Icariside II promotes the differentiation of human amniotic mesenchymal stem cells into dopaminergic neuron-like cells.
Published on Mar 15, 2021in In Vitro Cellular & Developmental Biology – Animal1.665
· DOI :10.1007/S11626-021-00556-8
The purpose of this study is to observe the effect of icariside II (ICS II) on the differentiation of human amniotic mesenchymal stem cells (hAMSCs) into dopaminergic neuron-like cells, the involvement of PI3K signaling pathway inhibitors. After identifying hAMSCs by flow cytometry, hAMSCs were induced and treated with ICS II at 10 μmol/L, 3 μmol/L, 1 μmol/L, and 0 μmol/L. hAMSCs in the LY294002+3μM ICS II group were pretreated with 20 μmol/L LY294002, a PI3K-specific inhibitor, for 1 h, and then hAMSCs were induced with 3 μmol/L ICS II. On the 21st day of induction, immunofluorescence was used to detect expression of the neuronal nuclei (NeuN), neuron-specific enolase (NSE), microtubule-associated protein-2 (MAP-2), glial fibrillary acidic protein (GFAP), and tyrosine hydroxylase (TH) antigens in each induced cell group. Western blotting was used to detect the relative protein expression of NSE, MAP-2, GFAP, and TH. ELISA was used to detect the dopamine concentration in the induction medium supernatant of each group. After 21 d of ICS II induction, immunofluorescence showed that GFAP expression was not obvious in any hAMSC group. The NeuN, NSE, MAP-2, and TH fluorescent proteins were expressed in each group. NeuN was expressed in the nucleus and cytoplasm, while NSE, MAP-2, and TH were mainly expressed in the cytoplasm. The positive cell rates of NeuN, NSE, MAP-2, and TH in the 10 μmol/L, 3 μmol/L, and 1 μmol/L ICS II groups were higher than those in the LY294002+3μM ICS II and control groups. After 21 d of induction, the Western blot results showed that the protein expression levels of NSE, MAP-2, and TH in the 10 μmol/L, 3 μmol/L, and 1 μmol/L ICS II groups were significantly higher than those in the LY294002+3μM ICS II and control groups. The MAP-2 protein expression levels in the 10 μmol/L and 3 μmol/L groups were higher than that in the 1 μmol/L group. After 21 d of induction, the dopamine concentrations in the culture supernatants of the 10 μmol/L, 3 μmol/L, and 1 μmol/L ICS II groups were higher than those in the LY294002+3μM ICS II and control groups. In our experiment, ICS II induced hAMSCs to differentiate into dopaminergic neuron-like cells, and the optimal concentration range of ICS II was 3–10 μmol/L. Moreover, the PI3K signaling pathway is involved in the above differentiation process.