Sirtuin 1 independent effects of resveratrol in INS-1E β-cells

- Sirt1 negatively regulates cell size of clonal insulin secreting β-cells.
- Resveratrol causes cell enlargement in a dose-dependent manner in both Sirt1 knockdown and negative-control cells.
- The cell size increase by resveratrol is mTOR cascade-dependent, while Sirt1 knockdown does not change the mTOR activity.
- Cell size is controlled by Sirt1 and mTOR signalling in an independent manner.

Resveratrol (Resv), a natural polyphenol, is suggested to have various health benefits including improved insulin sensitivity. Resv activates Sirtuin (Sirt1) in several species and tissues. Sirt1 is a protein deacetylase with an important role in ageing, metabolism and β-cell function. In insulinoma β-cells (INS-1E), Resv is previously shown to improve glucose-stimulated insulin secretion in a Sirt1-dependent mechanism and to protect against β-cell dedifferentiation in non-human primates, while inducing hypertrophy in myoblasts. Mammalian (mechanistic) Target of Rapamycin (mTOR), is a key regulator of cellular metabolism and regulates the cell size, β-cell survival and proliferation.In order to understand the interaction of Sirt1 and mTOR cascade activity with Resv-induced changes in the INS-1E cell line, we generated stable Sirt1-down-regulated INS-1E cells, and analysed Sirt1-dependent effects of Resv with respect to mTOR cascade activity. Sirt1-knockdown (KD) had a significant increase in cell size compared to negative-control (NEG CTR) cells. Resveratrol treatment increased cell size in both cell types in a dose-dependent manner at 24 h (Resv conc: 15-60 μM), and decreased the cell number (Resv conc: 30-60 μM). Cell area was increased in NEG CTR cells (Resv conc: 60 μM) at 24 h and KD cells at 48 h (Resv conc: 15-60 μM). Rapamycin, a specific mTOR inhibitor, counteracted the Resv-induced cell enlargement (both cell diameter and area). Furthermore, Sirt1-downregulation by itself did not affect the mTOR cascade activities as measured by Western blotting for total and phosphorylated Akt and mTOR. Rapamycin decreased the mTORC1 activity, while increasing the pAkt levels. Resveratrol did not interfere with the mTOR activity or with Sirt1 expression. Altogether, this work indicates that Sirt1 is a negative regulator of cell size. Moreover, the effect of Resv on cell size increase is mTOR-cascade dependent.

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