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. 2019 Jul 18;10(8):549. doi: 10.1038/s41419-019-1787-7

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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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Fig. 3 — a, b mRNA level (a) (n = 5–6) and protein expression (b) (n = 8) of Bnip3 in NRVMs transfected with non-targeting siRNA (CTRL siRNA) or siRNA targeting TCTP (TCTP siRNA #1) for 72 h. c–e NRVMs were transfected with non-targeting siRNA (CTRL siRNA) or siRNA targeting TCTP (TCTP siRNA #1), or Bnip3 (Bnip3 siRNA #1), or a mixture of both (TCTP siRNA #1 & Bnip3 siRNA #1). Cell death (c) (n = 4) and apoptosis (d) (n = 4–5) were determined by calcein-AM (green)/ethidium homodimer-1 (red) staining and flow cytometry after siRNA transfection for 72 h. Scale bar, 200 μm. e mPTP opening was assessed by co-loading with calcein/AM and CoCl2 after siRNA transfection for 24 h. Loss of green fluorescence is indicative of mPTP opening (n = 6). Scale bar, 40 μm. f NRVMs were infected with Ad-LC3-GFP (30 MOI) for 8 h and then transfected with non-targeting siRNA or siRNA targeting TCTP (TCTP siRNA #1), or Bnip3 (Bnip3 siRNA #1) or a mixture of both (TCTP siRNA #1 & Bnip3 siRNA #1) for 48 h. Green puncta indicate autophagosomes (n = 6). Scale bar, 25 μm. g Expression of LC3BII and GAPDH proteins in NRVMs transfection with siRNA for 72 h (n = 4). h NRVMs were infected with Ad-LC3-GFP (30 MOI) for 8 h and then transfected with non-targeting siRNA (CTRL siRNA) or siRNA targeting TCTP (TCTP siRNA #1) with or without BFA treatment for 6 h. Autophagosome formation was evaluated after 48 h. Green puncta indicate autophagosomes (n = 4). Scale bar, 25 μm. i Expression of LC3BII and GAPDH proteins in NRVMs transfection with siRNA for 72 h (n = 4). *P < 0.05, **P < 0.01, ***P < 0.001. Unpaired, two-tailed Student’s t-test (a, b) or One-way ANOVA followed by Tukey’s test (c–g) or two-way ANOVA followed by Bonferroni’s test (h, i)