关键词: 刚地弓形虫, IDO, 蜕膜免疫细胞, 不良妊娠结局

Abstract:

Objective To investigate the effect of Toxoplasma gondii infection on indoleamine 2,3-dioxygenase (IDO) expression in decidual immune cells at the maternal-fetal interface during pregnancy, and to examine its association with adverse pregnancy outcomes. Methods A total of 40 wild-type pregnant mice of the C57BL/6 strain were divided into the wild-type control group and the infection group, of 20 animals in each group, while 20 pregnant IDO gene-knockout mice were assigned as the IDO^-/- infection group. Mice in two infection groups were intraperitoneally injected with 250 T. gondii RH tachyzoites in 250 µl PBS on day 8 of gestation, and animals in the wild-type control group were intraperitoneally injected with an equal volume of PBS. The mental state of the pregnant mice was observed on day 14 of gestation, and placental weight of pregnant mice and fetal weight were recorded and the incidence of abnormal fetal mice was calculated. Clinical decidual tissues were obtained from healthy pregnant women who voluntarily chose to terminate their pregnancies during the first trimester, and mononuclear cell suspensions of human decidual tissues were prepared using the human lymphocyte separation solution. Cell suspensions were divided into the control group and T. gondii infection group, with 1.5 × 10⁷ cells in each group. No treatment was given in the control group, and cells in the T. gondii infection group were infected with T. gondii RH strain tachyzoites at a tachyzoite-to-cell ratio of 1 : 3. Cells were harvested in both groups 30 hours following incubation. The IDO expression was detected in decidual natural killer (dNK) cells, decidual macrophages (dMϕ), decidual dendritic cells (dDC), and decidual myeloid-derived suppressor cells (dMDSC) using flow cytometry. dNK cells, dMϕ, dDC, and dMDSC were separated and purified from human decidual mononuclear cells with CD3^-CD56⁺, CD14, DC, and CD33 magnetic microbeads separation kits, respectively. These types of cells were all divided into the control group and the T. gondii infection group, with 1 × 10⁷ cells in each group. Cells in the control group were given no treatment, while cells in the infection group were infected with T. gondii RH strain tachyzoites at a tachyzoite-to-cell ratio of 1 : 3. Cells in both groups were harvested 20 hours following culture. Total proteins were extracted from cells, and the IDO expression was determined in various types of decidual immune cells using Western blotting. All statistical analyses were performed using the software GraphPad Prism 10.1.2, and comparisons of means between groups were conducted with independent sample Student’s t-test. Results The pregnant mice exhibited poorer pregnancy outcomes, more listlessness, slower movements, more severe placental congestion, and placental dryness, collapse, and elasticity loss in the IIDO^-/- infection group relative to in the wild-type control group, and the pregnant mice in the IDO^-/- infection group had poorly developed fetuses and absorbed fetuses. The pregnant mice had lower placental [(55.73 ± 5.52) mg vs. (81.73 ± 9.35) mg; t = 6.329, P < 0.01] and fetal weights [(106.88 ± 37.33) mg vs. (198.66 ± 26.58) mg; t = 5.299, P < 0.01) in the wild-type infection group than in the wild-type control group, and the pregnant mice had lower placental [(39.87 ± 4.24) mg] and fetal weights [(51.67 ± 19.75) mg] in the IDO^-/- infection group than in the wild-type infection group (t = 5.726 and 3.241, both P values < 0.01). No abnormal fetal mice were seen in the wild-type infection group, while a higher incidence rate of abnormal fetuses was found in the wild-type infection group [(40.6 ± 20.69)%] than in the wild-type control group (t = 5.193, P < 0.01) and higher in the IDO^-/- infection group [(93.33 ± 10.80)%] than in the wild-type infection group (t = 5.600, P < 0.01). Flow cytometry detected no significant difference in the proportion of positive IDO expression in dNK cells between the infection and control groups [(8.02 ± 3.20)% vs. (9.65 ± 2.62)%; t = 2.025, P > 0.05], and higher proportions of positive IDO expression in dMϕ [(27.50 ± 6.24)% vs. (35.07 ± 8.11)%; t = 5.830, P < 0.05], dDC [(20.23 ± 3.20)% vs. (24.80 ± 4.13)%; t = 5.533, P < 0.05] and dMDSC [(3.14 ± 0.57)% vs. (6.81 ± 0.88)%; t = 5.644, P < 0.05] in the infection group than in the control group. The largest reduction in the proportion of positive IDO expression [(53.20 ± 11.56)%] was seen in dMDSC following T. gondii infection, followed by in dMϕ [(21.45 ± 3.65)%], dDC [(18.22 ± 4.17)%] and dNK cells [(17.84 ± 15.54)%]. Western blotting determined relatively lower IDO expression in dNK cells [(0.91 ± 0.10) vs. (1.04 ± 0.11); t = 13.360, P < 0.01], dMϕ [(0.62 ± 0.12) vs. (1.14 ± 0.09); t = 12.930, P < 0.01)], dDC [(0.75 ± 0.11) vs. (1.11 ± 0.06); t = 7.482, P < 0.05] and dMDSC [(0.47 ± 0.06) vs. (1.05 ± 0.03); t = 13.960, P < 0.01] in the infection group than in the control group. The largest reduction in the relative IDO expression [(55.22 ± 16.12)%] was seen in dMDSC following T. gondii infection, followed by in dMϕ [(46.30 ± 6.98)%], dDC [(33.53 ± 7.78)%] and dNK cells [(12.74 ± 0.67)%]. Conclusion T. gondii infection may downregulate the IDO expression in dNK cells, dMϕ, dDC and dMDSC at the maternal-fetal interface during the first trimester of pregnancy, thereby increasing the incidence of adverse pregnancy outcomes. These findings suggest that downregulation of IDO expression is closely associated with the occurrence of adverse pregnancy outcomes.

Key words: Toxoplasma gondii, Indoleamine 2,3-dioxygenase, Decidual immune cell, Adverse pregnancy outcomes