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. 2020 May 21;10:8418. doi: 10.1038/s41598-020-65423-0

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© The Author(s) 2020

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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Glomerular SIRT3 activation is associated with enhanced tubular Sirt3 expression. (a,b) Representative images (a) and quantification (b) of in situ hybridization for Sirt3 in kidney cortex from BTBR WT mice and BTBR ob/ob mice treated with vehicle or honokiol at 14 weeks of age. Scale bars: 20 μm. Insets show the localization of Sirt3 in tubular cells. Image-based quantitative software analysis was performed to evaluate tubular expression of Sirt3. Fiji Image J software (https://imagej.net/Fiji) was used for the quantification of the number of Sirt3 mRNA dots (representing single mRNA molecules). Orbit Image analysis software (http://orbit.bio) was used to count the number of cells. Sirt3 mRNA levels were expressed as average number of dots per cell. (c) Quantification of tubular acetylated SOD2 staining in BTBR WT mice (n = 9) and in BTBR ob/ob mice treated with vehicle (n = 6) or honokiol (n = 9) at 14 weeks of age. (d) qRT-PCR analysis of Nampt mRNA levels in kidney of BTBR WT mice (n = 9) and BTBR ob/ob mice treated with vehicle (n = 6) or honokiol (n = 9). (e) Representative images of NRF2 expression in BTBR WT mice (n = 9) and in BTBR ob/ob mice treated with vehicle (n = 6) or honokiol (n = 9) at 14 weeks of age. Scale bars: 50 μm. (f,g) qRT-PCR analysis of Nrf2 (f) and Keap1 (g) mRNA levels in kidney of BTBR WT mice (n = 9) and BTBR ob/ob mice treated with vehicle (n = 6) or honokiol (n = 9). Data are mean ± SEM and were analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test, *p < 0.05, ***p < 0.001 vs BTBR WT mice; ^##p < 0.01 vs BTBR ob/ob ⁺ vehicle.