Multiple mechanisms are involved in apoptotic cell death in the mouse uterus and vagina after ovariectomy
Introduction
Cell proliferation, differentiation, and regression of the uterus during the estrous cycle are regulated by ovarian estrogen (17β-estradiol; E2) and progesterone in rats [1]. In the endometrium of the human uterus, apoptosis is an integral component of the menstrual process [2]. Apoptosis can be induced by sex hormone withdrawal in the uterus and prostate in rats and mice [3], [4], [5], [6]. We have demonstrated that regression of the uterus and vagina after ovariectomy was accompanied by induction of new proteins [7], indicating that some specific factors could be involved in the regression mechanism in male and female reproductive organs. Transforming growth factor-β (TGF-β) was increased in the prostate on the first day after androgen removal in the rat [8] and addition of TGF-β1 to primary culture of rabbit uterine epithelial cells caused apoptosis in vitro [9]. Fas, a member of the NGF/TNF receptor family, transduces an apoptotic signal in immunocytes [10], [11] and is thought to be involved in cell death in the human endometrium [12]. The expression of tumor necrosis factor-α (TNF-α) mRNA and TNF-α protein in the human uterus was correlated with the menstrual cycle [13], [14] and mouse estrous cycle [15]. On the other hand, bcl-2 and related proteins also regulate the survival and death of cells [16], [17], [18], [19]. In human endometrium, expression of the bcl-2 gene family changes during the menstrual cycle [20], [21]. Menstruation is correlated with changes in bcl-2, bax, and TNF-α expression level [22], [23]. These results raise the question of whether these factors could be participating in apoptosis in the uterus and vagina after ovariectomy.
The present study was designed to further examine the regression mechanism of the mouse uterus and vagina induced by hormone withdrawal. C57BL/Tw female mice were ovariectomized at estrus, and histologic and biochemical changes were examined by the in situ 3′-end labeling method, electron microscopy, and RT-PCR. In order to study the involvement of Fas, histologic changes in the uterus and vagina were examined in lprcg female mice that were ovariectomized at estrus. The lprcg mice have a point mutation of the gene encoding Fas, resulting in an autoimmune disorder [24]. The lprcg mice are capable of reproduction, suggesting that their reproductive organs might be the under control of ovarian hormones. Our results indicate that apoptotic cell death is induced by ovariectomy through the mediation of both Fas- and TNF-α-systems; however, uterine and vaginal cells in CBA lprcg/lprcg mice lacking functional Fas showed apoptosis indicating that the Fas system may not be essential.
Section snippets
Animals
Female mice of the C57BL/Tw strain were kept under 12 h light/12 h dark at 23–25 °C and fed laboratory chow (CE-2, CLEA, Tokyo, Japan) and tap water ad libitum. Mice were killed at estrus, and 1, 2, 3, and 5 days after ovariectomy, which was carried out at estrus. The estrous cycle was determined by vaginal smears. The uteri and vaginae from five mice in each group were weighed and fixed in 4% buffered formaldehyde for histologic preparation. Portions of the organs were frozen in liquid nitrogen
BrdU labeling index and apoptotic index
Weights of uteri and vaginae decreased following ovariectomy (Fig. 1). BrdU labeling index in uterine luminal and glandular epithelia was low at estrus, but it increased significantly 5 days after ovariectomy (Fig. 2A). In vaginal epithelium, the labeling index was high at estrus but it decreased significantly 2 days after ovariectomy. BrdU labeling index was not altered in the stromal cells of either the uterus or vagina by ovariectomy (Fig. 2B).
Apoptotic index in uterine luminal epithelial
Discussion
Apoptotic cell death plays a critical role in tissue homeostasis. Morphologic changes of apoptosis are electron-microscopically characterized by compaction and segregation of chromatin in sharply circumscribed masses that abut the inner surface of the nuclear envelope [30]. The nuclear fragments and condensation of the cytoplasm are associated with extensive cell surface protrusion, followed by separation of the surface protuberances to produce membrane-bounded apoptotic bodies of varying sizes
Acknowledgments
We thank Emeritus Professor Noboru Takasugi at Yokohama City University, for his valuable advice and critical readings of this manuscript. This work was supported by a Grant-in-Aid for Scientific Research on Priority Areas (A) from the Ministry of Education, Culture, Sports, Science and Technology of Japan, a Research Grant from Kihara Memorial Yokohama Foundation for the Advancement of Life Sciences (to T.I. and T.S.), a Grant for Support of the Promotion of Research at Yokohama City
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2021, DifferentiationCitation Excerpt :Overexpression of ERBBs is associated with carcinogenesis in the reproductive organs (Dickson and Lippman, 1995) and ERBB2 is frequently amplified and overexpressed in cancer cells (Hynes and Stern, 1994). Apoptotic cell death occurs in vaginal epithelial cells and there is decreased DNA synthesis during the estrous cycle and after ovariectomy in intact rats and mice (Sato et al., 1997, 2003). In neonatally DES-exposed adult mice, however, ovariectomy did not induce reduction of DNA synthesis and only a slight increase in apoptotic cells in vaginal epithelium (Sato et al., 2004).
The Bax/Bcl-2 apoptotic pathway is not responsible for the increase in apoptosis in the RU486-treated rat uterus during early pregnancy
2013, Reproductive BiologyCitation Excerpt :The blocking of the glucocorticoid receptors by RU486 binding may account for the shift in the Bax/Bcl-2 ratio toward a more anti-apoptotic environment, however, this does not account for the increase in observed apoptotic activity within the RU486-treated rat endometrium. Sato et al. [25] demonstrated increased levels of TNF-α, and Fas and Fas-ligand (FasL) 24 h after ovariectomy within the endometrium. Both TNF-α and Fas and FasL have been reported to be involved in the up-regulation of apoptosis [28,29].
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2010, Journal of Sexual MedicineCitation Excerpt :Ovariectomy-induced reduction in the active form of AKT, which is also an anti-apoptotic factor [37], may rather indicate reduced anti-apoptotic capability of the vagina after estrogen withdrawal. Indeed, a considerable level of apoptosis has been observed in the mouse vagina as early as 1 day after ovariectomy [38]. At present, the exact mechanism of estrogen-mediated eNOS (Ser-1177) phosphorylation in the rat vagina is unknown.
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Present address: Division of Cerebral Structure, National Institute for Physiological Sciences, 38 Nishigonaka, Myodaiji, Okazaki 444-8585, Japan.