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. 2020 Jun 19;17:195. doi: 10.1186/s12974-020-01846-w

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© The Author(s) 2020

Open AccessThis 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 2 — HFD-induced hypothalamic reactive gliosis is reduced by astrocyte-specific Myd88 KO. a Diagram depicts the experimental design. TMX, tamoxifen. B/W, body weight. IHC, immunohistochemistry. b The expression of Myd88 mRNA was determined using a real-time PCR analysis with RNA samples immunoprecipitated with HA antibody from hypothalamic extracts of Myd88^△GFAP;Rpl22^HA mice that were generated by cross-breeding astrocyte-specific Myd88 KO mice (Myd88^△GFAP) with transgenic mice (Rpl22^HA) expressing HA-tagged Rpl22. Gfap-Cre;Rpl22^HA mice were used as the control (n = 4–5/group). c–i Control (Myd88^fl/fl) mice and Myd88^△GFAP mice were fed a STD or HFD for 16 weeks, and their astrocytes and microglia in the hypothalamic ARC were analyzed with immunohistochemistry using GFAP and Iba1 antibodies. Representative images (c, f) and calculated data (d, e, g, h, i) show the effect of astrocyte-specific Myd88 KO (Myd88^△GFAP) on the HFD-induced increase in the number and intensity of GFAP-positive cells (c–e) and Iba1-positive cells (f–h), and the size of Iba1-positive cells (i) in the ARC compared with the control Myd88^fl/fl mice (n=3–6 sections of 3–6 mice/group). j, k Representative images (j) and calculated data (k) indicate HFD-induced BrdU and Iba1 double-positive cells in the ARC of Myd88^△GFAP mice and control Myd88^fl/fl mice after HFD feeding for 8 weeks. White arrows indicate cells double positive for Iba1 and BrdU (n = 6 sections of 3 mice/group). l Real-time PCR analysis of RNA samples shows the expression of Gfap, Iba1, Tnf-α, Il-1β, Il-6, and Il-10 in the hypothalamus of Myd88^△GFAP mice and control Myd88^fl/fl mice after HFD feeding for 16 weeks (n = 3/group). m Hypothalamic expression of Tnf-α, Il-1β, Il-6, and Il-10 was determined using a real-time PCR analysis of RNA samples (immunoprecipitated with HA antibody) from hypothalamic extracts of Myd88^△GFAP;Rpl22^HA mice and control Gfap-Cre;Rpl22^HA mice after HFD feeding for 16 weeks (n = 3–4/group). Data are presented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. ns, not significant. Scale bar = 100 μm (20 μm for higher magnification view in insets)