Nutritional Stimulation by In-ovo Feeding Modulates Cellular Proliferation and Differentiation in the Small Intestinal Epithelium of Chicks

Abstract

BackgroundNutritional stimulation of the small intestine (SI) of chick embryos can be conducted by enriching the amniotic fluid with nutrients via the in-ovo feeding (IOF) methodology. The impact of IOF of specific nutrients on cellular proliferation and differentiation within the multipotent (MP) and differentiated cell niches of the developing SI have not yet been characterized. In the study, we examined the effects of IOF of 1% glutamine (IOF-Gln), 1% leucine (IOF-Leu) and 0.4% NaCl (IOF-NaCl), compared to non-injected controls, on the proportions and localizations of MP, progenitor and differentiated cells within the SI epithelium of peri-hatch chicks. MP, progenitor and differentiated cells were located and quantified in jejunum sections of all treatment groups at E17 and at E19, and post-hatch at days 0, 1, 3 and 7, by immunofluorescence of Sox9 and PCNA, in-situ hybridization of Lgr5 and PepT1 and histochemical goblet cell staining.ResultsAt E19, 48 h post IOF, the effects of IOF treatments, in comparison to Control embryos, were as follows: total cell counts increased by 40%, 33% and 19%, and MP cell counts increased by 52%, 50% and 38%, in IOF-Gln, IOF-Leu and IOF-NaCl embryos, respectively. Only IOF-Gln embryos exhibited a significant, 36% increase in progenitor cell counts. Lgr5+ stem cell localizations shifted to villus bottoms in IOF-treated embryos. The differentiated, PepT1+ region of the villi was 1.9 and 1.3-fold longer in IOF-Gln and IOF-Leu embryos, respectively, while goblet cell densities decreased by 20% in IOF-Gln embryos. Between hatch and D7, crypt and villi epithelial cell counts were significantly higher IOF-Gln chicks, compared to Control chicks (P<0.05).ConclusionsIOF promotes pre-hatch SI maturation through increased proportions and enhanced compartmentalization of the MP and differentiated cell niches. IOF of glutamine stimulates SI maturation to a greater extent than leucine and NaCl, and elicits further expansions of the crypt and villus epithelium during the first week post-hatch. These findings shed light on the link between primary nutritional stimulation and cellular maturation within the SI epithelium.

Intra-Amniotic Administration of l-Glutamine Promotes Intestinal Maturation And Enteroendocrine Stimulation In Chick Embryos

“…Though intra-amniotic administration of NaCl elicited some bene cial effects on enterocyte maturation, these effects were limited to post-hatch morphometric parameters. Since amniotic enrichment with NaCl was previously shown to promote proliferation in the small intestinal epithelium through activation of ion-dependent nutrient transporters during amniotic uid ingestion [19], It may be possible that NaCl promoted earlier morphological maturation of enterocytes, but did not stimulate L-cell signaling, and thus failed to enhance absorptive and barrier functions.…”

“…Recently, intra-amniotic administration of l-glutamine (Gln) was found to promote proliferation and differentiation in the small intestinal epithelium of chick embryos, resulting in higher quantities of enterocytes during the rst week post-hatch [19]. However, the effects of Gln on enterocyte morphology and function during this period, and the mechanism by which Gln stimulates small intestinal maturation, have not yet been characterized.…”