Time-dependent mRNA expression of selected pro-inflammatory factors in the endometrium of primiparous cows postpartum

Abstract

Background: Inflammatory processes and infections of the uterine wall must be accepted as a physiological event in dairy cows after calving. This might result in clinical or subclinical endometritis which is assumed to impair reproductive performance in the current lactation. Several cytokines and acute phase proteins have been discussed as local and systemic mediators of these inflammatory processes. The aim of the present study was to investigate the endometrial mRNA expression of the chemokine CXC ligand 5 (CXCL5), interleukin 1β (IL1B), IL6, IL8, tumour necrosis factor alpha (TNF), prostaglandin-endoperoxide synthase 2 (PTGS2) and haptoglobin (HP) in the postpartum period.

Methods: Endometrial samples were obtained from primiparous cows (n = 5) on days 10, 17, 24, 31, 38 and 45 postpartum (pp) using the cytobrush technique. Cytological smears were prepared from cytobrush samples to determine the proportion of polymorphonuclear neutrophils (PMN). Total RNA was extracted from endometrial samples, and real-time RT-PCR was performed.

Results: A time-dependent mRNA expression of the investigated factors was found for the course of the postpartum period. In detail, a significantly higher expression of these factors was observed on day 17 pp compared to day 31 pp. Furthermore, the proportion of PMN peaked between days 10-24 pp and decreased thereafter to low percentages (< 5%) on day 31 pp and thereafter. In addition, CXCL5, IL1B, IL8 and HP mRNA expression correlated significantly with the proportion of PMN (P < 0.05). A significantly higher CXCL5, IL1B, IL6, IL8, PTGS2 and TNF mRNA content was observed in samples from cows with an inflamed endometrium compared with samples from cows with a healthy endometrium (P < 0.05).

Conclusions: These results show that inflammatory cytokines and acute phase proteins are expressed in the bovine endometrium in a time-related manner during the postpartum period, with a significant expression peak on day 17 pp as a possible mucosal immune response in the uterus. The evaluation of the expression patterns of such candidate genes may reveal more information than only determining the percentage of PMN to judge the severity of an inflammation.

Figure 1

PMN content in endometrial samples. PMN content in bovine endometrial samples collected during the postpartum period on days 10, 17, 24, 31, 38 and 45, respectively (n = 5 animals). Sample from cow 4 on day 17 pp is missing due to technical reasons.

Figure 2

mRNA expression of CXCL5, IL1B, IL6 and IL8 in endometrial samples. mRNA expression of CXCL5 (A), IL1B (B), IL6 (C) and IL8 (D) in bovine endometrial samples collected during the postpartum period on days 10, 17, 24, 31, 38 and 45 pp, respectively (n = 5 animals). The contents of each mRNA are expressed as ratio relative to individual 18S rRNA values as an internal control. The asterisk indicates significant differences between day 17 pp and day 31 pp; P < 0.05.

Figure 3

mRNA expression of TNF, PTGS2 and HP in endometrial samples. mRNA expression of TNF (A), PTGS2 (B) and HP (C) in bovine endometrial samples collected during the postpartum period on days 10, 17, 24, 31, 38 and 45 pp, respectively (n = 5 animals). The contents of each mRNA were expressed as ratio relative to individual 18S rRNA values as an internal control. The asterik indicates significant differences between day 17 pp and day 31 pp; P < 0.05.