Regulation of Intestinal Development and Function in Early-Life Pigs

Abstract

The small intestine's plasticity allows for rapid structural and functional adaptions in response to external stimuli. The early postnatal phase is a critical developmental window, during which diet transitions and stress can have lasting effects on overall health and productivity. Attenuated growth performance and feed efficiency may exacerbate production losses and have significant economic implications on the swine industry. Historically, the industry used in-feed antibiotics as growth promoters, but their ban has left the need for nutritional or pharmacological strategies that mitigate intestinal inflammation. Additionally, knowledge gaps remain regarding how Wnt/ β-catenin and Notch signaling pathways influence intestinal remodeling during the early life of pigs. Collectively, these gaps highlight the need for targeted nutritional or molecular interventions during critical developmental phases during the early life of pigs. Topics examined herein focus on development changes of the gastrointestinal tract. The first study evaluated the effects of different concentrations of cannabidiol (CBD) supplementation on intestinal inflammation and function at weaning. Pre-weaned pigs received one of four treatments: low (10 mg/kg CBD, n=8), medium (25 mg/kg, n=8), high (50 mg/kg, n=8) and control (0 mg/kg, n=8) once a day for 5 days starting one day before weaning. The medium dose showed greater lactase (P = 0.003) and maltase (P = 0.042) activities in the jejunum, and increased lactase activity in the ileum (P = 0.020). Jejunal IL-β concentrations were greater in control pigs (P = 0.014) compared with other treatments, while mRNA abundance of intestinal integrity markers varied among treatment groups, with CLDN4 reduced in low pigs (P= 0.045), ZO-1 reduced across all treatments compared with controls (P=0.012), and CCL2 reduced in high pigs (P=0.018). CASP6 was elevated in low pigs (P = 0.012), and TNFα was reduced in medium pigs (P = 0.031). Morphology and goblet cell measurements had no differences. Overall, CBD supplementation had limited effects on intestinal inflammation and function in newly weaned pigs, but the medium dose showed the greatest changes in the jejunum. The second study focused on early life intestinal epithelial development by analyzing the duodenum, jejunum, and ileum of 70 pigs sampled at 0, 7, 20, 22, 25, and 28 days of age (n=10 pigs/day). Frozen tissues were assessed for brush border activity and found shifts in enzymatic activity coinciding with luminal content changes. Formalin- fixed tissues were used for histological analysis, cell phenotyping, and RNA in situ hybridization. Morphological analysis showed an in increased villus height and crypt depth in early life and decreased postweaning. Expression of Sox9 (duodenum, jejunum, ileum P <0.001) and Ki67 (duodenum P = 0.008; ileum P = 0.039) were increased at birth and postweaning reflecting intestinal growth and remodeling. Sucrase Isomaltase (duodenum P = 0.025; jejunum and ileum P <0.001) increased from birth to weaning, coinciding with changes in disaccharidase activity. Somatostatin (duodenum, jejunum, ileum P <0.001) was greatest at birth and at weaning, reflecting adaptions to changes in luminal contents. Goblet cells (duodenum, jejunum, ileum P <0.001) increased until weaning suggesting heightened mucus production in response to compromised barrier integrity. These results were supported by Ussing Chamber assays of fresh jejunal tissue. Reduced transepithelial resistance (jejunum P <0.001) combined with enhanced glucose (jejunum P <0.001) and glutamine (jejunum P <0.001) active transport at birth and postweaning described increased permeability and increased energy demands. Molecular signaling pathways, Wnt/β-catenin and Notch, were evaluated to better understand how stem cell proliferation and cell fate determination are regulated from birth to weaning. Wnt3 (duodenum P = 0.001) and β-catenin (duodenum, jejunum, ileum P <0.001) expression was increased at birth and postweaning, promoting stem cell proliferation to support intestinal remodeling. Notch-1 (duodenum and jejunum P <0.001; ileum P = 0.009) and Hes-1 (duodenum P = 0.007; jejunum P = 0.004; ileum P <0.001) expression increased at birth and postweaning, driving enterocyte differentiation. Together these findings highlight the rapid and coordinated intestinal remodeling pigs undergo during early life and the molecular pathways that drive intestinal development during early life of pigs, and potential targets to develop nutritional interventions that enhance nutrient utilization.