| Objective:Immunofluorescence was used to detect the expression of AQP1 and AQP6 in the decidua of recurrent abortion rats,and to clarify the effect of Shoutai Wan on aquaporin in the decidual tissue of recurrent abortion rats;From the perspective of fluid transport,the possible mechanism of Shoutai Pills in preventing and treating recurrent miscarriage is explored,which provides new ideas and theoretical basis for the prevention and treatment of recurrent spontaneous abortion by traditional Chinese medicine.Methods:1.Modeling60 healthy and mature SPF SD female rats of 8 weeks old and 1 week of quarantine were selected,weighing(216.09 ± 8.49)g;30 10-week old rats,weighing(253.87 ± 6.55)g SPF grade SD male rats,after a week of adaptive feeding,were fed in a cage according to the ratio of female rats: male rats =2: 1,and observed the mating situation.The female rats observed vagina in the vagina the next morning The plug was shed(ie,a milky white solid in the vagina),and a large number of sperm could be seen on the vaginal smear as the first day of pregnancy.After the pregnancy was determined,60 pregnant rats were divided into 6 groups according to the random number table method and marked.On the morning of the 1st to 9th day of pregnancy,the Shoutai pill high,middle and low groups and the dydrogesterone group were given intragastric administration respectively;the model group and normal pregnancy group were given normal saline every morning stomach.On the afternoon of the first to the 9th day of pregnancy,the pregnant rats in the normal pregnancy group were given 2m L / d of normal saline by intragastric administration,and the other groups of pregnant rats were intragastrically administered with hydroxyurea tablets in water(450 mg / Kg)· 2 m L / d;On the morning of the 10 th day,the pregnant mice in 6 groups were gavage(4.0mg / Kg)· 2m L / d with mifepristone tablet aqueous solution;the body weight of the pregnant mice was required to be measured on the 1st,4th and 9th day of modeling.2.MaterialOn the eleventh day of pregnancy,all pregnant rats were sacrificed after neck rupture,and the uterus of the pregnant rats was opened to observe and record the uterine morphology of each pregnant mouse.Decidual tissue,use filter paper to suck the residual blood on the decidual tissue,and then fix it for use.3.DetectionImmunofluorescence staining was used for film production and theexpression and location of aquaporin 1,6(AQP1,AQP6)in decidual tissue were detected.The fluorescence microscope was used for computer imaging,and the magnification was 40 * 10 times.Image J software was used for image analysis,and the IOD / Area value of each specimen was measured and calculated,and the IOD / Area value was used as the semi-quantitative statistical analysis parameter.Results:1.Embryo developmentOn the 11 th day of pregnancy(24 hours after the last dose),the pregnant rats were sacrificed and their uteruses were isolated by open dissection.Visually observe the morphology of the uterus of the pregnant rat: normal group: the uterus of the pregnant rat is light red,the size of the embryo tissue is the same,there is no slub-like change,there is no obvious congestion and stasis in the uterine cavity,the size of the implantation point is the same,the embryo tissue is not Model group: the uterus of pregnant rats is dark red,partly dark black,the embryo tissues vary in size,and there is obvious slub-like change,the congestion in the uterine cavity is obvious,the congestion spot can be seen at the implantation point,or the implantation is more normal The spots were significantly reduced,and the embryo was partially organized;the dydrogesterone group: the uterus of pregnant rats was light red or dark red,and the embryo tissues wereindividually small,without slub changes,and occasionally a small amount of congestion spots in the uterine cavity,and no implantation sites Significantly reduced,the embryonic tissue was not organized;the three groups of Shoutaiwan high-dose group,Shoutaiwan medium-dose group,and Shoutaiwan low-dose group were basically the same,and there were slight differences between the groups: the uterus of pregnant rats was pale Red or dark red,a small part of the embryos are not uniform in size,no slub changes,occasional congestion in the uterine cavity,the implantation point is not significantly reduced,and the embryonic tissue is not organized.2.Comparison of embryo abortion rate among pregnant rats in each groupThe miscarriage rates in the high,medium and low dose groups of Shoutai Pill,didyrogesterone group,model blank group and normal pregnancy group were: 26.92%,29.70%,30.28%,27.27%,55.68%,8.25%.From the perspective of miscarriage: the model blank group had the highest miscarriage rate,and the normal pregnancy group had the lowest miscarriage rate.The 4 groups with drug intervention from low to high were: Shoutai pill high-dose group,didyrogesterone group,and longevity Feiwan medium-dose group and Shoutaiwan low-dose group.It can be seen that,in terms of miscarriage rate,the high-dose group of Shoutai Wan has a better effect than the middle-and low-dose groups and thedydrogesterone group.3.Differences in expression of AQP1 and AQP6 in decidual tissues of pregnant ratsAQP1 expression: Compared with the normal pregnancy group,the model group P <0.01,the difference was statistically significant,suggesting that the expression of AQP1 in the uterine decidua of the model group was significantly lower than that of the normal group;after drug intervention,and Compared with the model group,the Shoutai pill high and middle dose groups and the dydrogesterone group all met P <0.05,and the difference was statistically significant,suggesting that the Shoutai pill high,middle dose group and the dydrogesterone group can make RSA rats The expression of AQP1 in the decidual tissue of the uterus increased.Among them,the high-dose group of Shoutaiwan P < 0.01,the difference was statistically significant;compared with the high-dose group of Shoutaiwan,the medium and low dose group of Shoutaiwan P < 0.05,the difference Statistically significant,indicating that the high-dose Shoutai pill group had a stronger effect on the expression of AQP1 than the middle-and low-dose groups;compared with the didyrogesterone group,the high-dose Shoutai pill group P <0.05,indicating that the high-dose Shoutai pill The improvement of AQP1 gene expression was higher than that of didyprogesterone.S > Taiwan pill middle-dose group P > 0.05,indicating that Shoutai pill middle-dose group and didyrogesterone group can increase AQP1 gene expression,but the efficacy of the two is equivalent.AQP6 expression: compared with the normal pregnancy group,the model group P <0.01,the difference was statistically significant,suggesting that the expression of AQP6 in the uterine decidua of the pregnant rats of the model group was significantly reduced;after drug intervention,compared with the model group,Shoutai Wan Both the high-dose group and the dydrogesterone group met P <0.05,of which the Shoutai pill high-dose group P <0.01,suggesting that both the high-dose Shoutai pill and dydrogesterone can make AQP6 in the decidual tissue of RSA rats.The expression increased;compared with the Dydrogesterone group,the Shoutaiwan high-dose group P <0.05,indicating that the Shoutaiwan high-dose improved AQP6 gene expression was higher than Dydrogesterone.Conclusion:1.Shoutai Wan can improve the intrauterine congestion of RSA rats,prevent the implantation site from shrinking,and significantly improve the slub-like changes of the aborted uterus,which is conducive to the implantation of embryos and reduces the abortion rate of RSA rats;2.Shou Tai Wan may achieve the effect of relieving the fetus byimproving the expression of AQP1 and AQP6 genes in the decidual tissue of RSA rats;3.The expression of AQP1 and AQP6 genes in the RSA rat model is positively correlated with the dose of Shoutai Pills,indicating that the high-dose upregulation of AQP1 and AQP6 is more significant,but the correlation between its placenta mechanism and dose still needs further research. |
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