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ObjectiveTo investigate the regulatory effect of Shoutaiwan on oxidative stress and pyroptosis in lipopolysaccharide （LPS）-induced human extravillous trophoblast （HTR-8/SVneo） cells and provide a new direction for deciphering the mechanism of action of Shoutaiwan. MethodLPS （100 μg∙L-1） was used to induce the injury of HTR-8/SVneo cells （modeling）. Five groups were designed in this study， including a blank group， a model group， a Shoutaiwan （10% Shoutaiwan-containing serum） group， an antioxidant （1 mmol·L-1 NAC） group， and NOD like receptor thermoprotein domain 3 （NLRP3） inhibitor （50 μmol·L-1 MCC950） group. Cell viability was detected by cell counting kit-8 （CCK-8） kit. Hochest 33342/PI double fluorescence staining and flow cytometry were employed to observe cell death. The levels of interleukin-18 （IL-18）， interleukin-1β （IL-1β）， malondialdehyde （MDA）， and superoxide dismutase （SOD） in cell supernatant was determined by enzyme-linked immunosorbent assay （ELISA）. DCFH-DA probe was used to measure the level of intracellular reactive oxygen species （ROS）. Western blot was employed to determine the protein levels of NLRP3， Caspase-1， gastermin D （GSDMD）， and IL-1β in cells， and Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR） to measure the mRNA levels of NLRP3 and Caspase-1 in cells. ResultCompared with the blank group， the modeling decreased the cell viability （P<0.01）， elevated the levels of IL-1β， IL-18， ROS， and MDA， and weakened the activity of SOD （P<0.01）. Furthermore， it up-regulated the protein levels of NLRP3， Caspase-1， GSDMD， and IL-1β and the mRNA levels of NLRP3 and Caspase-1 （P<0.01）. Compared with the model group， Shoutaiwan， NAC， and MCC950 increased the cell viability （P<0.01）. Further， Shoutaiwan and NAC lowered the levels of MDA and ROS and increased the activity of SOD （P<0.01）. Shoutaiwan and MCC950 reduced the IL-1β and IL-18 in cell supernatant （P<0.01）， and down-regulated the protein levels of NLRP3， Caspase-1， GSDMD， and IL-1β and the mRNA levels of NLRP3， Caspase-1， and IL-1β （P<0.05，P<0.01）. ConclusionShoutaiwan can regulate oxidative stress and pyroptosis to attenuate the LPS-induced damage of HTR-8/SVneo cells， which may be the mechanism of Shoutaiwan in preventing recurrent spontaneous abortion.
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ObjectiveTo explore difference in the mechanism of Shoutaiwan， a representative kidney-tonifying and abortion-preventing formula， and Juyuanjian， a typical spleen-invigorating and abortion-preventing formula in reversing the pathology of decidua of spontaneous abortion （SA） patients and to expound the connotation of "uterine collaterals connecting kidney" and "fetal collaterals connecting spleen" theory. MethodThe targets of SA were retrieved from GeneCards， followed by gene ontology-biological process （GO-BP） annotation. Based on Cytoscape and previous research， the main processes and core targets were screened out. High-performance liquid chromatography-mass spectrometry （HPLC-MS） was used to identify the potential active components of Shoutaiwan and Juyuanjian and the regulatory networks were constructed. SA was induced in rats and the model rats were treated with Shoutaiwan and Juyuanjian at the same unit. Hematoxylin and eosin （HE） staining， transmission electron microscopy （TEM）， enzyme-linked immunosorbent assay （ELISA）， immunohistochemistry （IHC）， immunofluorescence （IF）， and other methods were employed to verify the mechanisms against miscarriage. ResultThe dysregulation of cell adhesion， inflammatory response， cell death， and angiogenesis was the core pathological process of SA. A total of 13 potential specific active components of Shoutaiwan and 14 active components of Juyuanjian were screened out. The regulatory networks showed that the potential active components of the two prescriptions modulated vascular endothelial growth factor （VEGF）， interleukin （IL）-2， estrogen receptor （ESR）-1， matrix metalloproteinase-9 （MMP-9）， and other targets to regulate the pathological process of SA. The two can significantly improve the pregnancy rate and the integrity rate and blood supply of decidua cells， control the apoptosis morphology and the expression of estrogen （E2）， progesterone （P）， and its receptor， and down-regulate the expression of MMP-2， MMP-9， IL-2， and IL-6 in decidua tissue of SA rats. At the same time， they up-regulated the expression of anti-apoptotic protein B-cell lymphoma-2 （Bcl-2） and IL-4. Shoutaiwan significantly up-regulated the expression of VEGF， and Juyuanjian significantly down-regulated the expression of E-cadherin （E-Cad）. ConclusionBoth Shoutaiwan and Juyuanjian regulate the core pathological process of SA to prevent miscarriage. At the same unit， Shoutaiwan is overall superior to Juyuanjian. Shoutaiwan is better than Juyuanjian in regulating angiogenesis and Juyuanjian is superior to Shoutaiwan in regulating cell adhesion. This conclusion can partly explain the biological basis of "treating the same disease with different methods"， and provide objective data reference for the identification of quality marker （Q-marker） of anti-miscarriage Chinese medicine and further study of formula-syndrome metabolome.
