Anti-amyloidogenic constituents from roots of Dryopteris crassirhizoma in Alzheimer's disease cellular model

β-amyloid (Aβ), one of the main causes of Alzheimer's disease (AD) [1], is generated by the amyloidogenic pathway from amyloid precursor protein (APP) by β-secretases and γ-secretases [2]. The generated Aβ monomers aggregate into oligomers and fibrils, and induce neurotoxicity and neuronal cell death [3]. Thus, we investigated the anti-amyloidogenic compounds from natural sources using Chinese Hampster Ovary (CHO) cells stably over-expressing APP (APP-CHO cells). The methanol extract of Dryopteris crassirhizoma Nakai [Dryopteridaceae] roots efficiently reduced the Aβ production. The reduction was more pronounced by the n-butanol layer of the extract. Active constituents inhibiting Aβ production were isolated by activity-guided fractionation. Nine compounds were isolated including the flavonoids epicatechin (1), β-carboxylmethyl-(-)-epicatechin (2), eriodictyol (5), the chromones 7-methoxy-isobiflorin (3), biflorin (4) and noreugenin (6), and the phloroglucinols butyrylphloroglucinol (7), 2-propionyl-4-methylphloroglucinol (8) and 2-butyryl-4-methyl-phloroglucinol (9). All compounds were tested for the effects on the production of Aβ by ELISA and sAPPβ by Western blot analysis. Compounds 3-9 significantly inhibited the production of Aβ. Particularly, 20 µg/mL of compound 7 and 9 reduced 80% of Aβ production compared to controls (IC₅₀= 9.8 and 5.2 µg/mL, respectively). In addition, compounds 3-9 also significantly reduced the production of sAPPβ. Compound 7 and 9 (20 µg/mL) reduced the sAPPβ production by 60 and 80% compared to the controls, respectively. Furthermore, these results were accompanied with the reduced protein expression of β-secretases. Compound 7 and 9 (20 µg/mL) reduced about 30% of the expression of β-secretases compared to the controls. The results suggest that compounds 3-9 significantly reduced the Aβ production by inhibiting β-secretases and if they can cross the blood-brain barrier might be beneficial for AD as preventatives or therapeutics.

Acknowledgements: This work was supported by the research fund of the National Research Foundation of Korea (NRF-2010 – 000398, NRF-2011 – 000398).

Keywords: Dryopteris crassirhizoma, beta-amyloid, Alzheimer's disease, beta-secretase.

References:

[1] Tanzi RE, Moir RD, Wagner SL. Clearance of Alzheimer's Abeta peptide: the many roads to perdition. Neuron 2004; 43: 605 – 608

[2] Zhang H, Ma Q, Zhang YW, Xu H. Proteolytic processing of Alzheimer's beta-amyloid precursor protein. J Neurochem 2012; 120 (Suppl 1): 9 – 21

[3] Kawahara M, Kuroda Y. Molecular mechanism of neurodegeneration induced by Alzheimer's beta-amyloid protein: channel formation and disruption of calcium homeostasis. Brain Res Bull 2000; 53: 3893 – 3897

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