Tau immunoreactivity in the neocortex and hippocampus of the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 2

Immunocytochemical analysis of effects of CBL on levels of p-tau in 3R tau transgenic mice. a Immunocytochemistry with an antibody against p-tau (PHF1) in the 3–6 month group. b Image analysis of levels of p-tau immunoreactivity in the neocortex and hippocampus. c Immunocytochemistry with an antibody against p-Tau in the 6–9 month group. d Image analysis of levels of p-tau tau immunoreactivity in the neocortex and hippocampus of the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 3

Western blot analysis of effects of CBL on levels of tau in 3R tau transgenic mice a Representative immunoblots of samples from the 3–6 month group analyzed with antibodies against 3R Tau tTau and p-tau (PHF1). b, c Image analysis of levels of 3R Tau and ratio of p-tau/tTau in the brains of mice from the 3–6 month group. d Representative immunoblots of samples from the 6–9 month group analyzed with antibodies against 3R Tau, tTau and p-tau (PHF1). e, f Image analysis of levels of 3R tau and p-tau/tTau in the brains of mice from the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 4

Analysis of NeuN cell counts in 3R tau transgenic mice treated with CBL. a Immunocytochemistry with an antibody against NeuN in the 3–6 month group. b Stereological analysis with the dissector method to estimate the numbers of NeuN positive cells in the neocortex and hippocampus in the 3–6 month group. c Immunocytochemistry with an antibody against NeuN in the brains of the 6–9 month group. d Stereological analysis with the dissector method to estimate the numbers of NeuN positive cells in the neocortex and hippocampus in the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 5

Analysis of astrogliosis in 3R Tau transgenic mice treated with CBL a Immunocytochemistry with an antibody against GFAP in the 3–6 month group. b Computer aided image analysis of levels of GFAP immunoreactivity in the neocortex and hippocampus in the 3–6 month group. c Immunocytochemistry with an antibody against GFAP in the brains of the 6–9 month group. d Computer aided image analysis of levels of GFAP immunoreactivity in the neocortex and hippocampus in the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 6

Effects of CBL treatment in behavioral alterations in 3R Tau transgenic mice. Mice were evaluated for context-dependent learning in an open field area using a Kinder SmartFrame Cage Rack Station activity monitor system. a, b Levels of total activity and rearing in the cage in the 3–6 month group. c Nesting behavior in the cage of the 3–6 month group. d, e Levels of total activity and rearing in the cage in the 6–9 month group. f Nesting behavior in the cage of the 6–9 month group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 7

Effects of CBL on learning memory in the water maze in 3R Tau tg mice. a Water maze testing presented as distance traveled to find the platform, respectively in the 3–6 month group. During the cued portion of the test both the vehicle and CBL treated non-tg and 3R tau tg mice performed as expected. However during the hidden portion of the test, the vehicle treated 3R tau tg mice went a farther distance to find the platform compared to the non-tg, CBL prevented these effects. b At day 8, during the probe portion of the test (with platform removed) memory retention was evaluated in the 3–6 month group. c In the 3–6 month group, probe test with the visual platform confirmed that no visual alterations were detected. d Water maze testing presented as distance traveled to find the platform in the 6–9 month group. During the cued portion of the test both the vehicle and CBL treated non-tg and 3R tau tg mice performed as expected. However during the hidden portion of the test, the vehicle treated 3R Tau tg mice went a farther distance to find the platform compared to the non-tg. e At day 8, during the probe portion of the test (with platform removed) memory retention was evaluated in the 6–9 month group. f In the 6–9 month group, probe test with the visual platform showed visual alterations in the 3R tau tg group. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to vehicle-treated tg mice control using one way ANOVA with Dunnet’s post hoc test

Fig. 8

Western blot analysis of effects of CBL on levels of Akt/GSK3β in the 3R Tau transgenic mice. a Representative immunoblots of samples from the 6–9 month group analyzed with antibodies against p-GSK3β (Y216), t-GSK3β, t-Akt and p-Akt (Ser473). b, c Image analysis of the ration between p-GSK3β (Y216) and total and p-Akt (Ser473) and total respectively. For analysis, N = 10 non-tg and N = 10 3R Tau tg mice from each age and treatment group. *P < 0.05 when compared to non-tg control using one way ANOVA with Dunnet’s post hoc test. ^# P < 0.05 when compared to tg control using one way ANOVA with Tukey post hoc test