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Objective::To observe the efficacy of modified Shoutaiwan on recurrent abortion due to prethrombotic status with kidney deficiency and blood stasis and investigate its effect on coagulation factors. Method::The 100 cases were randomly divided into control group and observation group, 50 cases in each group. Bushen Yangxue capsule + enoxaparin sodium were given in control group, while modified Shoutaiwan + enoxaparin sodium were given in observation group, with a treatment course of 60 d in both groups. The serum clotting factor activated protein S (PS), activated protein C (PC), platelet aggregation function (PAF), platelet aggregation rate (PagT), antithrombin (AT), lupus anticoagulant material positive positive predictive value (LA), anti cardiolipin antibody positive rate of positive predictive value (ACA), plasminogen activator inhibitor-1(PAI-1), tissue factor (TF), fibrinogen (Fig), fibrinogen (FIB), fibrinogen alpha granule membrane glycoprotein-140 (GMP-140), fibrinogen fragment (F1+ 2), tissue fibrinogen activation factor (t-PA), D-dimer (D-D), activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) were observed. Clinical efficacy, pregnancy rate, traditional Chinese medicine(TCM) syndromes and adverse reactions were compared between the two groups. Result::4 cases withdrew from the study. The total effective rate in observation group was 97.9% (48/49), higher than 85.1% (40/47) in control group (χ2=5.713, P<0.05). The pregnancy rate in observation group was 89.8% (44/49), higher than 70.2% (33/47) in control group (χ2=6.351, P<0.05). The score of traditional Chinese medicine syndromes in observation group was lower than that in control group. The indexes of PS, PC, AT, TF, Fig, FPA, APTT and TT in observation group were significantly higher than those in control group (P<0.05), while PAF, PagT, LA, ACA, PAI-1, PAP, TAT, FIB, GMP-140, F1+ 2, t-PA, D-D and PT were lower than those in control group (P<0.05). The incidence of adverse reactions in observation group was 10.2% (5/49), lower than 35.4% (17/48) in control group (P<0.05). Conclusion::Modified Shoutaiwan can effectively improve the clinical symptoms of recurrent abortion due to prethrombotic status and improve the coagulation factors in patients with kidney deficiency and blood stasis.
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Objective:To observe the immune factors， coagulation and curative effect of modified Shoutaiwan with Si Junzitang combined with dydrogesterone tablets in advanced age patients with early threatened abortion， and to explore its mechanism of action. Method:The 90 advanced age patients with threatened abortion and kidney deficiency and blood stasis syndrome differentiation in traditional Chinese medicine （TCM） were randomly divided into control group and observation group by random number table， with 45 cases in each group. Both groups took oral dydrogesterone tablets for luteal support. The control group additionally received natural vitamin E soft capsules by oral administration， while observation group received modified Shoutaiwan with Si Junzitang. The course of treatment was 10 days in both groups. The clinical efficacy， TCM syndrome score， immune factors and coagulation factors of the two groups were compared before and after treatment. Result:There was no statistically significant difference in TCM symptom scores， immune factors， and coagulation factors between two groups before treatment. After treatment， the scores of TCM syndromes were reduced in both groups （P<0.05）， the proportion of helper T lymphocyte （Th）， Th/Ts ratio， D-dimer （D-D） level and fibrinogen （FIB） were reduced while prothrombin time （PT） and the ratio of suppressor T lymphocyte （Ts） were increased in observation group （P<0.05）. After treatment， the proportion of Th， Th/Ts， D-D， and FIB levels in observation group were lower than those in control group， while PT and the proportion of Ts were higher than those in control group （P<0.05）. The proportion of natural killer cells （NK） had no significant change after treatment， also with no significant difference between two groups. The total effective rate was 84.4%（38/45） in observation group， higher than 64.4%（29/45） in control group （χ2=4.398，P<0.05）. There was no obvious adverse reaction in both groups during the treatment. Conclusion:Modified Shoutaiwan with Si Junzitang combined with dydrogesterone tablets can improve symptoms and the therapeutic effect for fetal protection by regulating the immune balance and coagulation function in advanced age patients with threatened abortion.
