Glimepiride mitigates tauopathy and neuroinflammation in P301S transgenic mice: role of AKT/GSK3β signaling

Abstract

Background and objective: Tauopathy is a group of neurodegenerative diseases in which the pathogenesis processes are related to tau protein. The imbalances between the activities of kinases and phosphatases of tau protein lead to tau hyperphosphorylation and subsequent neurodegeneration. Numerous studies suggest a strong linkage between type 2 diabetes mellitus (T2D) and neurodegenerative diseases. Therefore, finding a drug with a dual therapeutic activity against T2D and neuroprotective will be a promising idea. Hence, the potential neuroprotective effect of Glimepiride (GPD) against tauopathy was evaluated in the current study.

Methods: P301S mice model was employed for tauopathy and C57BL/6 wild type mice (WT) was used as control. Phosphorylated and acetylated tau protein levels was assessed in cortex and hippocampus by western blot. Effect of GPD on tauopathy related enzymes, neuroinflammation, apoptotic markers were evaluated. Furthermore, the neuroprotective effects against anxiety like behavior and motor impairment was analyzed using Parallel rod floor and Open field tests.

Results: GPD significantly ameliorates motor impairment, anxiety like behavior and neurodegeneration in P301S mice. Phosphorylated tau and acetylated tau were significantly decreased in both cortex and hippocampus of P301S mice via decreasing GSK3β, increasing ratio of phosphorylated-AKT to total-AKT, increasing PP2A and normalization of CDK5 levels. Furthermore, GPD treatment also decreased neuroinflammation and apoptosis by reducing NF-kB, TNF-α and caspase 3 levels.

Conclusion: The current data suggests that GPD exerts a protective effect against tauopathy, behavioural consequences, neurodegeneration, neuroinflammation and apoptosis. GPD is therefore a promising agent for the treatment of neurodegenerative diseases associated with tauopathy.

Keywords: GSK3β; Glimepiride; Neuroinflammation; P301S; PP2A; Tauopathy.

Fig. 1

Study timeline in terms of days of administration of GPD for screening phase and subsequent mechanistic phase

Fig. 2

Results of open field neurobehavioral test. Study of the effect of GPD treatment with doses (1, 2, 4 mg/kg) for 21 days on different parameters; a Total distance travelled (m), b Maximum speed (msec⁻¹), c Time in the center zone (sec), d Time in the corners zone (sec). Data are presented as mean ± SD (n = 8) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups respectively (P < 0.05)

Fig. 3

Results of parallel rod floor neurobehavioral test. Study of the effect of GPD treatment with doses (1, 2, 4 mg/kg) for 21 days on different parameters; a Number of foot slips, b Total distance travelled (m), c Rotations of the animal’s body, d Number of line crossings. Data are presented as mean ± SD (n = 8) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups respectively (P < 0.05)

Fig. 4

Effect of GPD treatment on tauopathy induced histopathological changes represented by photomicrographs of H&E stained cortical and different regions of hippocampal sections (hippocampus Cornu Ammonis; CA1 region, hippocampus CA2 region, hippocampus CA3 region and hippocampus Dentate Gyrus DG) in both P301S and WT treated with GPD for 21 days. Photomicrographs showing severe neuronal degeneration and necrosis of several neurons in P301S group. In contrast, normal cortical and hippocampal neurons in WT group and WT received (4 mg/kg) GPD. Further, cerebral and hippocampal sections from P301S groups received GPD with doses (1, 2, 4 mg/kg) showed dose dependent reduction in neuronal degeneration and number of apoptotic neurons in cortex and hippocampus (400× scale bar = 50 μm). Degeneration of neurons are represented by yellow arrows, vacuoles are represented by black arrow, apoptotic neurons are represented by blue arrow, neurofibrillary loss is represented by red arrow and necrosis of several neurons that lose their nuclei in CA2 region is represented by white arrows

Fig. 5

Effect of GPD treatment on tauopathy induced neurodegenerative changes represented by photomicrographs of Nissl blue stained hippocampal and cortical sections in both p301S and WT treated with GPD for 21 days; a photomicrographs showing Nissl granules (black arrows) which obviously decreased in density and distribution in vehicle treated p301s mice and increased with GPD treatment (400X scale bar = 50 μm); b quantification of Nissl bodies by Image J software in the cerebral cortex, c quantification of Nissl bodies by Image J software in hippocampal CA, and d quantification of Nissl bodies by Image J software in DG regions which is significantly decreased in p301S when compared with WT group and p301s mice received (1, 2, 4 mg/kg) GPD. Meanwhile, Number of Nissl positive cells in cerebral cortex, hippocampal CA, and DG regions is significantly increased in p301s mice receiving (1, 2, 4 mg/kg) GPD particularly in p301s group receiving (4 mg/kg) GPD when compared with vehicle treated p301s group. Data are presented as mean ± SD (n = 10) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, #: Statistically significant from P301S and WT groups respectively (P < 0.05)

Fig. 6

a Microscopic pictures of Congo red stained cerebral sections showing presence of positively red stained β-amyloid in vehicle treated P301S mice and P301S mice receiving (1, 2 mg/kg) GPD for 21 days (400X scale bar = 50 μm); b Quantitation of numbers of plaques by Image J software showed absence of plaques in P301S treated with(4 mg/kg) GPD and significantly less in the cortex of groups P301S receiving (1, 2 mg/kg) GPD when compared with vehicle treated P301S group. Data are presented as mean ± SD (n = 10) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups respectively (P < 0.05)

Fig. 7

Effect of GPD doses (1, 2 and 4 mg/kg) on phosphorylated tau deposition in P301S mice with or without treatment for 21 days detected by western blot. Panels (a–c) represent western blot of phosphorylated tau in cortex and its quantitation. Panel (b, d) Represents western blot of phosphorylated tau in hippocampus and its quantitation. Data are presented as mean ± SD (n = 3) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups, respectively (P < 0.05)

Fig. 8

Effect of GPD doses (1, 2 and 4 mg/kg) on acetylated tau protein deposition in P301S mice with or without treatment for 21 days detected by western blot. Panels (a–c) represent western blot of acetylated tau protein in cortex and its quantitation. Panel (b, d) Represents western blot of acetylated tau protein in hippocampus and its quantitation. Data are presented as mean ± SD (n = 3) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups, respectively (P < 0.05)

Fig. 9

Effect of GPD treatment with dose (4 mg/kg) for 21 days on tauopathy related kinases and phosphatases in P301S mice; a GSK3β level, b ratio of (p-AKT/t-AKT) level, c CDK5 level, d PP2A level. Data are presented as mean ± SD (n = 3) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups, respectively (P < 0.05)

Fig. 10

Effect of GPD treatment with dose (4 mg/kg) for 21 days on tauopathy induced Neuroinflammation and apoptosis in P301S mice; a NF-kB level, b TNF-alpha level, c Caspase 3 level. Data are presented as mean ± SD (n = 3) Statistical Analysis was carried out using one-way ANOVA followed by Tukey’s post-hoc test. *, # Statistically significant from P301S and WT groups, respectively (P < 0.05)