Repopulation of autophagy-deficient stromal cells with autophagy-intact cells after repeated breeding in uterine mesenchyme-specific Atg7 knockout mice

Abstract

Objective: Autophagy is highly active in ovariectomized mice experiencing hormone deprivation, especially in the uterine mesenchyme. Autophagy is responsible for the turnover of vasoactive factors in the uterus, which was demonstrated in anti-Müllerian hormone receptor type 2 receptor (<i>Amhr2</i>)-Cre-driven autophagy-related gene 7 (<i>Atg7</i>) knockout (<i>Amhr-Cre/Atg7<sup>f/f</sup></i> mice). In that study, we uncovered a striking difference in the amount of sequestosome 1 (SQSTM1) accumulation between virgin mice and breeder mice with the same genotype. Herein, we aimed to determine whether repeated breeding changed the composition of mesenchymal cell populations in the uterine stroma.Methods: All female mice used in this study were of the same genotype. <i>Atg7</i> was deleted by <i>Amhr2</i> promoter-driven Cre recombinase in the uterine stroma and myometrium, except for a triangular stromal region on the mesometrial side. <i>Amhr-Cre/Atg7<sup>f/f</sup></i> female mice were divided into two groups: virgin mice with no mating history and aged between 11 and 12 months, and breeder mice with at least 6-month breeding cycles with multiple pregnancies and aged around 12 months. The uteri were used for Western blotting and immunofluorescence staining. Results: SQSTM1 accumulation, representing <i>Atg7</i> deletion and halted autophagy, was much higher in virgin mice than in breeders. Breeders showed reduced accumulation of several vasoconstrictive factors, which are potential autophagy targets, in the uterus, suggesting that the uterine stroma was repopulated with autophagy-intact cells during repeated pregnancies. Conclusion: Multiple pregnancies seem to have improved the uterine environment by replacing autophagy-deficient cells with autophagy-intact cells, providing evidence of cell mixing.