Liraglutide Ameliorates Hyperhomocysteinemia-Induced Alzheimer-Like Pathology and Memory Deficits in Rats via Multi-molecular Targeting

Abstract

Hyperhomocysteinemia (Hhcy) is an independent risk factor for Alzheimer's disease (AD), and insulin-resistance is commonly seen in patients with Hhcy. Liraglutide (Lir), a glucagon-like peptide that increases the secretion and sensitivity of insulin, has a neurotrophic or neuroprotective effect. However, it is not known whether Lir ameliorates the AD-like pathology and memory deficit induced by Hhcy. By vena caudalis injection of homocysteine to produce the Hhcy model in rats, we found here that simultaneous administration of Lir for 2 weeks ameliorated the Hhcy-induced memory deficit, along with increased density of dendritic spines and up-regulation of synaptic proteins. Lir also attenuated the Hhcy-induced tau hyperphosphorylation and Aβ overproduction, and the molecular mechanisms involved the restoration of protein phosphatase-2A activity and inhibition of β- and γ-secretases. Phosphorylated insulin receptor substrate-1 also decreased after treatment with Lir. Our data reveal that Lir improves the Hhcy-induced AD-like spatial memory deficit and the mechanisms involve the modulation of insulin-resistance and the pathways generating abnormal tau and Aβ.

Keywords: Glucagon-like peptide-1 receptor; Hyperhomocysteinemia; Liraglutide; Tau; β-Amyloid.

Fig. 1

Liraglutide (Lir) decreases plasma Hcy level and ameliorates memory deficits in Hhcy rats. A Administration of Lir restored plasma Hcy level as measured by ELISA. B Administration of Lir improves spatial learning, as shown by a decreased escape latency to find the hidden platform in the Morris water maze test. C, D Administration of Lir improves spatial memory as shown by the decreased time to reach the platform location (latency) and the increased total distance in the target quadrant after removal of the platform on day 7. E Administration of Lir did not affect the swimming speed of the mice, suggesting no changes in motor function. L, low Lir (150 μg/kg per 12 h); M, medium Lir (300 μg/kg per 12 h); H, high Lir (450 μg/kg per 12 h). The data are expressed as mean ± SEM (n = 10/group). *P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05, ^##P < 0.01 vs Hcy.

Fig. 2

Liraglutide (Lir) rescues Hhcy-inhibited spine formation and expression of synapse-associated proteins in rat hippocampus. A, B Representative Golgi-Cox images and statistics showing that administration of Lir preserves spine density in cortical neurons (scale bar, 10 μm; n = 2/group). C, D Western blots and statistics showing that Lir restores the levels of synapse-associated proteins in hippocampus (GAPDH was used as loading control; n = 6/group). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05, ^##P < 0.01 vs Hcy.

Fig. 3

Liraglutide (Lir) attenuates Hhcy-induced tau hyperphosphorylation. A–C Administration of Lir attenuates Hhcy-induced tau hyperphosphorylation at several AD-associated epitopes as measured by Western blotting (A and B, n = 6/group) and immunohistochemistry (n = 2/group; scale bar, 100 μm). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05; ^##P < 0.01 vs Hcy.

Fig. 4

Liraglutide (Lir) activates PP2A by decreasing the inhibitory phosphorylated and demethylated PP2Ac. A, B Lir restores Hhcy-induced PP2A inhibition shown by the reduced phosphorylation (pPP2A) and de-methylation (Dm-PP2A) measured by Western blotting. Data are expressed as mean ± SEM (n = 4/group; *P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05; ^##P < 0.01 vs Hcy).

Fig. 5

Liraglutide (Lir) attenuates Hhcy-induced Aβ overproduction with modulated APP phosphorylation and the activity of secretases. A, B Administration of Lir arrests the Hhcy-induced Aβ_1-40 and Aβ_1-42 elevation in hippocampal extracts measured by ELISA (n = 6/group). C, D Lir attenuates Hhcy-induced elevation of pS668-APP, BACE1, and PSEN1 with restoration of ADAM10 assessed by Western blotting and the quantitative analysis (n = 4/group). Data are expressed as mean ± SEM (*P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05, ^##P < 0.01 vs Hcy).

Fig. 6

Liraglutide (Lir) restores Hhcy-induced impairment of insulin sensitivity. A, B Administration of Lir at the middle (M) and high (H) doses increases the expression level of insulin receptor (IRS) and glucagon-like peptide-1 receptor (GLP-1R) as assessed by Western blotting (n = 4/group). C, D Administration of Lir attenuated Hhcy-induced plasma glucose elevation measured at 1 h and 2 h after glucose gavage (C) without significantly affecting the plasma insulin level (D). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01 vs Ctrl; ^#P < 0.05, ^##P < 0.01 vs Hcy.