Pregnancy outcomes following the administration of high doses of dexamethasone in early pregnancy

Abstract

Objective: In the present study, we aimed to evaluate the effects of high doses of dexamethasone (DEX) in early pregnancy on pregnancy outcomes.

Methods: Pregnant BALB/c mice were treated with high-dose DEX in the experimental group or saline in the control group on gestational days (GDs) 0.5 to 4.5. Pregnant mice were sacrificed on GDs 7.5, 13.5, or 18.5 and their peripheral blood, placentas, fetuses, and uterine tissue were collected. Decidual and placenta cell supernatants were examined to evaluate the effect of DEX on the proliferation of mononuclear cells, the quantity of uterine macrophages and uterine natural killer (uNK) cells, and levels of progesterone and 17β-estradiol, as determined by an 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide assay, immunohistochemistry, and enzyme-linked immunosorbent assay, respectively. We also were measured fetal and placental growth parameters on GD 18.5.

Results: We found that high doses of DEX were associated with an increased abortion rate, enhancement of the immunosuppressive effect of the decidua, alterations in placental growth parameters, decreased progesterone and 17β-estradiol levels, and a reduced frequency of macrophages and uNK cells.

Conclusion: Our data suggest that the high-dose administration of DEX during early pregnancy negatively affected pregnancy outcomes.

Keywords: Abortion; Dexamethasone; Estradiol; Macrophage; Natural killer cell; Pregnancy; Progesterone.

Figure 1. The effects of high doses of DEX in early pregnancy on pregnancy outcomes and appearance of the uterus on gestational day 13.5. (A) Treatment of pregnant BALB/c mice with DEX in early pregnancy significantly the increased abortion rate in comparison with PBS-treated pregnant BALB/c mice (^*p<0.01). (B) No significant difference was found in the number of implantation sites between DEX- and PBS-treated pregnant BALB/c mice. (C) Aborted fetuses in the uteri of DEX-treated pregnant BALB/c mice were identified by their small size accompanied by a necrotic appearance in comparison with normal embryos and placentas. (D) The nine viable fetuses and one aborted fetus in the uterine horns were obtained from PBS-treated pregnant BALB/c mice on gestational day 13.5. Results are expressed as mean±standard error of measurement from seven independent experiments and were analyzed using the nonparametric Mann-Whitney U test in two independent experiments. DEX, dexamethasone; PBS, phosphate buffer saline.

Figure 2. The effects of DS and PS on the proliferation of mitogen-stimulated mouse spleen mononuclear cells. (A) DS from DEX-treated pregnant BALB/c mice significantly suppressed the proliferation of PHA- and LPS-stimulated mononuclear cells in comparison with DS from PBS-treated pregnant BALB/c mice. (B) PS from DEX-treated pregnant BALB/c mice and PS from PBS-treated pregnant BALB/c had a similar inhibitory effect on PHA- and LPS-stimulated mononuclear cell proliferation. All data presented in these charts were obtained by the treatment of mouse spleen mononuclear cells with a 5% concentration of DS or PS. The stimulation index was calculated by dividing the optical density test of each group by the optical density control for each group. The results are expressed as mean±standard error of measurement from seven independent experiments. DS, decidual cell supernatant; PS, placenta cell supernatant; DEX, dexamethasone; PHA, phytohemagglutinin; LPS, lipopolysaccharides; PBS, phosphate-buffered saline. ^*p<0.05, ^**p<0.01 was used for the statistical analysis.

Figure 3. The effects of DEX treatment in early pregnancy on serum levels of progesterone and 17β-estradiol. Pregnant BALB/c mice were injected with DEX or PBS on gestational days 0.5 to 4.5 and sacrificed on gestational day 7.5. Levels of progesterone (ng/mL) and 17β-estradiol (pg/mL) were measured by enzyme-linked immunosorbent assay in serum samples from pregnant BALB/c mice. The serum concentration of 17β-estradiol (B), especially progesterone (A), was decreased in DEX-treated pregnant BALB/c mice compared with the PBS-treated pregnant BALB/c mice. Data are reported as mean±standard error of measurement from seven independent experiments and the unpaired Student's t-test with ^*p<0.05, ^**p<0.01 was used for the statistical analysis. DEX, dexamethasone; PBS, phosphate-buffered saline.

Figure 4. Immunohistochemical comparative analysis of macrophages in the uteri of pregnant BALB/c mice on gestational day 7.5. Cryosections were prepared from the uteri of pregnant BALB/c mice and stained with F4/80 antibody. Tissues were also counterstained with hematoxylin. Positive cells were counted and expressed as a percentage of the total number of nucleated cells. Macrophages were present in the myometrium, mesometrium, and decidua basalis. The frequency of uterine macrophages significantly decreased in pregnant BALB/c mice treated with DEX (A, B) when compared with pregnant BALB/c mice treated with PBS (C, D). DEX, dexamethasone; PBS, phosphate-buffered saline; M, mesometrial side; DB, decidual basalis; My, myometrium.

Figure 5. Immunohistochemical comparative analysis of NK cells in the uteri of pregnant BALB/c mice on day 7.5 of gestation. Cryosections were prepared from the uterus of pregnant BALB/c mice and stained with biotinylated dolichos biflorus agglutinin lectin. Tissues were also counterstained with hematoxylin. Positive cells were counted and expressed as a percentage of the total number of nucleated cells. The mesometrial regions are at the top of each image. The NK cells accumulated in the decidua basalis and around the glandular and luminal epithelial layers. The frequency of uterine NK cells significantly decreased in pregnant BALB/c mice treated with DEX (A, B) when compared with pregnant BALB/c mice treated with PBS (C, D). NK, natural killer; DEX, dexamethasone; PBS, phosphate-buffered saline; M, mesometrial side; My, myometrium; L, luminal epithelium; DB, decidual basalis; GE, glandular epithelium.

Figure 6. The in vitro quantification of fetal and placental growth parameters in pregnant BALB/c mice treated with DEX or PBS on gestational day 18.5 (^*p<0.05; ^**p<0.01). The results are expressed as mean concentrations±standard error of the mean. DEX, dexamethasone; PBS, phosphate-buffered saline; WU, weight of the uterus; WP, weight of placenta; PD, placenta diameter; BWE, body weight of embryo; CRL, crown-rump length; BPD, biparietal diameter.