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. 2019 Nov 4;9:15927. doi: 10.1038/s41598-019-51799-1

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Effects of siRNA-induced NAPE-PLD silencing on NAPE and FAE levels in SH-SY5Y cells. (A) Napepld transcription and (B1, B2) NAPE-PLD protein levels in SH-SY5Y cells treated for 24 h with scrambled (C, open bar) or siRNA oligonucleotide (closed bar) (n = 8); (B1) membrane and (B2) cytosolic fractions: top, representative blot; bottom, densitometric quantification (expressed as percent control) (n = 3–5). Full-length blots/gels are presented in Supplementary. *P < 0.05, **P < 0.01, ***P < 0.001 two-tailed Student’s t-test. (C,D) NAPE levels in scrambled (C) and NAPE-PLD silenced (D) SH-SY5Y cells after 8 h of 6-OHDA incubation (n = 15 and 10 respectively). Open bars: control cells; color-coded bars: 6-OHDA-treated scrambled cells (blue), 6-OHDA-treated NAPE-PLD-silenced cells (red). (E) Total NAPE levels in SH-SY5Y cells treated for 24 h with scrambled (C) or siRNA oligonucleotide (siRNA). (F) SEA levels in SH-SY5Y cells treated for 24 h with scrambled (C) or siRNA oligonucleotide (siRNA). Open bars: control cells; color-coded bars: 6-OHDA-treated scrambled cells (blue), 6-OHDA-treated NAPE-PLD-silenced cells (red). *P < 0.05, **P < 0.01, ***P < 0.001 two-way ANOVA compared to scramble or vehicle-treated cells; °p < 0.05 °°p < 0.01 two-way ANOVA, Bonferroni post hoc test.