We have demonstrated that the pattern recognition toll-like receptors (TLR) 2&4 are upregulated during pregnancy in human cervical epithelium. We sought to determine whether oestradiol (E2) alters the immunological responses of cultured human ecto-cervical epithelial cells (HECECs) to the ligands of TLR2 (peptidoglycan, PGN) and TLR4 (lipopolysaccharides, LPS).
Methods and Material Fresh ectocervical tissues were collected from hysterectomy specimens, epithelia isolated and cultured after collagenase and trypsin digestion, cell purity being validated by immunofluorescence. The expression of TLRs 2&4, and E2 receptor genes (ERs α&β) were demonstrated by flow cytometry and RT-PCR respectively. HECECs were co-incubated with varying E2 concentrations (0.1, 1, 10, 100 nM) and PGN (50 μg/ml) or LPS (100 ng/ml) for 10 min, 2 hrs and 18 hrs. Cytokines were assayed in supernatant fluid by Cyto-Bead Array.
Result In response to PGN, HECECs co-incubated with E2 released lower levels of IFN-γ, IL-6 and IL-8, whilst after 18 hrs, RANTES expression increased to low E2 but decreased to high E2. In response to LPS, HECECs co-incubated with E2 demonstrated increased expression levels of IL-10, IL-12p70, RANTES and TNFα at 10 min and 2 hrs. IL-6 and IL-8 levels increased at 2 hrs but were suppressed or unchanged after 18 hrs. Markedly increased levels of RANTES were detected after 18 hrs, an effect obtunded by culturing HECECs in E2.
Discussion The complex immunomodulatory effects of E2 on cultured HECECs appear to vary with bacterial ligands and duration of exposure, suggesting direct non-genomic vs. genomic mechanisms. The resulting cytokine profiles may contribute to cervical remodelling during pregnancy.
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