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. 2022 Mar 9;15(3):443–458. doi: 10.1038/s41385-022-00494-y

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© The Author(s) 2022

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. 1 — a Representative magnification of the intestine of either Gpr35^wt or Gpr35b^−/− zebrafish larvae showing Alcian blue staining. Scale bars, 100 μm. b Quantification of the Alcian blue area in (a) performed by using automatic Alcian blue color deconvolution in ImageJ. 1 dot = 1 larva. c Representative magnification of the intestine of Gpr35^wt zebrafish larvae untreated or treated with either zaprinast or LPA and stained with Alcian blue. Scale bars, 100 μm. d Quantification of the Alcian blue area in (c) performed by color deconvolution. 1 dot = 1 larva. e Representative AB/PAS staining of proximal colon sections obtained from Gpr35^f/fVil⁺ and Gpr35^wt co-housed littermates. Scale bars, 50 μm. f Cell count of goblet cells in (e) performed blindly by two different investigators in at least 30 crypts. g Transmission electron microscopy analysis of goblet cell morphology in the proximal colon of Gpr35^f/fVil⁺ and Gpr35^wt co-housed littermates. Red arrowheads indicate fused granules. Scale bars, 5 μm. h Quantification of the number of mucin granules per goblet cell in (g) performed blindly by two different investigators in at least 30 cells. i Representative images of proximal colon sections obtained from Gpr35^f/fVil⁺ and Gpr35^wt co-housed littermates and stained for Muc2 protein by immunohistochemistry. Scale bars, 50 μm. j mRNA expression levels of Muc2 measured by RT-qPCR in proximal colon samples obtained from Gpr35^f/fVil⁺ (n = 4) and Gpr35^wt co-housed littermates (n = 4). k Representative images of proximal colon sections obtained from Gpr35^f/fVil⁺ and Gpr35^wt co-housed littermates and stained for Agr2 protein by immunohistochemistry. Scale bars, 50 μm. mRNA expression levels of (l) Atoh1, Hes1, (m) Gfi1 and Spdef measured by RT-qPCR in proximal colon samples obtained from Gpr35^f/fVil⁺ (n = 4) and Gpr35^wt co-housed littermates (n = 4). Each dot represents one animal with medians. Data are represented as mean ± SEM, ns not significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001 by Mann–Whitney U test.