Neurogastroenterology & Motility. 2020;00:e13990. wileyonlinelibrary.com/journal/nmo | 1 of 15 https://doi.org/10.1111/nmo.13990 © 2020 John Wiley & Sons Ltd Received: 10 October 2019 | Revised: 12 August 2020 | Accepted: 25 August 2020 DOI: 10.1111/nmo.13990 ORIGINAL ARTICLE Supernatants of intestinal luminal contents from mice fed high- fat diet impair intestinal motility by injuring enteric neurons and smooth muscle cells Yvonne Nyavor 1 | Catherine R. Brands 1 | Jessica Nicholson 1 | Sydney Kuther 1 | Kortni K. Cox 1 | George May 1 | Christopher Miller 1 | Allysha Yasuda 1 | Forrest Potter 1 | Joshua Cady 1 | Heino M. Heyman 2 | Thomas O. Metz 2 | Timo D. Stark 3 | Thomas Hofmann 3 | Onesmo B. Balemba 1 Abbreviations: ALC10 +11, HPLC subfractions 10 and 11 of the aqueous SPE1 fractions of ICS; ANNA1, anti-neuronal nuclear antibody 1; DHE, dihydroethidium; ENS, enteric nervous system; GI, gastrointestinal; GS, anti-glutathione synthase antibody; HFD, high-fat diet; HFD-ICS, ileocecal supernatants from HFD mice; HFD-SPE1, aqueous solid-phase extraction fraction of HFD-ICS; HPLC, high-performance liquid chromatography; ICS, ileocecal supernatants; IJPs, inhibitory junction potentials; IL-6, Interleukin 6; iNOS, inducible nitric oxide synthase; LPS, lipopolysaccharide; MCP1, monocyte chemo-attractant protein 1; MLC1, HPLC subfraction 1 of the methanolic SPE4 fractions of ICS; nNOS, neuronal nitric oxide synthase; PAI-1, plasminogen activator inhibitor-1; SCD- SPE1, aqueous solid-phase extraction fraction of SCD-ICS; SCD, standard chow diet; SCD-ICS, ileocecal supernatants from SCD mice; SMCs, smooth muscle cells; SPE, solid-phase extraction; T2D, type 2 diabetes; TNFα, Tumor necrosis factor-alpha; VIP, vasoactive intestinal peptide. 1 Department of Biological Sciences, University of Idaho, Moscow, ID, USA 2 Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland, WA, USA 3 Lehrstuhl für Lebensmittelchemie und Molekulare Sensorik, Technische Universität München, Freising, Germany Correspondence Onesmo B. Balemba, Department of Biological Sciences, University of Idaho, 875 Perimeter Drive, LSS 252, Moscow, ID 83844, USA. Email: obalemba@uidaho.edu Funding information This study was supported by the University of Idaho—Dyess Faculty Fellowship and National Institutes of Health grant number S10OD018044 to Deborah Stenkamp. Additional support was by Institutional Development Awards (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant numbers P20 RR016454 and the National Center for Research Resources grant number P20 GM103408 through IDAHO INBRE. Abstract Background: Damage to enteric neurons and impaired gastrointestinal muscle contrac- tions cause motility disorders in 70% of diabetic patients. It is thought that enteric neuropathy and dysmotility occur before overt diabetes, but triggers of these abnor- malities are not fully known. We tested the hypothesis that intestinal contents of mice with and without high-fat diet- (HFD-) induced diabetic conditions contain molecules that impair gastrointestinal movements by damaging neurons and disrupting muscle contractions. Methods: Small and large intestinal segments were collected from healthy, standard chow diet (SCD) fed mice. Filtrates of ileocecal contents (ileocecal supernatants; ICS) from HFD or SCD mice were perfused through them. Cultured intact intestinal mus- cularis externa preparations were used to determine whether ICS and their fractions obtained by solid-phase extraction (SPE) and SPE subfractions collected by high-per- formance liquid chromatography (HPLC) disrupt muscle contractions by injuring neu- rons and smooth muscle cells. Key Results: ICS from HFD mice reduced intestinal motility, but those from SCD mice had no effect. ICS, aqueous SPE fractions and two out of twenty HPLC subfractions of aqueous SPE fractions from HFD mice blocked muscle contractions, caused a loss of nitrergic myenteric neurons through inflammation, and reduced smooth muscle excit- ability. Lipopolysaccharide and palmitate caused a loss of nitrergic myenteric neurons but did not affect muscle contractions. Conclusions & Inferences: Unknown molecules in intestinal contents of HFD mice trig- ger enteric neuropathy and dysmotility. Further studies are required to identify the toxic molecules and their mechanisms of action.