Mammary inflammation around parturition appeared to be attenuated by consumption of fish oil rich in n-3 polyunsaturated fatty acids

Abstract

Background: Mastitis endangers the health of domestic animals and humans, and may cause problems concerning food safety. It is documented that n-3 polyunsaturated fatty acids (PUFA) play significant roles in attenuating saturated fatty acids (SFA)-induced inflammation. This study was therefore conducted to determine whether mammary inflammation could be affected by consumption of diets rich in n-3 PUFA.

Methods: Forty-eight rats after mating began to receive diets supplemented with 5% fish oil (FO) or 7% soybean oil (SO). Blood and mammary tissue samples (n = 6) at day 0 and 14 of gestation and day 3 postpartum were collected 9 hours after intramammary infusion of saline or lipopolysaccharide (LPS) to determine free fatty acids (FFA) concentration and FA composition in plasma and inflammation mediators in mammary tissues.

Results: At day 14 of gestation and day 3 postpartum, the FO-fed rats had lower plasma concentrations of C18:2n6, C20:4n6, total n-6 PUFA and SFA, and higher plasma concentrations of C20:5n3 and total n-3 PUFA than the SO-fed rats. Plasma C22:6n3 concentration was also higher in the FO-fed than in the SO-fed rats at day 3 postpartum. Compared with the SO-fed rats, the FO-fed rats had lower mammary mRNA abundance of xanthine oxidoreductase (XOR) and protein level of tumor necrosis factor (TNF)-α, but had higher mammary mRNA abundances of interleukin (IL)-10 and peroxisome proliferator-activated receptor (PPAR)-γ at day 14 of gestation. Following LPS infusion at day 3 postpartum, the SO-fed rats had increased plasma concentrations of FFA, C18:1n9, C18:3n3, C18:2n6 and total n-6 PUFA, higher mammary mRNA abundances of IL-1β, TNF-α and XOR but lower mammary mRNA abundance of IL-10 than the FO-fed rats.

Conclusions: Mammary inflammation around parturition appeared to be attenuated by consumption of a diet rich in n-3 PUFA, which was associated with up-regulated expression of IL-10 and PPAR-γ.

Figure 1

Plasma FFA concentration of rats fed different diets at different reproductive stages. Plasma FFA concentration was determined by ELISA using plasma collected from rats fed the SO or FO diet at day 0 of gestation, day 14 of gestation and day 3 postpartum. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 2

Relative mRNA abundances of rats fed different diets at different reproductive stages. mRNA abundances of IL-8 (A), XOR (B), IL-10 (C) and PPAR-γ (D) was determined by RT-PCR with mammary tissues collected from rats fed the SO or FO diet at day 0 and 14 of gestation. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 3

Immunohistochemical localization of IL-1β in udder of rats fed different diets at different reproductive stages. The microphotograph from one rat with the positive primary IL-1β antibody was visualized with DAB reaction. The area positive for IL-1β in mammary tissues of rats fed the SO diet (B) or FO diet (C) at day 0 and 14 of gestation was quantified by Easy Image 3000 software. IL-1β production is presented as the average percentage of the positively stained areas (A). Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 4

Immunohistochemical localization of TNF-α in udder of rats fed different diets at different reproductive stages. The microphotograph from one rat with the positive primary TNF-α antibody was visualized with DAB reaction. The area positive for TNF-α in mammary tissues of rats fed the SO diet (B) or FO diet (C) at day 0 and14 of gestation was quantified by Easy Image 3000 software. TNF-α production is presented as the average percentage of the positively stained areas (A). Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 5

Histopathology of mammary glands of rats fed different diets at different reproductive stages. Hematoxylin and eosin stained slides were made with mammary tissues collected from rats fed the SO diet (B) or FO diet (C) at day 0 and 14 of gestation. PMN prevalence (A) in alveoli was estimated by using light microscopic (Olympus BH2, Japan) analysis at a magnification of 400×. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 6

Plasma FFA concentration of rats fed different diets and challenged with different stimulus. Plasma FFA concentration was determined by ELISA using plasma collected from rats fed the SO or FO diet and challenged with saline or LPS. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 7

Relative mRNA abundances of rats fed different diets and challenged with different stimulus. mRNA abundances of IL-1β (A), TNF-α (B), XOR (C), IL-10 (D) and PPAR-γ (E) was determined by RT-PCR with mammary tissues collected from rats fed the SO or FO diet and challenged with saline or LPS. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).

Figure 8

Histopathology of mammary glands of rats fed different diets and challenged with different stimulus. Hematoxylin and eosin stained slides were made with mammary tissues collected from rats fed the SO diet (B) or FO diet (C) and challenged with saline or LPS. PMN prevalence (A) in alveoli was estimated by using light microscopic (Olympus BH2, Japan) analysis at a magnification of 400×. Statistics are shown as means ± SE. Statistics with no common letters differ significantly (P < 0.05).