Effect Of N-acetyl-p-phenylenediamine On Decidualization Of Endometrial Stromal Cells And Pregnancy In Mice

Objective:Changes in the external environment may have an irreversible effect on pregnancy.During pregnancy,the female endometrium plays a vital role as the "foundation" of embryo implantation,and stromal cell decidualization is one of the important processes.Therefore,the study of environmental harmful substances on endometrial stromal cell decidualization and pregnancy outcome and related mechanisms will be conducive to the promotion of human reproductive health.Hair colorant has become an important part of human life,and the skin diseases caused by the use of hair colorants have been widely confirmed.In addition,epidemiological analysis also suggests that it is a high-risk factor for the occurrence of tumors.P-phenylenediamine is one of the main components of industrial colorants and a synthetic material of various hair colorants.When p-phenylenediamine enters the body through skin contact or respiratory contact,it is metabolized in the liver into N-acetyl-p-phenylenediamine and N,N-diacetyl-p-phenylenediamine.Studies have shown that n-acetyl-p-phenylenediamine in vivo has an obvious toxic effect on male reproduction in mice,but whether it has an effect on female pregnancy outcomes has not been confirmed.In recent years,the childbearing age of women has been increasingly delayed,and with the implementation of the two-child policy in China,the childbearing age also tends to be delayed.For elderly pregnant women,the adverse pregnancy outcomes such as birth defects and abortion are significantly higher than that of women of normal childbearing age due to changes in embryonic genetic material caused by genetic factors,endometrial aging and other reasons.Are there differences in the sensitivity to environmental toxins at different gestational ages? There are few literature reports.In this study,the toxic effects of N-acetyl on phenylenediamine on endometrial stromal cells and the effects on decidualization of mouse stromal cells were studied by establishing a mouse young and old endometrial stromal cell infection model,laying a foundation for the establishment of an animal infection model.The influence of N-acetyl-p-phenylenediamine on reproductive function and pregnancy outcome of female mice was studied by establishing the infection model of young and old female mice,and the related mechanism was preliminarily discussed,so as to provide a certain research basis for finally exploring the influence of hair colorants on female reproduction and pregnancy outcome.Methods:1 A model of endometrial stromal cell infection in young and old female mice was established.Different age groups were divided into two groups: the exposure groups(50 ?M,100 ?M,150 ?M and 200 ?M groups)and a control group.Immunohistochemistry(IHC),Western blot,PCR and other techniques were employed to detect the expression of related markers such as endometrial stromal cell activity,oxidative stress,apoptosis and autophagy in mice.In addition,the decidua markers were detected by PCR in different exposure groups.2 A subacute infection model of young and old female mice was established and divided into the exposure groups(26.88 mg/kg/day,53.74 mg/kg/day,107.5 mg/kg/day and 215 mg/kg/day groups)and the control group.The embryo implantation rate and birth rate of mice were counted in the mating experiment.The uterus samples of mice in each group were collected respectively.Immunohistochemistry,Western blot,PCR and other techniques were used to detect the autophagy and other related molecular markers of mice endometrium.Results:1 P16,a marker of endometrial aging,was significantly higher in the old group than in the young group.2 The cell toxicity model showed that there was no significant difference in ccK8 between n-acetyl-p-phenylenediamine 50 ?M and 100 ?M groups,and that ccK8 in n-acetyl-p-phenylenediamine 150?M and 200 ?M groups was significantly higher than that in control group.The production of reactive oxygen species(ROS)in different groups has an obvious dose-dependent effect,that is,the production of ROS gradually increases with the increase of toxic concentration.Bax in the exposure group was significantly higher than that in the control group,and the highest factor was 200?M.ATG5,an autophagy marker in the 50?M and 100?M groups was significantly higher than that in the control group,and ATG5 in the 100 ?M group were the highest,while it in the 150?M and 200?M groups were significantly lower than that in the 100?M group.3 The model of induced decidua in stromal cell showed that the expression of DTPRP,the decidual marker,was significantly lower in the elderly group than in the younger group.After 48 hours of decidua induction,the exposure group was significantly lower than the unexposed group.4 The mouse model showed that there was no difference in the embryo implantation rate and birth rate between different age groups and between different dose groups.Conclusions1 N-acetyl-p-phenylenediamine had no effect on the activity of mouse endometrial stromal cells in a certain low dose range,but the higher dose had obvious inhibitory effect,leading to the occurrence of stromal cell oxidation emergency and promoting cell apoptosis.However,the level of autophagy was increased at a lower dose and significantly decreased at a higher dose,indicating that autophagy plays a protective role on stromal cells under certain conditions to avoid being damaged by environmental poisons.2 The decidua ability of endometrial stromal cells in aged mice was significantly lower than that in young mice,but N-acetyl-p-phenylenediamine had no effect on embryo implantation and pregnancy.The decidualization of mouse endometrial stromal cells was obviously inhibited by N-acetylp-phenylenediamine.3 N-acetyl-p-phenylenediamine had no effect on pregnancy in mice,and autophagy may play a protective role in the female toxicity of N-acetyl-p-phenylenediamine.