Bovine Endometrial Epithelial Cells Scale Their Pro-inflammatory Response In vitro to Pathogenic Trueperella pyogenes Isolated from the Bovine Uterus in a Strain-Specific Manner

Abstract

Among different bacteria colonizing the bovine uterus, Trueperella pyogenes is found to be associated with clinical endometritis (CE). The ability of cows to defend against T. pyogenes infections depends on the virulence of invading bacteria and on the host's innate immunity. Therefore, to gain insights into bacterial factors contributing to the interplay of this host pathogen, two strains of T. pyogenes were included in this study: one strain (TP2) was isolated from the uterus of a postpartum dairy cow developing CE and a second strain (TP5) was isolated from a uterus of a healthy cow. The two strains were compared in terms of their metabolic fingerprints, growth rate, virulence gene transcription, and effect on bovine endometrial epithelial cells in vitro. In addition, the effect of the presence of peripheral blood mononuclear cells (PBMCs) on the response of endometrial epithelial cells was evaluated. TP2, the strain isolated from the diseased cow, showed a higher growth rate, expressed more virulence factors (cbpA, nanH, fimE, and fimG), and elicited a higher mRNA expression of pro-inflammatory factors (PTGS2, CXCL3, and IL8) in bovine endometrial epithelial cells compared with TP5, the strain isolated from the healthy cow. The presence of PBMCs amplified the mRNA expression of pro-inflammatory factors (PTGS2, CXCL3, IL1A, IL6, and IL8) in bovine endometrial epithelial cells co-cultured with live TP2 compared with untreated cells, especially as early as after 4 h. In conclusion, particular strain characteristics of T. pyogenes were found to be important for the development of CE. Furthermore, immune cells attracted to the site of infection might also play an important role in up-regulation of the pro-inflammatory response in the bovine uterus and thus significantly contribute to the host-pathogen interaction.

Microbial communities and inflammatory response in the endometrium differ between normal and metritic dairy cows at 5–10 days post-partum

“…Nevertheless, the biopsy samples from the metritic cows seem to form a subgroup of the swab metritic communities. This observation can be explained by the different tissue penetrating abilities certain bacteria possess, by which only some members of the metritic bacterial communities found in the lumen and barrier epithelium can actually invade the uterine wall and cause disease [ 20 , 61 , 62 ].…”

Influence of intrauterine administration of Lactobacillus buchneri on reproductive performance and pro-inflammatory endometrial mRNA expression of cows with subclinical endometritis

“…In a former study was shown that the strain L. buchneri DSM 32407 did neither influence the viability of endometrial epithelial cells nor provoke a pro-inflammatory response in contrast to other Lactobacillus strains 16 . Pathogenic ( Trueperella pyogenes ) or potential pathogenic strains ( Bacillus pumilus ) caused cell death even within 16–24 h 33 , 34 . Therefore, the strain L. buchneri DSM 32407 seems suitable to serve as a probiotic strain within the bovine uterus to modulate the immune response and the reproductive performance of dairy cows.…”

Nisin reduces uterine inflammation in rats by modulating concentrations of pro‐ and anti‐inflammatory cytokines

“…In the first passage, endometrial epithelial cells were cultured in 96‐well plates until their density reached 3 × 10 5 cells/mL in a humidified incubator at 37°C and 5% CO 2 for 24 hours to make the cells adjust to conditions. In the second passage, the uterine epithelial cell population was observed to reach a 100.0% pure epithelial cell population, the medium was removed again, and the endometrial epithelial cells were washed three times with PBS as described by Ibrahim et al After 72 hours of incubation, nisin and kanamycin were diluted with PBS to 6.22 μg/mL and 0.25 μg/mL, respectively, and suspended in an antibiotic‐free epithelial cellculture medium before being cocultured with the endometrial epithelial cells and S aureus . ELISA kit and RT‐PCR were used to determine the cytokine levels of the uterine epithelial cell culture supernatants (UEC).…”

“…The RT‐PCR conditions consisted of 95°C for 10 minutes (45 cycles), followed by 45 cycles at 95°C for 15 seconds, 20 seconds of annealing at a temperature specified for each gene, and a last step at 72°C for 30 seconds. This was followed by a melting curve program (50‐90°C) and a final cooling step at 40°C for 1.0 minutes …”