助孕3号方对自然流产T淋巴细胞亚群、NK细胞、Fas/FasL表达干预的研究
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摘要
目的
本课题通过研究助孕3号方对自然流产患者外周血T淋巴细胞亚群、NK细胞的影响以及助孕3号方对肾虚黄体抑制流产模型孕鼠外周血T淋巴细胞亚群、蜕膜组织Fas／FasL表达干预的影响，深入探讨自然流产的免疫学发病机制和中药助孕3号方安胎的机理。
方法
1．临床研究：所有受试病例必须按照诊断标准、纳入标准和排除标准进行筛选，选取早期自然流产脾肾虚型患者30例做为实验组与正常早孕妇女15例做为对照组，检测正常早孕妇女外周血中CD3+、CD4+、CD8+、CD4+／CD8+及NK细胞百分含量，并比较实验组服用助孕3号方前和服药两周后外周血中CD3+、CD4+、CD8+、CD4+／CD8+及NK细胞百分含量，进一步探讨助孕3号方治疗自然流产脾肾虚型的作用机理。
2．实验研究：以羟基脲+米非司酮建立肾虚黄体抑制病证结合流产模型。随机将46只雌性受孕SD大鼠分为4组：助孕3号方组、黄体酮组、模型组、正常妊娠组，通过检测这四组孕鼠孕第11天外周血中CD3+、CD4+、CD8+、CD4+／CD8+和蜕膜组织Fas和FasL的表达，进一步研究自然流产的发病机制，探讨助孕3号方治疗自然流产的免疫学作用机理。
研究结果
1．临床研究：实验组在服用助孕3号方前和对照组比较，CD3+变化无显著性差异(P＞0.05)；CD4+、CD4+／CD8+增高，有非常显著性差异(P＜0.01)；CD8+降低，有显著性差异(P＜0.05)；NK细胞百分含量增高，有显著性差异(P＜0.05)；血β-HCG定量明显降低，有非常显著性差异(P＜0.01)。服用助孕3号方两周后，治疗组30例，其中29例有效，1例妊娠维持超过既往流产天数后再次自然流产，总有效率96.7％。实验组用药后比用药前CD3+变化无显著性差异(P＞0.05)；用药后CD8+增高，CD4+、CD4+／CD8+、NK细胞百分含量降低，有非常显著性差异(P＜0.01)；血β-HCG定量用药后比用药前明显升高，有非常显著性差异(P＜0.01)。
2．实验研究：四组比较，CD3+、CD8+的差别无显著性差异(P＞0.05)。造模组CD4+、CD4+／CD8+最高，与其他三组比较有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与造模组比较，CD4+、CD4+／CD8+明显降低，有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与正常妊娠组比较，CD4+、CD4+／CD8+无显著性差异(P＞0.05)。造模组与正常妊娠组比较，蜕膜组织Fas表达的平均染色指数明显增高，有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与造模组比较Fas表达均明显减低，有明显的差异性(P＜0.01)。助孕3号组、黄体酮组与正常妊娠组比较，Fas变化不明显，差异不具有显著性(P＞0.05)。造模组与正常妊娠组比较，FasL表达的平均染色指数降低，有显著性差异(P＜0.05)。助孕3号组、黄体酮组与造模组比较FasL表达均增高，有显著性差异(P＜0.05)。助孕3号组、黄体酮组和正常妊娠组比较，FasL染色强度指数变化不明显，差异不具有显著性(P＞0.05)。
结论
1．早期自然流产患者外周血中CD4+含量和CD4+／CD8+比值明显上升，CD8+含量下降，NK细胞百分数增多。T淋巴细胞亚群比例的失调，外周血NK细胞含量的升高可能是自然流产的发病原因之一。本研究显示，补肾健脾复方助孕3号方可以使早期自然流产患者外周血中CD8+增高、使CD4+、CD4+／CD8+和NK细胞含量降低。
2．肾虚黄体抑制流产孕鼠与正常早孕大鼠比较，外周血中CD3+、CD8+无明显变化，CD4+、CD4+／CD8+有明显升高，蜕膜组织的Fas表达增高，FasL表达降低，提示早期流产淋巴细胞亚群比率失调，细胞免疫功能过度，Fas／FasL介导的AICD机制平衡失调，导致妊娠丢失。
3．助孕3号方可以降低肾虚黄体抑制流产孕鼠外周血CD4+、CD4+／CD8+，纠正淋巴细胞亚群比率失调，还可以增加蜕膜组织FasL表达、降低Fas表达。由此推出，助孕3号方可能通过纠正淋巴细胞亚群比率失调，恢复CD8+T淋巴细胞和CD4+T淋巴细胞在机体免疫应答过程中的免疫协调作用，并通过增加蜕膜组织FasL表达，降低Fas表达，使受异体抗原刺激增生的Fas阳性T淋巴细胞及时凋亡，Pas／FasL介导的AICD机制恢复平衡协调，从而起到安胎的作用。
Objective To research the Prescription of Zhuyun-Ⅲon T lymphocyte subset、NK cell、Fas/FasL of Spontaneous Abortion patient.To observe the effect ofprescription Zhuyun-Ⅲon T lymphocyte subset、NK cell、Fas/FasL ofSpontaneous Abortion rats by set up the animal model of renal-deficiency andluteal insufficiency abortion rats, investigate immunology nosogenesis ofSpontaneous Abortion and tocolysis mechanism of Zhuyun-Ⅲ.