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Objective: To observe the effect of Shoutaiwan and Shaoyao Gancao Tang on recurrent spontaneous abortion due to spleen and kidney Qi deficiency. Method: Totally 80 eligible patients were randomly divided into the control group and the treatment group, with 90 cases in each group. The control group was treated with lymphocyte immune; the Shoutaiwan and Shaoyao Gancao Tang combined with lymphocyte immune was applied in the treatment group. Coagulation function[the part plasminogen activator inhibitor-1 (PAI-1), fibrinogen (FIB), type plasminogen activator (t-PA), prothrombin time (PT), D-dimer (D-D) and tissue factor (TF), fibrin peptide A (FPA), fibrinogen (Fig), Plasminase-antifibrinolytic complex (PAP), glycoprotein platelet particle-140 (GMP-140), thrombin-antithrombin complex (TAT)], serum closed antibody (CD3+,CD4+,CD8+,CD4+/CD8+), inflammatory factor interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), Interleukin-1β (IL-1β), interleukin-10 (IL-10)]were scored after the treatment. The rates of birth and adverse reactions in each group were compared. The traditional Chinese medicine(TCM) syndrome differentiation of abortion spleen and kidney Qi deficiency was scored. Result: 5 cases were lost during the study period. The term delivery rate was 92.0%in the treatment group, which was higher than 73.6%in control group (PP+, CD4+, CD4+/CD8+), inflammatory factors (IFN-γ,TNF-α,IL-1β) and TCM syndrome scores were significantly lower in the treatment group than that in control group (P+), inflammatory factors IL-10 were significantly increased compared with the control group (PPConclusion: Shoutaiwan and Shaoyao Gancao Tang have a remarkable effect on recurrent spontaneous abortion case by spleen and kidney Yang deficiency.
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Objective To observe the effect of Shoutaiwan on the expression ofα-enolase in the decidua tissue of recurrent abortion mice. Methods The abortion-prone CBA/J × DBA/2 matings were established as the model of recurrent abortion and the nonabortion-prone CBA/J×BALB/C matings were used as the model of normal pregnancy. The model of recurrent abortion CBA/J × DBA/2 of pregnant mice were randomly divided into model group and Shoutaiwan high-, medium-, low-dose groups, pregnant mice of every group were orally administrated in different doses. On the 14th day of pregnancy, the mice were killed. The expression ofα-enolase was detected by using immunohistochemical method and Western Blot. Results α-enolase expression in the model group was significantly higher than the normal pregnancy group (P0.05). Conclusion Shoutaiwan could down-regulate the expression ofα-enolase in the decidua tissues of recurrent abortion mice, which may be one of its mechanisms of preventing miscarriage.
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Objective To study the effects of Shoutaiwan on the STAT signal transduction pathway of CD4+ T cells in maternal-fetal interface of mice with recurrent spontaneous abortion(RSA).Methods CBA/J?BALB/C mating combination and CBA/J?DBA/2 mating combination were used as normal pregnancy and RSA model respectively.Mice with RSA were randomly assigned to model group and Shoutaiwan group.After 14 days of administration,the STAT signal transduction pathway of CD4+ T cells in mice decidual and placental tissues were detected by Springbio 720 antibody microarrays.Results Shoutaiwan could increase the phosphorylation level of STAT3 and STAT6 protein in the STAT signal transduction pathway.At the same time,Shoutaiwan could decrease the phosphorylation level of STAT1 protein.Conclusion The activation of STAT3 and STAT6 signal transduction pathway might play a important role in the therapy of RSA by Shoutaiwan.
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Objective To observe the effect of Shoutaiwan on improving clinical pregnancy rate of in vitro fertilization-embryo transfer (IVF-ET). Method Eighty people, who were easy to be followed up and pregnant with the assistant of IVF-ET by the tube-baby centre of Shenzhen Urology Surgical Hospital of Zhongshan Medical University from 2001 to 2003, were divided into two groups randomly with 40 people for each. Treatment group, after embryo transfer, were normally injected progesterone and auxiliary treated of traditional Chinese Medicine according to syndrome differentiation. The comparing group were only injected progesterone. The curative effect was observed and compared. Result In 40 cases of treatment group, 23 cases were pregnant, clinical pregnant rate was 57.5%, the average level of progesterone was (22.45?1.31)?g/L, pregnant abortion rate was 13.0%. In 40 cases of comparing group, 12 cases were pregnant, clinical pregnant rate was 30.0%, the average level of progesterone was (10.18?0.89)?g/L, pregnant abortion rate was 25%. There were significant differences in the clinical pregnant statistical rate, the average level of progesterone and pregnant abortion rate between the two groups. Conclusion It’s more effective that the syndrome differentiation method based on Shoutaiwan than progesterone treatment only in improving clinical pregnant rate and reducing pregnant abortion rate of IVF-ET.