Abstract
Endometritis is a prevalent reproductive disease in dairy cows, and is a superficial inflammation of the endometrium. S100 calcium-binding protein A4 (S100A4) is suggested to be implicated in the progression of inflammation. However, to our knowledge, no study has reported the changes of S100A4 during bovine endometritis. The objective of this study was to investigate S100A4 gene expression and protein levels in the uterus with endometritis in dairy cows. Vaginal mucus samples were collected for diagnosis of the severity degree of endometritis and the detection of S100A4 protein content. Blood samples and endometrial biopsies were collected and divided into the control (CN), mild endometrtis (M), and severe endometritis (S) groups according to the characteristics of the vaginal mucus type. The isolated bovine endometrial epithelial cells (BEECs) were challenged with E. coli (2 × 106 CFU/mL, 2 × 107 CFU/mL) or lipopolysaccharide (LPS, 3 and 10 μg/mL) as an inflammatory model. RT-qPCR was used to detect the gene expression levels of S100A4 and cytokines, including interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-10 (IL-10), and tumour necrosis factor-alpha (TNF-α), in tissues or cells. Enzyme-linked immunosorbent assay (ELISA) was used for S100A4 protein level detection in tissues, cells, cell supernatant, vaginal mucus, and serum samples. The results showed that S100A4 gene and protein levels decreased in bovine endometrium with endometritis and in E. coli- or LPS-stimulated BEECs. We failed to detect S100A4 in the cell supernatant, vaginal mucus, and serum samples. This study suggested that S100A4 is a pathogenesis-related protein of endometritis, and decreased expression of S100A4 may pave the way for the development of endometritis in dairy cows.
Highlights
As a result of milk accounting for a large proportion of the human diet, the health conditions of dairy cows have attracted great attention
The gene and protein expression of S100A4 in the classified tissue samples was detected, and the results showed that both the gene expression level and protein content of S100A4 were significantly decreased in endometrial tissues with mild or severe endometritis (Fig. 2) and negatively correlated with the severity of endometritis
Bovine endometrial epithelial cells (BEECs), TNF-α, IL-1β, IL-6, and IL-10 were upregualted significantly when BEECs were stimulated with E. coli (2×106 CFU/mL, 2×107 CFU/mL), with the exception of IL-6 levels after 2×106 CFU/mL E. coli stimulation (Fig. 3A-D)
Summary
As a result of milk accounting for a large proportion of the human diet, the health conditions of dairy cows have attracted great attention. Reproductive disease, especially endometritis, is one of the most important diseases compromising animal welfare and causing increased economic loss in the dairy industry [1]. Endometritis is highly prevalent in dairy cows and can result in other kinds of reproductive diseases, including pyometra, anovulation and pregnancy loss, as well as reduce productive performance [2, 3]. It has been reported that S100A4 promoted the progression of endometritis, while deficiency of S100A4 reduced the mRNA expression levels of IL-1β and TNF-α in the endometrium of mice [11]. To investigate the roles of S100A4 in endometritis of dairy cows, we determined the gene and protein expression of S100A4 in endometrial tissues, vaginal mucus, serum, E
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