Ileum tissue single-cell mRNA sequencing elucidates the cellular architecture of pathophysiological changes associated with weaning in piglets

Abstract

Background In mammals, transitioning from sole milk uptake to the intake of solid feed results in dramatic developmental changes in intestinal function and immunological status. In fact, weaning stress is often accompanied by intestinal inflammatory processes. To develop effective intervention strategies, it is necessary to characterize the developmental pattern and immune response that occurs on weaning, as we have done in this study for piglets. Results To comprehensively delineate cell heterogeneity in ileum tissues and the underlying mechanisms in weaning-induced intestinal inflammation of piglets, we have analyzed the transcriptomes of 42,149 cells from ileum mucosa of normally suckling and post-weaned piglets. There were 31 cell subtypes including epithelial, stromal, and immune cells. A bifurcating trajectory was inferred to separate secretory and absorptive lineages. Integrated cross-species datasets showed well-conserved cellular architectures and transcription signatures between human and pig. Comparative analyses of cellular components, cell–cell communications, and molecular states revealed that T cell subpopulations were significantly altered in weaned piglets. We found that T helper (Th) 17 functional plasticity across changes in the cytokine milieu and the enrichment of granzyme B (GZMB)-expressing cytotoxic T cells potentially exacerbate mucosal inflammation via mitochondrial dysfunction in epithelial cells. Conclusions Our work has elucidated the single-cell molecular characteristics of the piglet ileum before and after weaning. We have provided an atlas that portrays the landscape of the intestinal pathophysiological inflammatory process associated with weaning, finding a level of conservation between human and pig that support the use of piglets as a model for human infants.

Intraepithelial lymphocytes in the pig intestine: T cell and innate lymphoid cell contributions to intestinal barrier immunity

“…-Human intestinal physiology, nutrition, and microbiota more similar to pigs than mice (188)(189)(190)(191)(192)(193)(194) -Proportions of gdTCR T-IELs, CD8ab + abTCR T-IELs, and CD8aa + abTCR T-IELs in human intestine more similar to pigs than mice (1,7,25,29,66,67,(195)(196)(197) -Transcriptional similarities between human and porcine intestinal immune and epithelial cells identified via high-resolution transcriptomics (57,(198)(199)(200) -Biomedical applications using pig models have cross-utility to improve pig health and global food security Frontiers in Immunology frontiersin.org assess transcriptomes of epithelial cells in duodenum, jejunum, and ileum of the porcine small intestine, finding again transcriptional similarities between humans and pigs for the majority of epithelial cells (200). Thus, high-resolution transcriptomic analyses support transcriptional conservation between immune and epithelial cells of human and porcine intestine, but scRNA-seq studies have so far been limited to only the small intestine.…”

“…In addition to phenotyping a limited number of cellular markers at the protein level to identify proportions of specific IEL populations, high-resolution transcriptional studies suggest pig-to-human similarities of intestinal cells. A recent scRNA-seq study identified conserved expression of several cell type-canonical genes between pig and human epithelial and immune cells that were sparsely expressed in mice ( 198 ). An additional scRNA-seq study defining lymphocyte subsets (including T cells and ILCs) at enhanced resolution also identified transcriptional similarities, similar localization patterns, and conserved inferred functions of lymphocytes from porcine and human ileum ( 57 ).…”

Development of an intestinal epithelial cell line and organoids derived from the same swine and characterization of their antiviral responses

“…In addition, our group has previously established porcine intestinal epitheliocyte (PIE) cells to study the immune responses of swine intestinal epithelial cells via toll-like receptors (TLRs) and reported on their innate immune responses [11]. Considering that the intestinal tract is composed of various cell types, including absorptive epithelial cells, goblet cells, Paneth cells, enteroendocrine cells, BEST4 cells, and intestinal stem cells [12,13], it may be difficult to accurately reproduce the response and function of the intestinal tract when relying solely on cell lines composed of a single cell type. Intestinal organoids, which can self-organize in three dimensions and reproduce the heterogeneity and behavior of the intestinal mucosa, are attractive tools to study in vitro the complex interactions between the different cell types and foreign agents, such as commensal or pathogenic microorganisms [14,15].…”

Construction of a multi-tissue cell atlas reveals cell-type-specific regulation of molecular and complex phenotypes in pigs

“…The next step is to explore the cell-type-dependent biological consequences of trait-associated variants as tissues contain numerous cell types 24 . Although some studies have conducted single-cell/nucleus RNA-seq (scRNA-seq and snRNA-seq) analyses in pigs, they primarily focused on elucidating the cellular heterogeneity and trajectories of lineage speci cation in a limited range of tissue types 25,26,27,28,29,30,31,32 . The cell-type-speci c biological impacts of genetic variants on complex traits by integrating single-cell RNA-sequencing with large-scale pig genetics data still need to be explored.…”