Date of Graduation
Davis College of Agriculture, Natural Resources and Design
Animal and Nutritional Sciences
Robert A Dailey
Scott A Bowdridge
Eugene E Felton
An immunological balance has to be established during pregnancy that protects the mother yet tolerates the semi-allogenic fetus. To understand the innate immune response during bacterial infections that may cause early embryonic loss, a lipopolysaccharide (LPS) treated sheep model was used. Two objectives of this study were to examine if omega-3 PUFAs in the form of supplementary whole flaxseed could reduce the inflammatory response to an LPS challenge and to examine if there is a differential immune response to LPS in Suffolk and Dorset ewes. A total of 3 experiments were conducted; two investigating the effect of supplement and one investigating the effect of breed. Estrus was synchronized by CIDR insertion for 5 days, followed by 20 mg of PGF 2alpha at CIDR withdrawal. Early pregnant ewes received via the jugular vein, either phosphate buffered saline (PBS) (3 ml) or LPS (2.5 mug/kg). Blood was collected via jugular venipuncture at hour: 0, 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 5, 6, 9, 12, and 24. Whole blood samples were used to determine white blood cell counts (WBCs) before centrifugation to collect white blood cells for RNA extraction. Rectal temperature and change in behavior/physical (lethargy, coughing, nasal discharge, absence of eating) appearance were recorded hourly. Real-time PCR was performed for expression of cytokines (CXCL8, IL6, TNFalpha, IFNgamma, IL-10, and TGFbeta), receptors (TLR4, MRC1), enzymes (COX2, SOD2), transcription factors (NF-kappaB, PPARgamma, and Foxp3) and complement component 3. In all experiments, temperature increased in response to LPS, peaking at hour 4 before returning to normal by hours 6-9; WBCs dropped by hour 1 before returning to normal by hours 6-9. In trial 1 of the supplement study (flaxseed versus a control supplement) (Dorset ewes flaxseed + LPS n=3; flaxseed + PBS n=3; control + LPS n=5; control + PBS n=5), LPS increased haptoglobin and cortisol levels and affected gene transcription of CXCL8, IFN?, TLR4, MRC1, SOD2, Foxp3 (by hour), and C3.There was a diet effect with regard to cortisol and gene expression of CXCL8, and TLR4. There was a diet x LPS interaction with regard to temperature, WBCs (by hour), haptoglobin, serum amyloid A and gene expression of CXCL8, IL-6, and TLR4. In trial 2 of the supplement study (Dorset ewes flaxseed + LPS n=11, flaxseed + PBS n=10, control + LPS n=11, control + PBS n=10), LPS increased haptoglobin and cortisol and affected gene expression of CXCL8, TLR4, MRC1, SOD2, PPARgamma, Foxp3, and C3. There was a diet effect on cortisol and gene expression of C3.There was a diet x LPS interaction with regard to temperature and cortisol. In the breed effects study (Dorset + LPS n=11; Dorset + PBS n=10; Suffolk + LPS n=16; Suffolk + PBS n=16), LPS increased cortisol and affected gene transcription of CXCL8, TLR4, MRC1, SOD2, PPARgamma, and C3. There was an effect of breed on temperature; haptoglobin, serum amyloid A (by hour), and cortisol levels; and gene transcription of IL-6, IFNgamma, IL-10, TLR4, COX2, SOD2, PPARgamma, Foxp3, and C3. There was a breed x LPS interaction on change in temperature from hour 0, the frequency of behavior/physical changes; haptoglobin, serum amyloid A (by hour), and cortisol; and gene transcription of IL-6 and C3. Pregnancy status was assessed at 25 dpc with transrectal ultrasound and progesterone was measured in plasma samples. In trial 1 of the supplement study, flaxseed increased progesterone but it did not differ between groups in trial 2. There were no differences in progesterone between the breeds tested. The number of ewes that lambed in each treatment group was not different in any of the experiments. In summary, acute infections may cause embryonic loss by shifting the environment to be pro-inflammatory. There was no clear benefit of supplementary o-3 PUFAs in reducing the inflammatory response. Suffolk ewes had an elevated inflammatory response to LPS compared to Dorset ewes and may be more susceptible to embryonic loss in response to infection.
Hadfield, Jessalyn Marie, "Characterization of the Immune Response to Lipopolysaccharide in Early Pregnant Ewes as a Model to Study Bacterial Infection Induced Embryonic Loss" (2017). Graduate Theses, Dissertations, and Problem Reports. 5731.