Method
Clinical Research:According to internalize criteria, experiment groupconcluded 30 patients who were diagnosed as spontaneous abortion of both kidneyand spleen deficiency in the first trimester. Control group concluded 15 normalwomen in early pregnancy who requested artificial abortion. To dicover themechanism of action of Zhuyun-Ⅲon spontaneous abortion of both kidney andspleen deficiency, the content of CD3~+、CD4~+、CD8~+、CD4~+/CD8~+、NK cell in bloodof 15 normal pregnancy women were not only detected but also of 30 spontaneoussuffers by before and after two weeks of therapy with Zhuyun-Ⅲ.
Experimental Research: Setting up the animal model of renal-deficiency andluteal-insufficiency abortion rats with Hydrea and Mifepristone and groupingthem at random in 4 teams: model with Zhuyun-Ⅲ、model with flavolutan、modelwithout herbs、controlled group.
Comparing the content of CD3~+、CD4~+、CD8~+、CD4~+/CD8~+ and expression of Fas/FasLin the rats' blood and counting the rate of abortion, to investigate themechanism of spontaneous abortion and the immunology mechanism of action ofPrescription on Zhuyun-Ⅲ.
Result
Clinic study: Comparing with before and of after therapy withPrescription on Zhuyun-Ⅲof experiment group, variation of CD3+ is notsignificantly (P＞0.05). Comparing to control group heighten significantly (P＜0.01). Comparing to control group degrade significantly (P＜0.05).Comparing to control group odds ratio of odds ratio heighten significantly(P＜0.01). Comparing to control group heighten significantly (P＜0.05).Comparing to control groupβ-hCG of quantitative experiment group in blooddegrade significantly (P＜0.01). With Prescription Zhuyun-Ⅲtwo weeks later,there were 29 effective cases in the control group and 1 case which happenedSpontaneous Abortion again with total effective rate 96.7%.Comparing withbefore and of after therapy with Prescription on Zhuyun-Ⅲof experiment groupratio of CD3~+ didn't (P＞0.05).After prescription Zhuyun-Ⅲratio of CD8~+heighten significantly (P＜0.01), while ratio of CD4~+ and CD4~+/CD8~+ and percentcontent of NK cells degrade significantly (P＜0.01).
Experimental study:Comparing with other 3 groups CD3~+ is the highest incontrolled group which isn't significant (P＜0.05), while CD4~+ and CD4~+/CD8~+is the highest in controlled group which is significant(P＜0.01)and CD8~+ don' tdegrade significantly(P＞0.05).Comparing to controlled group CD4~+ and CD4~+/CD8~+degrade significantly in the model with Zhuyun-Ⅲand flavolutan (P＜0.01).Comparing with model without herbs and model with Zhuyun-Ⅲand flavolutanCD4~+ and CD4~+/CD8~+ area' t significantly (P＞0.05).Comparing to model withoutherbs index of average dyeing intensity of expression of index FasL degradesignificantly and Fas heighten significantly in the controlled group(P＜0.05). Comparing to controlled group index ofexpression of FasL heightensignificantly and Fas degrade significantly in the model with flavolutan(P＜0.05). Comparing to model without herbs index of dyeing intensity indexof expression of FasL don' t change significantly in the model with Zhuyun-Ⅲand flavolutan (P＞0.05).
Conclusion
1. It was conformed that content of CD4~+ and ratio of CD4~+/CD8~+ raiseobviously,with decreased content of CD8~+ and raised percentage of NK cellsspontaneous abortion in the first trimester.The research also demonstratedthat imbalance of proportional T lymphocyte subset and raised percentage ofNK cells might be one reason of spontaneous abortion.
2.1t was disclosed that Comparing to normal rats in early pregnancythere has been not marked change for CD3~+, while content of CD4~+ and ratio ofCD4~+/CD8~+ raise obviously and index of positive staining of expression of FasLheighten significantly in renal-deficiency and luteal-insufficiency abortion rats. It also provide that proportion disturbance of lymphocyte subset andexcess of immunologic function land raised content of CD4~+ and ratio of CD4~+/CD8~+ lead to spontaneous abortion in the first trimester.
