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Primary intestinal epithelial cells are a powerful tool to study tight junction protein function and barrier integrity of epithelial cells. However, primary cells are costly, difficult to isolate, have a finite ability to replicate, and require the usage of live tissue. An alternative to primary cells is immortalized cell lines, which are cost effective, easily cultured, stored, and maintain the physiology of primary cells. Several immortalized lines are commercially available, yet there remains a need for cell lines for livestock research, specifically bovine. Here we report an immortalized bovine intestinal epithelial cell (IBEC) line was established by transfection with exogenous human telomerase reverse transcriptase ( hTERT ). Primary bovine intestinal epithelial cells (PBECs) were isolated from a 2-day old mixed breed calf. PBECs were transfected with hTERT to create an IBEC line. The transfected cells were positive for intestinal epithelial markers while testing negative for fibroblast markers. There was no significant difference in IBEC morphology or physiology post immortalization. In Trans-Epithelial Electrical Resistance (TEER assays), PBECs and IBECs behaved similarly to intestinal porcine enterocyte cells (IPEC-J2) under treatment of Iipopolysaccharide and calcium butyrate. This newly-developed cell line will be an essential and useful tool for in vitro bovine and ruminant intestinal research.

Heat stress (HS) is a major environmental stressor primarily affecting swine performance through negative effects on intestinal health. Zinc and butyric acid supplementation help maintain intestinal integrity and barrier function, and has been shown to be beneficial to swine during stress conditions. We tested a novel formulation of zinc butyrate (ZnB) to study whether it has protective effects toward swine using pig intestinal epithelial cells (IPEC-J2) and in a grower swine HS trial. IPEC-J2 cells were grown either under an inflammatory challenge (Escherichia coli lipopolysaccharide) or HS (41.5 [degrees]C for 48 h) using Transwell plates. The tight junction integrity of the cells under various treatments, including ZnB, zinc sulfate, and calcium butyrate, was followed over a period of 36 to 48 h by measuring transepithelial electrical resistance (TER). During inflammatory challenge, ZnB-treated cells had the greatest TER (P < 0.05) at 36 h. When the cells were exposed to HS at 41.5 [degrees]C, ZnB-treated cells had similar TER to the cells incubated at 37.0 [degrees]C, indicating significant protection against HS. In the swine trial (two dietary treatments, control and an encapsulated form of 40% zinc butyrate [E-ZnB] in hydrogenated palm oil pearls, 12 pigs per treatment), grower gilts (35 [+ or -] 1 kg) were supplemented with E-ZnB for 24 d before being subjected to biphasic HS for 7 d, 30 to 32 [degrees]C for 8 h and 28 [degrees]C for 16 h, for a total duration of 56 h of HS. At the end of the HS phase, half the pigs were euthanized from each treatment (n = 6 per treatment), and growth performance was calculated. During the HS phase, average daily gain (ADG; 0.53 vs. 0.79 kg) and gain-to-feed ratio (G:F; 0.33 vs. 0.43) were greater in the E-ZnB group (P < 0.05). Although in vivo intestinal permeability increased during the HS phase (P < 0.05), no differences were observed in the present study for the intestinal health parameters measured including TER, villus height:crypt depth ratio, and in vivo and ex vivo intestinal permeability between the two treatment groups. In conclusion, results presented here demonstrate that E-ZnB supplementation during HS improves ADG and G:F in grower pigs. Although we could not measure any differences, the mode of action of butyric acid and zinc suggests that the performance improvements are related to improved intestinal health. Key words: butyric acid, heat stress, intestinal integrity, pigs, swine, zinc

Abstract Heat stress (HS) is a major environmental stressor primarily affecting swine performance through negative effects on intestinal health. Zinc and butyric acid supplementation help maintain intestinal integrity and barrier function, and has been shown to be beneficial to swine during stress conditions. We tested a novel formulation of zinc butyrate (ZnB) to study whether it has protective effects toward swine using pig intestinal epithelial cells (IPEC-J2) and in a grower swine HS trial. IPEC-J2 cells were grown either under an inflammatory challenge (Escherichia coli lipopolysaccharide) or HS (41.5 °C for 48 h) using Transwell plates. The tight junction integrity of the cells under various treatments, including ZnB, zinc sulfate, and calcium butyrate, was followed over a period of 36 to 48 h by measuring transepithelial electrical resistance (TER). During inflammatory challenge, ZnB-treated cells had the greatest TER (P < 0.05) at 36 h. When the cells were exposed to HS at 41.5 °C, ZnB-treated cells had similar TER to the cells incubated at 37.0 °C, indicating significant protection against HS. In the swine trial (two dietary treatments, control and an encapsulated form of 40% zinc butyrate [E-ZnB] in hydrogenated palm oil pearls, 12 pigs per treatment), grower gilts (35 ± 1 kg) were supplemented with E-ZnB for 24 d before being subjected to biphasic HS for 7 d, 30 to 32 °C for 8 h and 28 °C for 16 h, for a total duration of 56 h of HS. At the end of the HS phase, half the pigs were euthanized from each treatment (n = 6 per treatment), and growth performance was calculated. During the HS phase, average daily gain (ADG; 0.53 vs. 0.79 kg) and gain-to-feed ratio (G:F; 0.33 vs. 0.43) were greater in the E-ZnB group (P < 0.05). Although in vivo intestinal permeability increased during the HS phase (P < 0.05), no differences were observed in the present study for the intestinal health parameters measured including TER, villus height:crypt depth ratio, and in vivo and ex vivo intestinal permeability between the two treatment groups. In conclusion, results presented here demonstrate that E-ZnB supplementation during HS improves ADG and G:F in grower pigs. Although we could not measure any differences, the mode of action of butyric acid and zinc suggests that the performance improvements are related to improved intestinal health.