ABSTRACT
Objective Endoplasmic reticulum stress may be involved in the menstrual process, but the detailed mechanism is unclear. This article is to explore the effect of progesterone replantation at different periods on the endoplasmic reticulum stress-induced autophagy and apoptosis in mice during menstruation. Methods Mice were divided into 12h replantation group, 16h replantation group and the withdrawal group according to random number table, with 10 mice in each group. 12h (12h replantation group) and 16h (16h replantation group) after the withdrawal of progesterone, replanted respectively with progesterone tube, and withdrawal group were not replanted. Mice in each group were sacrificed 24h after the withdrawal of progesterone and were collected bilateral uterine horns. The expression and localization of t-PERK, p-PERK, t-eIF2α, p-eIF2α, ATF4, CHOP and Caspase12 mRNA and proteins in mouse endometrial tissues were detected by qPCR, Western blot and immunohistochemistry. Results At the mRNA level, the PERK and eIF2α(1.000 ± 0.000) of the 12h replantation group were significantly higher than those of the 16h replantation group (0.450 ± 0.049, 0.330 ± 0.015) and the withdrawal group (0.260 ± 0.233, 0.195 ± 0.014). The difference was statistically significant (P<0.05), and the 16h replantation group was higher than the withdrawal group (P<0.05). At the protein level, p-PERK / t-PERK (0.606 ± 0.051) and p-eIF2α / t-eIF2α (0.795 ± 0.074) in 12h replantation group were significantly higher than those in 16h replantation group (0.367 ± 0.019, 0.503 ± 0.038) and withdrawal group (0.243 ± 0.020, 0.293 ± 0.020). The difference was statistically significant (P<0.05) and the 16h replantation group was significantly higher than the withdrawal group. At the mRNA and protein levels, ATF4 of 12h replantation group were higher than 16h replantation group and the withdrawal group (P <0.05) , and the 16h replantation group was higher than the withdrawal group. The proteins of p-PERK, p-eIF2α and ATF4 are mainly expressed in the cytoplasm of decidual stromal cells which located at the junction of basal layer and decidualized stromal cells. At mRNA and protein levels, the expression levels of CHOP and Caspase12 in 12h replantation group were significantly lower than those in 16h replantation group , were also lower than the withdrawal group, and the 16h replantation group was lower than the withdrawal group (P<0.05). The proteins of CHOP and Caspase12 are mainly localized in cytoplasm of adequately decidualized stromal cells and the glandular epithelium cells. Conclusion Replantation of progesterone before the critical period of menstruation (12h) can effectively block the onset of menstruation in mice, which may be achieved by maintaining the expression of PERK/eIF2α/ATF4 signal and blocking the expression of apoptotic signaling pathway members CHOP and Caspase 12 in endometrial stromal cells. And this effect can only be partially achieved by progesterone replantationing after the critical period of menstruation (16h).
ABSTRACT
<p><b>OBJECTIVE</b>The effect of angelica sinensis polysaccharides (ASP) on the production of reactive oxygen specie (ROS), the capability of total anti-oxidant (T-AOC), and the expression of p16 in mRNA level in mice hematopoietic stem cells (HSCs) were observed to explore the underlying mechanism that ASP delay aging of HSCs in vivo.</p><p><b>METHOD</b>C57BL/6J mice were randomly divided into normal group, aging group, and the above groups treated with ASP. Mice were uniformly explored in X-ray (3.0 Gy/8 F) to erect model of aging. Normal and aging ASP intervention groups mice were treated with ASP by intragastric administration, while normal and aging groups were treated with equal-volume NS during X-ray irradiation. Mice HSCs were isolated by magnetic cell sorting and cultured in vitro. Senescence-associated beta-galactosidase (SA-beta-Gal) staining was used to detect aging HSCs. Cell cycles analysis and CFU-Mix cultivation were used to evaluate the capability of self-renewing and colony forming in HSCs. The production of ROS in HSCs was evaluated by flow cytometry analysis and immunofluorescence assess, respectively. T-AOC was detected by chemical colorimetric method. The expression of p16 was determined by real-time quantitative PCR (qRT-PCR).</p><p><b>RESULT</b>Exogenous X-ray irradiation induced HSCs aging was compared with normal group without irradiation. Biological feature of HSCs in aging group with X-ray irradiation as follows: The percentage of SA-beta-Gal positive cells, the ratio of G1 stages and the production of ROS were significantly increased , the expression of p16 in mRNA level was also upregulated. The capacility of colony forming and T-AOC in HSCs were decreased. ASP could significantly decrease the percentage of SA-beta-Gal positive cells, the ratio of G1 stages and the production of ROS in HSCs, and downregulate the expression of p16 in mRNA level in HSCs contrast to aging group without ASP treatment. In addition, ASP could remarkably increase T-AOC and the capacility of colony forming in HSCs compared with aging group without ASP treatment.</p><p><b>CONCLUSION</b>X-ray (3.0 Gy/8 F) could induce mice HSCs aging. ASP could delay senescence HSCs aging which maybe partly ascribed to the inhibition of oxidative damage and the downregulation of p16 mRNA expression.</p>