Uterine cyclin A2–deficient mice as a model of female early pregnancy loss
Fatimah Aljubran, Katelyn Schumacher, Amanda Graham, Sumedha Gunewardena, Courtney Marsh, Michael Lydic, Kristin Holoch, Warren B. Nothnick
Fatimah Aljubran, Katelyn Schumacher, Amanda Graham, Sumedha Gunewardena, Courtney Marsh, Michael Lydic, Kristin Holoch, Warren B. Nothnick
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Research Article Reproductive biology

Uterine cyclin A2–deficient mice as a model of female early pregnancy loss

Abstract

Proper action of the female sex steroids 17β-estradiol (E2) and progesterone (P4) on the endometrium is essential for fertility. Beyond its role in regulating the cell cycle, cyclin A2 (CCNA2) also mediates E2 and P4 signaling in vitro, but a potential role in modulating steroid action for proper endometrial tissue development and function is unknown. To fill this gap in our knowledge, we examined human endometrial tissue from fertile and infertile cisgender women for CCNA2 expression and correlated this with pregnancy outcome. Functional assessment of CCNA2 was validated in vivo using a conditional Ccna2 uterine-deficient mouse model, while in vitro function was assessed using human cell culture models. We found that CCNA2 expression was significantly reduced in endometrial tissue, specifically the stromal cells, from women undergoing in vitro fertilization who failed to achieve pregnancy. Conditional deletion of Ccna2 from mouse uterine tissue resulted in an inability to achieve pregnancy, which appeared to be due to alterations in the process of decidualization, which was confirmed using in vitro models. From these studies, we conclude that CCNA2 expression during the proliferative/regenerative stage of the menstrual cycle allows for proper steroid responsiveness, decidualization, and pregnancy. When CCNA2 expression levels are insufficient, there is impaired endometrial responsiveness, aberrant decidualization, and loss of pregnancy.

Authors

Fatimah Aljubran, Katelyn Schumacher, Amanda Graham, Sumedha Gunewardena, Courtney Marsh, Michael Lydic, Kristin Holoch, Warren B. Nothnick

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Figure 7

CCNA2 expression in human and mouse uterine stromal cells and uterine tissue prior to and during decidualization.

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CCNA2 expression in human and mouse uterine stromal cells and uterine ti...
(A–C) qRT-PCR assessment of (A) CCNA2, (B) PRL, and (C) IGFBP1 transcript expression during in vitro decidualization in nontransfected t-HESCs. (D) Ccna2 expression prior to and during late decidualization in mouse uterine tissue. Data are normally distributed and analyzed by 1-way ANOVA followed by Bonferroni’s post-hoc analysis. Different letters indicate statistical significance among the groups. In D, a versus b = P < 0.001. P < 0.001 for n =6 mice for dpc 0.5 and 2.5 and n = 5 mice for dpc 6.5. (E–G) qRT-PCR analysis of (E) CCNA2, (F) PRL, and (G) IGFBP1 transcript expression in NT-siRNA and CCNA2-siRNA transfected t-HESC prior to (day 0), and after a decidualization stimulus. Different block letters indicate statistical significance among the means across days of culture/treatment groups for NT-siRNA-transfected cells while different letters in italics indicate statistically significant differences among the means in the CCNA2-siRNA-transfected cells using 1-way ANOVA followed by Tukey-Kramer multiple comparison tests. In E, a versus b = P < 0.001. a versus b = P < 0.001; in F, a versus b = P < 0.001, a versus c = P < 0.001, b versus c = P < 0.001; a versus b = P < 0.001, a versus c = P < 0.001, b versus c = P < 0.01; in G, a versus b = P < 0.001, a versus c = P < 0.001, b versus c = P < 0.001; a versus b = P < 0.001, a versus c = P < 0.001, b versus c = P < 0.05. ***P < 0.001 between transfection groups within each time point using unpaired, 2-tailed t tests. For Figure 7, B and C, as well as Figure 7, E and F, values are expressed as the log10 value of the FC from the appropriate control for each transcript. Data are from 4 separate experiments (n = 4 replicates with different cell passage numbers). Within each graph of the box and whisker plot, X indicates the mean values while the whisker end points indicate the highest and lowest values.