Antioxidant and anti-inflammatory activities of pomegranate (Punica granatum) on Eimeria papillata-induced infection in mice

Abstract

Coccidiosis is the most prevalent disease causing widespread economic loss, especially in poultry farms. Here, we investigated the effects of pomegranate peel extract (PPE) on the outcome of coccidiosis caused by Eimeria papillata in mice. The data showed that mice infected with E. papillata and treated with PPE revealed a significant decrease in the output of oocysts in their faeces by day 5 p.i. Infection also induced inflammation and injury of the jejunum. This was evidenced (i) as increases in reactive oxygen species, (ii), as increased neutrophils and decreased lymphocytes in blood (ii) as increased mRNA levels of inducible nitric oxide synthase (iNOS), Bcl-2 gene, and of the cytokines interferon gamma (IFN-γ), tumour necrosis factor-α (TNF-α), and interleukin-1β (IL-1β), and (iv) as downregulation of mucin gene MUC2 mRNA. All these infection-induced parameters were significantly altered during PPE treatment. In particular, PPE counteracted the E. papillata-induced loss of the total antioxidant capacity. Our data indicated that PPE treatment significantly attenuated inflammation and injury of the jejunum induced by E. papillata infections.

Figure 1

Different developmental stages of E. papillata in the jejunum of a mouse on day 5 p.i. (a, b, and c). Meront (M), microgamont (Mi), macrogamont (Ma), and developing oocyst (DO). See Table 1 for quantification. Bar: 25 μm.

Figure 2

Reactive oxygen species (ROS) levels in jejunal homogenates of mice. Values are means ± SD. a: significant change at P < 0.01 with respect to noninfected (−PPE) mice. a, b: significant change at P < 0.01 with respect to infected (−PPE) mice.

Figure 3

Total antioxidant capacity levels in jejunal homogenates of mice. Values are means ± SD. a: significant change at P < 0.01 with respect to noninfected (−PPE) mice. a, b: significant change at P < 0.01 with respect to infected (−PPE) mice.

Figure 4

Immunohistochemical localization of Bcl-2 in the jejuna of mice. (a) Noninfected jejunum. (b) Noninfected PPE treated mouse jejunum. (c) E. papillata infected jejunum with increased number of Bcl-2 positive cells. (d) Infected treated mouse with decreased number of Bcl-2 positive cells. Bar: 25 μm.

Figure 5

Quantitative RT-PCR analysis of Bcl-2 mRNA in the jejunum. Expression was analysed on day 5 p.i., normalized to 18S rRNA signals, with the relative expression given as fold increase compared to the uninfected control mice. Values are means ± SD. a: significant change at P < 0.01 with respect to noninfected (−PPE) mice. a, b: significant change at P < 0.01 with respect to infected (−PPE) mice.

Figure 6

Quantitative RT-PCR analysis of MUC2 mRNA in the jejunum. Expression was analysed on day 5 p.i., normalized to 18S rRNA signals, with the relative expression given as fold increase compared to the uninfected control mice. Values are means ± SD. a: significant change at P < 0.01 with respect to noninfected (−PPE) mice. a, b: significant change at P < 0.01 with respect to infected (−PPE) mice.

Figure 7

Quantitative RT-PCR analysis of TNF-α, iNOS, IFN-γ, and IL-1β in the jejunum. Expression was analysed on day 5 p.i., normalized to 18S rRNA signals, and relative expression is given as fold increase compared to the uninfected control mice. Values are means ± SD. a: significant change at P < 0.01 with respect to noninfected (−PPE) mice. a, b: significant change at P < 0.01 with respect to infected (−PPE) mice.