Doll, C. L. | Search bioRxiv

bioRxiv2026

Ceramide Synthases Regulate Myristate-Induced Intestinal IRE1α Activation

Background & AimsHigh-fat diets (HFDs) are a major modifiable risk factor for intestinal health. Current research focuses primarily on palmitate (C16:0); however, myristate (C14:0, rich in dairy products) has been minimally investigated. HFDs increase ceramide generation which drives endoplasmic reticulum (ER) stress; with both sphingolipids and ER stress being key contributors to intestinal biology. Whether different fatty acids uniquely impact sphingolipid metabolism and ER stress in intestinal biology has not been well defined. MethodsHuman colon epithelial cells were utilized to determine the role of ceramide synthases (CerS) 5 and 6 on myristate-induced ER stress using pharmacologic inhibitors and siRNA. Intestinal epithelial cell specific CerS5 and/or CerS6 knockout mice of both sexes were fed a control, high milk-fat, or high lard-fat diet for 16 weeks. Cells and colon tissues were analyzed for lipids, mRNA, and protein. ResultsMyristate treatment increased C14:0-ceramide and induced IRE1-dependent ER stress. Inhibition of CerS suppressed these effects, yet knockdown of CerS5/6, the primary enzymes generating C14:0-ceramide, unexpectedly exacerbated IRE1 activation both in vitro and in vivo, potentially due to depletion of dihydro(dh)sphingosine. ConclusionsCerS are required for myristate-induced IRE1 activation and restoration of the sphingoid base pool provides partial protection from intestinal ER stress. SYNOPSISThis study identifies a new mechanism linking dietary fats to intestinal cell stress. Ceramide synthases drive ER stress triggered by myristate, a dairy-derived fat, while restoring sphingoid bases partially protects cells, revealing a new role for sphingolipids in shaping intestinal responses to diet. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/728542v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@d66503org.highwire.dtl.DTLVardef@181bffaorg.highwire.dtl.DTLVardef@162f675org.highwire.dtl.DTLVardef@22a511_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2026

Loss of Acid Ceramidase in Myeloid Cells Protects from Chronic Colitis in IL10-Deficent Mice

Background & AimsPatients with inflammatory bowel disease (IBD) exhibit elevated expression of acid ceramidase (AC), a sphingolipid metabolism enzyme. Recent studies have shown that myeloid cells contribute to the elevated expression of AC, such that the conditional loss of AC is protective in IBD. MethodsBone marrow derived macrophages (BMDMs) and neutrophils (BMDNs) were utilized to assess the role of AC in immune cell mediated inflammation. We then crossed conditional ASAH1 LyzMCRE knockout mice with IL10 knockout mice to determine the role of AC in a model of spontaneous colitis. Colon tissues were analyzed for lipids, mRNA, and protein. We performed flow cytometry to determine the role of myeloid AC in recruiting effector T cells in disease. ResultsIn this study, we found that loss of AC impaired secretory and migratory functions in BMDMs, but not BMDNs. Further, the conditional loss of AC protected from spontaneous, chronic colitis. Loss of AC reduced inflammatory markers, increased colon ceramides, and reduced the inflammatory metabolite sphingosine-1-phosphate (S1P). Recruitment of immune cells into intestinal tissue was significantly impaired, namely neutrophils and effector Th1/Th17 T cells. ConclusionsLoss of AC reduced inflammation and impaired immune cell recruitment in chronic colitis. Targeting AC may serve as a promising therapeutic potential for patients with IBD by modulating immune cell sphingolipid metabolism. WHAT YOU NEED TO KNOWO_ST_ABSBACKGROUND AND CONTEXTC_ST_ABSAcid ceramidase expression is increased in immune cells in patients with inflammatory bowel disease, specifically in macrophages. NEW FINDINGSWe determined that loss of acid ceramidase (AC) in macrophages, but not neutrophils, impairs inflammatory functions in vitro, and that loss of AC in myeloid cells partially protects from spontaneous colitis in vivo by reducing immune cell recruitment into intestinal tissue. LIMITATIONSThe IL10 knockout model of colitis exhibits highly variable onset and severity of disease, which may be challenging to distinguish the extent of protection. CLINICAL RESEARCH RELEVENCEThis study identifies AC as a promising therapeutic target for treating inflammatory bowel disease. BASIC RESEARCH RELEVENCEThis study contributes to our understanding of the role that AC plays in inflammation within immune cells, specifically myeloid cells. Additionally, this study underscores the role of immune cell sphingolipid metabolism in inflammatory bowel disease. LAY SUMMARYLoss of acid ceramidase in myeloid cells protects from chronic colitis by decreasing inflammation, altering immune cell function, and impairing the recruitment of effector immune cells to the colon.

physiology↗