Date of Award

2015

Document Type

Thesis

First Advisor

Worku, Dr. Mulumebet

Abstract

This study was conducted to evaluate the effect of a water extract of Sericea Lespedeza (SL), the lipid raft inhibitor NYS and known pathogen associated molecular patterns (LPS and PGN) on gene activation in ruminant (cow, sheep and goat) blood. Sericea Lespedeza, a high tannin containing legume has been shown to be a useful component of feed for control of gastro intestinal nematodes in ruminants. Blood was collected from four adult female Holstein Friesian cows (N=4), Spanish x Boer goats (N=4) and St Croix sheep (N=4) and incubated with 100 ng /mL-1 of SL in the presence and absence of LPS, PGN or NYS. Whole blood was also incubated with 100 ng mL-1 ng/ml of LPS, PGN or NYS without addition of SL. Phosphate-buffered saline (PBS) served as negative control. Total protein concentration, migration pattern, secretion of WNT5a, TLR2 and prostaglandin E2 alpha in plasma was also determined. Total RNA was isolated from cells using Trizol. Samples with RNA integrity number >7 were used for cDNA synthesis. Transcription of TLR2, TLR4, WNT, FZD, β catenin, TNFα and GAPDH was evaluated in cow, sheep and goat blood. The effect of Nystatin on expression of 84 genes on the cow WNT signaling pathway and human innate and adaptive immunity arrays was assessed in cow blood using real time PCR. Fold change in transcript abundance was calculated using the Livak method. Agarose gel electrophoresis and ethidium bromide staining was used to visualize amplicons. TLR2 and FZD were up regulated in response to PAMPS. Species specific gene expression and modulation by PAMPS was observed. Water extracts of SL reduced PGE2a in sheep and cows. Transcription and translation of genes involved in innate and adaptive immunity and the WNT signaling pathway in ruminant blood is responsive to diverse PAMPS and can be modulated by SL.

Share

COinS