Expression of retinoic acid-related orphan receptors and interleukin-17 in mice with vaginal candidiasis

Abstract

Objective To investigate the role of T helper 17 cells/interleukin-17 (Th17/IL-17) axis in the occurrence of vaginal candidiasis in mice. Methods A total of 120 female BALB/c mice were randomly and equally divided into Ei, En, Ci and Cn groups. Three days before vaginal inoculation, estrogen (Ei and En) groups and control (Ci and Cn) groups received subcutaneous injection of 0.05 mg estradiol and 0.1 ml sterilized soybean oil at the hind legs, respectively, and then the hormone treatment continued every other day until the end of experiment. Infected (Ei and Ci) groups and noninfected (En and Cn) groups were inoculated intravaginally with 10 μl (5 × 104 conidia) of Candida albicans suspension and 10 μl of sterilized phosphate-buffered saline, respectively. Ten mice were randomly selected from each group and sacrificed on day 3, 7 and 14 after inoculation. The intact vagina tissues were resected and then frozen in liquid nitrogen or embedded in paraffin. Real-time fluorescence-based quantitative PCR (qRT-PCR) and immunofluorescent staining were performed to measure mRNA expression and immunofluorescence intensities of retinoic acid-related orphan receptorγt (RORγt), RORα and IL-17, respectively. Western blot analysis was conducted to determine protein expression of RORγt and IL-17. Results Laser scanning confocal microscopy showed that RORγt, RORα and IL-17 immunofluorescence was mainly located at inflammatory cells of the lamina propria and blood vessels in En and Cn groups, at mucosal epithelium, adherent hyphae, and inflammatory cells of the lamina propria and blood vessels in Ci group, and at mucosal epithelium, vaginal canal and endocytosed hyphae, and inflammatory cells of the lamina propria and blood vessels in Ei group. qRT-PCR and immunofluorescent staining uncovered that mRNA expression and immunofluorescence intensities of RORγt, RORα and IL-17 were significantly higher in En, Ci and Ei groups than in Cn group at the same time points (all P 0.05). Moreover, RORγt and IL-17 protein expression in Ci and Ei groups was obviously up-regulated on day 7 after inoculation (RORγt: F = 13.137, P < 0.001; IL-17: F = 11.182, P < 0.001), but was not increased further on day 14. Conclusion Vaginal candida infection can up-regulate the expression of RORγt, RORα and IL-17, suggesting that Th17/IL-17 axis may be involved in the occurrence of vaginal candidiasis in BALB/c mice. Key words: Candidiasis, vulvovaginal; Candida albicans; Estrogens; Nuclear receptor subfamily 1, group F, member 3; Interleukin-17; Mice; Retinoic acid-related orphan receptors