Exfoliated epithelial cell transcriptome reflects both small and large intestinal cell signatures in piglets

Abstract

Assessing intestinal development and host-microbe interactions in healthy human infants requires non-invasive approaches. We have shown that the transcriptome of exfoliated epithelial cells in feces can differentiate breast and formula-fed infants and term and preterm infants. However, it is not fully understood which regions of the intestine that the exfoliated cells represent. Herein, the transcriptional profiles of exfoliated cells with that of the ileal and colonic mucosa were compared. We hypothesized that exfoliated cells in the distal colon would reflect mucosal signatures of more proximal regions of the gut. Two-d-old piglets (n=8) were fed formulas for 20 d. Luminal contents and mucosa were collected from ileum (IL), ascending (AC) and descending (DC) colon, mRNA was extracted and sequenced. On average, ~ 13,000 genes were mapped in mucosal tissues and ~10,000 in luminal contents. The intersection of detected genes between 3 mucosa regions and DC exfoliome indicated an approximately 99% overlap. On average, 49% of the genes in IL, AC and DC mucosa were present in the AC and DC exfoliome. Genes expressed predominantly in specific anatomic sites (stomach, pancreas, small intestine, colon) were detectable in exfoliated cells. Additionally, gene markers for all intestinal epithelial cell types were expressed in the exfoliome representing a diverse array of cell types arising from both the small and large intestine. Genes were mapped to nutrient absorption and transport and immune function. Thus, the exfoliome represents a robust reservoir of information in which to assess intestinal development and responses to dietary interventions.

Critical Steps for Human Gut Exfoliome RNA Profiling Analysis Using Non-Invasive Stool Samples

“…As limitations of this study, we should state that we did not have the results obtained validated since this was a pilot study; when this technique is used in populations, it would be advisable to validate the results with targeted RT-PCR of specific genes or other physiological biomarkers. Gene markers for all intestinal epithelial cell types were expressed in the exfoliome representing a diverse array of cells [21]. Nevertheless, we did not perform that confirmation by specific cell types in the present study.…”

“…More recently, a relationship was also established between the human intestinal exfoliome and microbiome [20]. The transcriptome of exfoliated epithelial cells in stool contains gene signatures from both small- and large-intestinal mucosa, affording a non-invasive approach to assess gut health and function [21] which may complement current gut evaluation tests. However, certain technical issues concerning sample collection, freezing procedure, and poly-A mRNA purification should be evaluated and standardized since they may affect the amount of human mRNA isolated, gene expression evaluation, and the cost of analysis.…”

Colonic Dysregulation of Major Metabolic Pathways in Experimental Ulcerative Colitis

“…However, several metabolomics studies indicate discrepancies of metabolite levels between blood/urine and colon biopsy samples [13,14,17,18]. It has been suggested that exfoliated cells in feces may serve as a noninvasive method to assess mucosal signatures for early detection/diagnosis of intestinal health [34]. In our study, we analyzed colonic tissue directly and employed a proteomics methodology to assess the alteration of metabolic pathways under colitis.…”

Integrated analysis of gut metabolome, microbiome, and exfoliome data in an equine model of intestinal injury

“…Further, host transcriptomic data was based on analysis of the equine exfoliome. We and others have used this approach in rodents [5], pigs [6], people [7], human neonates [8], and horses [9], and demonstrated that exfoliomic data mimic those in tissues but further validation of the exfoliomic methods in horses is warranted. Finally, the portion of our mechanistic hypothesis generated by evaluation of the exfoliome is based on the function of a small number of genes that are not master regulators of the pathways we identified.…”

“…One such approach is the use of exfoliomics. This platform has been utilized in rodents [ 5 ], pigs [ 6 ], adult humans [ 7 ], and human neonates [ 8 ]. We recently also validated this approach in horses [ 9 ].…”