The Role Of Homocysteine In Recurrent Abortion

Objective: Recurrent spontaneous abortion(RSA)is a failure of three or more pregnancies,most of which occur in early pregnancy.RSA has many pathogenic factors,such as thrombosis,immunity,heredity and anatomical abnormality.At present,the pathogenesis of recurrent miscarriage and the mechanism of pathogenesis are more complex.Homocysteine(Hcy)is a sulfhydryl amino acid.It is an important intermediate product in the process of methionine metabolism.A large number of clinical epidemiological and laboratory data show that the increase of Hcy is closely related to cardiovascular and cerebrovascular diseases and thrombotic diseases.Recent studies also suggest that elevated plasma Hcy level is a risk factor for recurrent spontaneous abortion.Therefore,through the detection of clinical samples,this paper will verify the relationship between plasma Hcy level and RSA.Through cell biology and molecular biological experiments,we will study the role of Hcy on vascular endothelial cells and explore the molecular mechanism of Hcy changes in RSA plasma.Object: 1.Normal pregnancy group: 40 cases were treated in the Department of gynaecology and obstetrics department of The Second Hospital Affiliated to Suzhou University for the same period.There was no history of spontaneous abortion or childbearing generation.The average age was(27.5 ± 4.3)years old.The increase of HCG in blood and B-mode ultrasound showed that pregnancy was confirmed by pregnancy and yolk sac.The average time of withdrawal was(54.4 ± 4.1)days.The following abnormality was excluded in the subjects of this study: 1.The history of spontaneous abortion or fetal parause,the history of basic diseases before;2.There is a history of basic disease before pregnancy;3.Pregnancy,and the signs of threatened abortion,such as vaginal bleeding and abdominal pain,were found in.2.RSA group: 40 cases were treated in the The Second Hospital Affiliated to Suzhou University reproductive center from January 01 to August 31,2017.In the past,three or more embryos with the same spouse had stopped developing abortion.The average age was(31.2 ± 5.8)years old,and the pregnancy sac and the embryo were observed during the visit.The pregnancy was confirmed by B-mode ultrasound.It was identified by ultrasound as empty pregnancy sac and embryo free fetal heart.The average time of stopping bleeding was(64.2 ± 6.7)days before the abortion.All the subjects included in the group were excluded from the following abnormalities before this pregnancy: 1.Malformation of the genital tract;2.There is a history of basic disease before pregnancy;3.Hypofibrinemia and thrombocytosis;4.Other autoantibodies,such as anti nuclear antibody and antiphospholipid antibody;5.Thyroid hormone anomaly and anti thyroid antibody positive;6.Prethrombosis state;7.Mycoplasma,Ureaplasma urealyticum and chlamydia of vaginal discharge were positive;8.The couples had abnormal karyotype.Method: 1.Peripheral blood was collected from normal pregnancy and RSA women.Plasma was centrifugally obtained and plasma Hcy concentration was detected by homocysteine kit.2.Labeling the Microparticles(MPs)with the antibody of cell surface marker.The total number of MPs and the number of Endothelial(EMPs)in normal pregnancy and RSA women plasma were analyzed by Flow cytometry(FCM).3.Nucleic acid combined with fluorescent dye PicoGreen was used to detect plasma DNA level in normal pregnancy and RSA women.4.HUVEC was stimulated with different concentration of Hcy,and the effect of Hcy on the growth of HUVEC was detected by CCK8.The number of positive cells was detected by FCM(Propidium iodide,PI),and the number of PI staining positive cells was detected by FCM.5.The levels of cytokines in normal pregnancy and RSA plasma were detected by flow cytometry.6.The villous tissues of normal pregnant women and RSA women were collected,and the Quantitative real-time polymerase chain reaction(qRT-PCR)method was used to detect the mRNA expression level of MTHFR,CBS,MAT1 A and BHMT.Results: 1.Compared with normal pregnant women,the plasma homocysteine concentration of RSA women increased significantly.2.Compared with normal pregnant women,the total number of plasma MPs and EMPs and the level of plasma DNA increased significantly in RSA women.There was a positive correlation between total plasma MPs and plasma free DNA level.3.HUVEC cells were treated with different concentrations of Hcy(0,1.5,1,2,4,8 mmol/L).Compared with the normal control group,the survival rate of HUVEC cells decreased significantly after Hcy stimulation,the number of PI positive cells increased significantly,and the number of EMPs released by HUVEC cells increased significantly.4.The plasma cytokines were detected and the plasma IL-2 and TNF-alpha(Th1 secretion)in RSA women were significantly higher than those of normal pregnant women.There was a Th1/Th2 immunoregulation imbalance and a bias to Th1.5.Real-tim quantiative reverse transcription PCR(q-PCR)was used to detect Hcy metabolism related protease in placental villi.It was found that the expression level of MAT1 A mRNA in villous tissue of RSA group was significantly lower than that of normal control group.Conclusion: Through the above experimental studies,we found that the plasma Hcy level in RSA women is elevated,and the high Hcy level may be an independent risk factor for RSA.The deficiency of placental villi MAT1 A mRNA in abortion patients may lead to the increase of Hcy during the metabolic process,which leads to the increase in plasma Hcy level in RSA women,to stimulate endothelial cell oxidative stress and to damage vascular endothelial cells.In addition,the imbalance of Th1/Th2 regulation may be one of the reasons for the occurrence of RSA.Therefore,this study elaborated the role and molecular mechanism of Hcy in recurrent spontaneous abortion,and provided new ideas and therapeutic targets for early diagnosis and prevention of RSA.