| Background and objectivesThe definition of recurrent spontaneous abortion(RSA)varies in different countries.The criteria of the American Society for Reproductive Medicine defines RSA as twice or more pregnancy failures;in 2016,the Department of Obstetrics And Gynecology of Chinese Medical Association established the diagnostic criteria for RSA as three times or more spontaneous abortions.Moreover,abortion refers to pregnant termination due to unviable embryo or fetus.Different countries and regions have different definitions for miscarriage weeks.In China,it refers to pregnancies that are less than 28 weeks or fetuses that weight less than 1000g.Miscarriages which occur before 12 weeks are known as early abortions,then,miscarriages after 12 weeks are known as late abortions.31%of embryos develop to spontaneous abortions after implantation,80%of which are early abortions,among them,nearly 2/3 are clinically silent miscarriages,also called biochemical pregnancies,which occur before the menstrual period.Although recurrent spontaneous abortion is defined as three times or more spontaneous abortions,most experts consider that two successive abortions ought to be considered and evaluated because the risk of recurrent miscarriage is similar to that of three times.Synthesizing the definition and conprehension of RSA at domestic and overseas,It finally reaches the criteria of RSA in this study:twice or more early spontaneous abortions that occur successively with the same sexual partnerThe causes of recurrent miscarriage are very complexed of genetic factors(embryo or couple chromosomal abnormalities),anatomical factors(cervical insufficiency,congenital uterine dysplasia,uterine lesions,etc.),endocrine abnormalities(hypercapsule dysfunction,polycystic ovary syndrome,diabetes,thyroid dysfunction,hyperprolactinemia,etc.),infection(chlamydia,mycoplasma,toxoplasma,herpes simplex virus,cytomegalovirus and human parvovirus B19,etc.),immune factors(autoimmune and allotype),prethrombotic state(congenital and acquired)and other factors(bad environment,bad psychology,smoking,alcohol abuse,etc.).However,more than 50%cases of RSA can’t make out final causes,which is called unexplained recurrent spontaneous abortion(URSA)[1].This study established an explant culture model by collecting the villi of URSA patients during early pregnancy.This study conducted explant culture on the villi tissue of aborted villi from URSA patients and the villi from patients undergoing artificial abortion in normal pregnancy,aiming to explore the effects of low-molecular weight heparin(LMWH)and hydroxychloroquine(HCQ)on the growth of villous tissues,and to study its application value in URSA patients and the guiding significance of re-pregnancy[2].Materials and methodsSix patients of unexplained recurrent spontaneous abortion who were admitted to the Maternal and Child Health Hospital of Shandong Province from September 2018 to March 2019 were randomly selected as the case group.Meanwhile,6 other abortion patients at the same gestation age were selected as the control groupThis study divided the case group embryo villi into three groups.after successful culture into explants,they were divided into blank control group,HCQ group and LMWH group according to the different media addition.The drug treatment group was divided into low dose group,treatment dose group and high dose group according to the drug concentrationThe concentration of low molecular weight heparin was set to 2.5 u/ml in the normal dose group,25 u/ml in the overdose group,and 0.25 u/ml in the low dose group.The concentration of hydroxychloroquine group was set lOug/ml in the normal dose group,1ug/ml in the low dose group,50ug/ml in the overdose group).Another group was supplemented with medium containing no drug.The villus explants were observed for 72 hours.The images were collected at 72h,ImageJ software was used to quantify exogenous migration and migration distance,which were statistically analyzed by SPSS 20.0 statistical software.Chi-square analysis was used to correct the chi-square analysis method.P<0.05 was considered as the difference.The purpose of this study was to analyze the significant difference in the migration distance of explants between case group and control group,or between different drugs in the case group,or among different concentrations of the same drug in case group.Results:The migration distance of villi explants in the normal control group was(333.38±112.89)μm,and the migration distance of villous explants in the blank control group of the case group was(73.10±21.28)pm,the migration distance of HCQ low dose group(1±g/ml)was(125.37±34.66)μm,and the migration distance of treatment dose group(0±g/ml)was(235.00±89.46)μm.The migration distance of the high-dose group(50μg/ml)was(135.25±42.05)μm,the migration distance of the LMWH low-dose group(0.25U/ml)was(110.19±18.32)μm,and the migration distance of the therapeutic dose group(2.5U/ml)was(216.33±85.11)μm,and the migration distance of the high-dose group(25U/ml)was(109.99±18.33)μm1.The migration distance of explants in the normal control group after 72 hours of culture was greater than that of the URSA blank control group after 72 hours,and there was a statistical difference between the two groups(P<0.01).2.The in vitro migration distance of HCQ and LMWH treatment doses of embryonic villus explants of URSA patients in the intervention group was greater than the migration distance of the explants in the blank control group,and there was a statistical difference between the two groups(P<0.05).3.The migration distance of villous explants under the treatment doses of HCQ and LMWH was long in the low-dose group and the high-dose group,and there was a statistical difference between the groups(P<0.05)4.URSA villous explants were treated with a therapeutic dose of HCQ for 72h,and the migration distance of explants was not statistically different from that of the normal control group.After 72h of LMWH treatment,it was statistically significant compared with the normal control group.Both HCQ and LMWH There was no significant difference in migration distance between the two drug treatment groups after 72 hours of treatment.Conclusions:1.Compared with normal villi,embryonic villi explants in URSA patients have a shorter in vitro migration distance and poor migration ability2.The therapeutic dose of HCQ and LMWH can improve the insufficiency of villi migration in URSA patients,but there is still a gap compared with normal pregnancy patients.3.The therapeutic dose of HCQ and LMWH has better effect on the exophytic growth of URSA patients than the high-dose and low-dose groups. |
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