Date of Graduation

5-2020

Document Type

Thesis

Degree Name

Bachelor of Science in Agricultural, Food and Life Sciences

Degree Level

Undergraduate

Department

Poultry Science

Advisor/Mentor

Dridi, Sami

Committee Member/Reader

Rosenkrans, Charles F. Jr.

Committee Member/Second Reader

Greene, Nicholas

Abstract

Heat stress hampers gut health by impeding its ability to absorb nutrients which leads to a lower feed efficiency and sicker birds. Therefore, there is a critical need to identify mechanism-based strategies to alleviate these effects. Polynesian poultry farmers have been supplementing broiler feed with Morinda Citrifolia (Noni), a medicinal plant high in anti-oxidants, to address this problem. The purpose of the present study was to evaluate the effect of Morinda Citrifolia (Noni)-supplementation on the expression of tight junction proteins. Male Broilers (Cobb 500, 3 wks, n=480) were subjected to 2 environmental conditions (TN, 24°C vs. HS, 35°C, 1 wk exposure), and fed two diets (control vs. 0.2% Noni) in a 2x2 factorial design. Chickens received water and feed ad libitum. Functional in vitro studies were conducted using IPEC-J2 and primary chicken gut epithelial cells. Once cells reached 80% confluence, cells were exposed to HS conditions (45°C) for 2h and 30 min. Control cells were maintained at 37°C. The expression of target genes and proteins were determined by quantitative real-time PCR using 2-ΔΔCT method and Western blot, respectively. A significant increase (P < 0.05) in the expression of either heat-shock protein 70 or 90 was observed in both in vivo and in vitro studies to indicate stress status. One-week HS broilers experienced a downregulation of ZO-2 and occludin tight junction protein expression. Interestingly, HS broilers fed a Noni diet had upregulated ZO-1, ZO-2 gene expression, but both ZO-1 and ZO-2 protein expression were downregulated in the HS Noni-fed broilers. ZO-1 and ZO-2 are essential scaffolding proteins that are key components of tight junction proteins. They allow tight junction proteins to bind to specific location between cells to function as a permeability barrier. Their absence could be responsible for an increase in gut permeability. Noni’s effect on interleukin expression under TN and HS conditions was also measured in which there was a tendency (yet not significant) to decrease IL-18 gene expression and significant reduction in IL-10 gene expression in both TN and HS broilers fed a Noni diet. No decreases in tight junction protein expression was visualized via western blotting in HS IPEC-J2 cells. Nonetheless, there is clear fragmentation of Claudin 1, Occludin, and Claudin 5 tight junctions which was visualized via immunofluorescence staining in heat stressed cells. When quercetin was added into the media of IPEC-J2 cells, ZO-2 protein expression was severely downregulated in cells regardless of containing quercetin. In the primary gut epithelial cells, there were no changes in any protein or genes except for a downregulation in Cadherin gene expression in heat stressed cells. Further studies to confirm and expand the results obtained in this study as well as the further characterization of the primary gut epithelial cells deserves further investigation.

Keywords

Noni; Heat stress; tight junction proteins; chicken; intestinal epithelial cells

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