Journal: bioRxiv

Article Title: Cellular transcriptomics reveals evolutionary adaptation and rumination of vertebrate stomachs

doi: 10.64898/2026.03.09.710455

Figure Lengend Snippet: In situ hybridization of LUC7L transcripts in four stomach chambers (rumen, reticulum, omasum, and abomasum) of sheep and equine stomach, with a scale bar of 100 μm. b, Quantification of the cell proliferation between negative control (NC) and Luc7l -knockdown (si Luc7l ) A7r5 cells, as measured by CCK-8 assay. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-way ANOVA, where *** indicates P < 0.001. c, Migration of A7r5 cells in a comparison of NC and si Luc7l . Wound healing of A7r5 cells was assessed in images (left) at 36 hours (h) post-scratch, relative to the 0-h time point. Wound edge is highlighted by the dashed white line. Migration rate was calculated and compared between NC and si Luc7l . Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t -test, where ** indicates P < 0.01, with a scale bar of 100 μm. d, Cytoskeleton F-actin of A7r5 cells in a comparison of NC (at 0 h) and si Luc7l (at 72 h). The images were all taken from randomly selected fields. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t -test, where ** indicates P < 0.01, with a scale bar of 25 µm. e, Volcano plot showing the up- and down-regulated differentially expressed genes (DEGs) in bulk RNA-seq between the NC and si Luc7l . f, GO cellular component (CC) enrichment analysis of the down-regulated DEGs. g, Schematic diagram illustrating the mechanism that LUC7L regulates the phenotypes of smooth muscle cells (SMCs). The up-regulated expression of LUC7L promotes the switch from synthetic to contractile phenotypes and lowers rates of SMC proliferation and migration. The expression of synthetic genes (e.g., PDGF , FGF , and SERPINE1 genes) are up-regulated significantly in si Luc7l cells, while the contractile genes (e.g., TGF-β and SMAD2 ) are down regulated, compared to NC. h, Representative ventral images of fluorescence emission of abdomen. Mice received a 600 mg/kg body weight of 4-kDa FITC dextran imaging agent in wild-type (WT) mice and homozygous Luc7l knockout ( Luc7l −/− ) mice. Representative ventral images of anesthetized mice are shown at baseline (blank), 5 and 7 min post-gavage. All images were acquired using the same pseudocolor scale of radiance to show relative changes in bioluminescence emission over time. Time to first appearance of fluorescence in the abdomen compared between WT and Luc7l −/− mice. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t-test, where * indicates P < 0.05. i, Gastric emptying 20 min after gavage of 0.5 ml of phenol red solution in WT and Luc7l −/− mice. Data are presented as the mean ± SEM (6 biological replicates). P values are determined by two-tailed unpaired t -test, where *** indicates P < 0.001.

Article Snippet: A7r5 vascular smooth muscle cells (hereafter A7r5, Procell Life Science & Technology) and STC-1 mouse intestinal enteroendocrine cells (hereafter STC-1, Shanghai Jinyuan Biotechnology Co., Ltd.) were maintained in DMEM (Gibco, 11995073) supplemented with 10% fetal bovine serum (FBS) (Gibco, A5670701) and 1% penicillin-streptomycin (Beyotime, C0222).

Techniques: In Situ Hybridization, Negative Control, Knockdown, CCK-8 Assay, Migration, Comparison, Two Tailed Test, RNA Sequencing, Expressing, Fluorescence, Imaging, Knock-Out