Curcumin-induced upregulation of the anti-tau cochaperone BAG2 in primary rat cortical neurons

Abstract

Alzheimer's disease (AD) is a progressive, neurodegenerative disease characterized by extracellular deposits of amyloid beta (Aβ) protein and intracellular neurofibrillary tangles of hyperphosphorylated tau protein. Various studies suggest that the tau tangle pathology, which lies downstream to Aβ pathology, is essential to produce AD-associated clinical phenotype and thus treatments targeting tau pathology may prevent or delay disease progression effectively. In this context, our present study examined three polyphenol compounds (curcumin, EGCG and resveratrol) for their possible activity against two endogenous proteins (BAG2 and LAMP1) that are shown to play a vital role in clearing tau tangles from neurons. Human epidemiological and animal data suggest potential positive effects of these polyphenols against AD. Here, primary rat cortical neurons treated with these polyphenols significantly up-regulated BAG2 levels at different concentrations, while only EGCG upregulated LAMP1 levels, although at higher concentrations. Importantly, curcumin doubled BAG2 levels at low micromolar concentrations that are clinically relevant. In addition, curcumin also downregulated levels of phosphorylated tau, which may be potentially attributed to the curcumin-induced upregulation in BAG2 levels in the neurons. The present results demonstrate novel activity of polyphenol curcumin in up-regulating an anti-tau cochaperone BAG2 and thus, suggest probable benefit of curcumin against AD-associated tauopathy.

Keywords: (−)-epigallocatechin-3-gallate; AD; ADAM10; AMP-activated protein kinase; AMPK; APP; Alzheimer's disease; Aβ; BACE1; BAG2; BCL2-associated athanogene 2; Curcumin; EGCG; LAMP1; LDH; NFTs; OA; Polyphenol; ROS; Tau; a disintegrin and metalloproteinase domain 10; amyloid beta; amyloid precursor protein; beta-site APP cleaving enzyme-1; lactate dehydrogenase; lysosomal-associated membrane protein 1; neurofibillary tangles; okadaic acid; reactive oxygen species.

Figure 1

Lactate dehydrogenase (LDH) release. Treatment for 24 h with 50 μM curcumin and 50 μM or higher EGCG caused increased LDH release from neurons as compared with controls. In contrast, resveratrol at all concentrations had no effect on LDH release. Data are taken from three different experiments and are expressed as mean ± SD.

Figure 2

Dose-dependent effect of curcumin on BAG2 levels in neurons. Immunoblots show significant up-regulation of BAG2 in a dose-dependent manner, while LAMP1 remained unaffected. Histogram represents quantitative determinations of intensities of the relevant bands of BAG2 normalized to actin. Data represent mean ± SD of three independent experiments (Student’s t test- *, p < 0.05 compared with control).

Figure 3

Effect of EGCG on BAG2 and LAMP1 in neurons. Immunoblots show significant up-regulation of BAG2 at 100 μM, while LAMP1 increased in a dose-dependent manner. Histograms represent mean ± SD of three independent experiments (Student’s t test- *, p < 0.05 compared with respective control).

Figure 4

Effect of resveratrol on BAG2 in neurons. Immunoblots show significant up-regulation of BAG2 at 100 μM, while LAMP1 remained unaffected. Histogram data represent mean ± SD of three independent experiments (Student’s t test- *, p < 0.05 compared with control).

Figure 5

(A) The okadaic acid (OA)-induced pathological hyperphosphorylation of tau in neurons. The AT8 immunoblot shows significant up-regulation in phospho-tau levels in a dose-dependent manner. (B) Inhibition of OA-induced tau hyperphosphorylation by curcumin. Primary rat cortical neurons were treated with 10 nM OA for 24 h, followed by 12.5 μM curcumin for 24 h (post-treatment) or preceded by 12.5 μM curcumin for 24 h (pre-treatment) or together with 12.5 μM curcumin (co-treatment). The AT8 immunoblot shows curcumin efficiently countered OA-induced tau hyperphosphorylation. Histogram data represent mean ± SD of three independent experiments (Student’s t test- *, p < 0.05 compared with control).

Figure 6

(A) The microRNA miR-9 predicted target site on LAMP1 3′UTR. (B) The microRNA analysis. The quantitative real time PCR (qRT-PCR) study showed significant down-regulation of both miR-128 and miR-9 levels in EGCG-treated cells, while curcumin had no significant effect on miR-128 levels. Relative expressions shown are normalized to RNU6B and average control (n=9) expressions (*, p < 0.05 compared with control).