Phytomedicine Plus (Aug 2022)
Sirtuin inhibition and neurite outgrowth effect as new biological activities for Areca catechu nut alkaloids
Abstract
Background: Areca catechu is distributed in Southeast Asia, and its nut is called “Binrouji” in Japanese. A. nut contains alkaloids typified by arecoline (1), which are reported to exhibit nicotine-like activities such as parasympathetic nerve excitatory activity. A. nut alkaloids, 1, arecaidine (2), guvacoline (3) and guvacine (4), have a basic skeleton similar to nicotinamide (5) which inhibits nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylases [sirtuins (SIRTs)], epigenetic enzymes involved in an acquired gene modification. We therefore hypothesized that the A. nut alkaloids might have an inhibitory activity against SIRTs as does nicotinamide (5). Purpose: The present study aimed to examine the potency of these alkaloids as SIRT inhibitors by the methods involving the assays of enzymatic inhibition, cellular acetylated lysine (Ac-Lys) accumulation, chemical pull-down and Western blot, as well as the test of neurite outgrowth. Methods: Inhibitory activity of the A. nut alkaloids against SIRT1–3 was measured utilizing the fluorometric SIRT1, 2, or 3 deacetylase assay. Pull down of SIRT1 from the lysate of the neuron-like cell, Neuro 2a, was conducted using arecaidine (2) immobilized with magnetic FG-NH2 beads as bait. The effect of arecoline (1) on the accumulation of acetylated histone was evaluated using an anti-Ac-Lys-antibody. The neurite outgrowth activity of arecoline (1) was assessed by using microscope. Results: Arecoline (1) and arecaidine (2) showed an inhibitory activity stronger than and equal to the representative SIRT inhibitor, nicotinamide (5), respectively. In support for this finding, SIRT1 was pulled down from the Neuro 2a lysate with arecaidine (2) immobilized on the FG NH2-beads, and accumulated acetyl-lysine was detected from the lysate of arecoline (1)-incubated Neuro 2a cells. Additionally, alkaloid 1 showed a neurite outgrowth effect though at the concentration range which did not inhibit SIRT1–3. Conclusion: We found out that arecoline (1) had the SIRT1–3 inhibition activities higher than the typical SIRT inhibitor, nicotinamide (5), while arecaidine (2) had the activities comparable to those of 5. Additionally, we disclosed that alkaloid 1 has another beneficial effect, the neurite outgrowth activity, though not related to SIRT inhibitory activity.
