Abstract
Here, we describe the use of monolayers of intestinal epithelial cells derived from intestinal organoids and transcriptomics to investigate the direct effects of dietary protein sources on epithelial function. Mechanically dissociated 3D organoids of mouse duodenum were used to generate a polarized epithelium containing all cell types found in the tissue of origin. The organoid-derived cell monolayers were exposed to 4% (w/v) of ‘undigested (non-hydrolysed)-soluble’ fraction of protein sources used as feed ingredients [soybean meal (SBM) and casein], or alternative protein sources (spray dried plasma protein, and yellow meal worm), or controls for 6 h prior to RNA isolation and transcriptomics. All protein sources altered expression of unique biological processes in the epithelial cells. Exposure of intestinal organoids to SBM downregulated expression of retinol and retinoid metabolic processes as well as cholesterol and lipid biosynthetic pathways, consistent with the reported hypotriglyceridaemic effect of soy protein in vivo. These findings support the use of intestinal organoids as models to evaluate complex interactions between dietary ingredients and the intestinal epithelium and highlights some unique host effects of alternative protein sources in animal feed and potentially human food.Graphical abstractSchematic representation of the study. 3-dimensional organoids were generated from mouse duodenum (1). The organoids were subsequently dissociated into single cells (2) and grown as 2-dimensional polarised monolayers (3). Polarized monolayers of organoid cells were exposed to different protein sources [CAS, SBM, SDPP, YMW, or medium control (MC)] for 6 h (4) and further processed for imaging (5) gene expression (6), and biochemical assays (7), to investigate the effects of undigested protein sources on the duodenal epithelium.
Highlights
There is much interest in alternative sources of protein for animal feed, driven by the expected growth in the human population and increasing demand for animal protein
The genome-wide transcriptomics data revealed expression of other genes associated with specific cell lineages of intestinal epithelium including columnar base cells (CBC), and enteroendocrine cells (EEC)(Fig. S2)
Different undigested sources of dietary protein have distinct biological effects on organoids Organoid monolayers were exposed to different undigested protein sources (4% w/v, soybean meal (SBM), CAS, spray-dried plasma protein from porcine blood (SDPP), and yellow mealworm larvae (YMW)) or Dulbecco’s modified Eagle medium (DMEM) as a medium control (MC) for 6 h
Summary
There is much interest in alternative sources of protein for animal feed, driven by the expected growth in the human population and increasing demand for animal protein. Cheaper sources of feed protein are being explored to sustain increased animal production. Alternative sources of protein include insect larvae and blood plasma. In vitro models commonly used to study intestinal responses to luminal. To overcome these issues, we used three-dimensional (3D) organoid cultures from isolated crypts of the duodenum [5, 6]. Intestinal organoids can be generated within 2 weeks, maintained for > 2 years in culture or cryopreserved, and when differentiated contain all the epithelial cell lineages found at the location of origin [7, 8]. We and others have described methods for growing polarised monolayers of 3D organoid cells on semipermeable membrane supports, enabling apical exposure to
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