3. The research indicated that Prescription Zhuyun-Ⅲcan invigorate thekidney and spleen, so that it can retrieved the proportion disorder oflymphocyte subset by decreasing the content of CD4~+ and ratio of CD4~+/CD8~+tomake CD4~+ and CD8~+ T lymphocyte educe immune coordination in immune responseof body.It also provided the scientific evidence that Prescription Zhuyun-Ⅲcan heighten expression of FasL of decidua tissue in rats which werespontaneous abortion in the first trimester to make Fas lymphocyte apoptosispromptly so that protect nutrient cell from aggression of lymphocyte andcontinue gestation.
本课题通过研究助孕3号方对自然流产患者外周血T淋巴细胞亚群、NK细胞的影响以及助孕3号方对肾虚黄体抑制流产模型孕鼠外周血T淋巴细胞亚群、蜕膜组织Fas／FasL表达干预的影响，深入探讨自然流产的免疫学发病机制和中药助孕3号方安胎的机理。
方法
1．临床研究：所有受试病例必须按照诊断标准、纳入标准和排除标准进行筛选，选取早期自然流产脾肾虚型患者30例做为实验组与正常早孕妇女15例做为对照组，检测正常早孕妇女外周血中CD3+、CD4+、CD8+、CD4+／CD8+及NK细胞百分含量，并比较实验组服用助孕3号方前和服药两周后外周血中CD3+、CD4+、CD8+、CD4+／CD8+及NK细胞百分含量，进一步探讨助孕3号方治疗自然流产脾肾虚型的作用机理。
2．实验研究：以羟基脲+米非司酮建立肾虚黄体抑制病证结合流产模型。随机将46只雌性受孕SD大鼠分为4组：助孕3号方组、黄体酮组、模型组、正常妊娠组，通过检测这四组孕鼠孕第11天外周血中CD3+、CD4+、CD8+、CD4+／CD8+和蜕膜组织Fas和FasL的表达，进一步研究自然流产的发病机制，探讨助孕3号方治疗自然流产的免疫学作用机理。
研究结果
1．临床研究：实验组在服用助孕3号方前和对照组比较，CD3+变化无显著性差异(P＞0.05)；CD4+、CD4+／CD8+增高，有非常显著性差异(P＜0.01)；CD8+降低，有显著性差异(P＜0.05)；NK细胞百分含量增高，有显著性差异(P＜0.05)；血β-HCG定量明显降低，有非常显著性差异(P＜0.01)。服用助孕3号方两周后，治疗组30例，其中29例有效，1例妊娠维持超过既往流产天数后再次自然流产，总有效率96.7％。实验组用药后比用药前CD3+变化无显著性差异(P＞0.05)；用药后CD8+增高，CD4+、CD4+／CD8+、NK细胞百分含量降低，有非常显著性差异(P＜0.01)；血β-HCG定量用药后比用药前明显升高，有非常显著性差异(P＜0.01)。
2．实验研究：四组比较，CD3+、CD8+的差别无显著性差异(P＞0.05)。造模组CD4+、CD4+／CD8+最高，与其他三组比较有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与造模组比较，CD4+、CD4+／CD8+明显降低，有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与正常妊娠组比较，CD4+、CD4+／CD8+无显著性差异(P＞0.05)。造模组与正常妊娠组比较，蜕膜组织Fas表达的平均染色指数明显增高，有非常显著性差异(P＜0.01)。助孕3号组、黄体酮组与造模组比较Fas表达均明显减低，有明显的差异性(P＜0.01)。助孕3号组、黄体酮组与正常妊娠组比较，Fas变化不明显，差异不具有显著性(P＞0.05)。造模组与正常妊娠组比较，FasL表达的平均染色指数降低，有显著性差异(P＜0.05)。助孕3号组、黄体酮组与造模组比较FasL表达均增高，有显著性差异(P＜0.05)。助孕3号组、黄体酮组和正常妊娠组比较，FasL染色强度指数变化不明显，差异不具有显著性(P＞0.05)。
结论
1．早期自然流产患者外周血中CD4+含量和CD4+／CD8+比值明显上升，CD8+含量下降，NK细胞百分数增多。T淋巴细胞亚群比例的失调，外周血NK细胞含量的升高可能是自然流产的发病原因之一。本研究显示，补肾健脾复方助孕3号方可以使早期自然流产患者外周血中CD8+增高、使CD4+、CD4+／CD8+和NK细胞含量降低。
2．肾虚黄体抑制流产孕鼠与正常早孕大鼠比较，外周血中CD3+、CD8+无明显变化，CD4+、CD4+／CD8+有明显升高，蜕膜组织的Fas表达增高，FasL表达降低，提示早期流产淋巴细胞亚群比率失调，细胞免疫功能过度，Fas／FasL介导的AICD机制平衡失调，导致妊娠丢失。
3．助孕3号方可以降低肾虚黄体抑制流产孕鼠外周血CD4+、CD4+／CD8+，纠正淋巴细胞亚群比率失调，还可以增加蜕膜组织FasL表达、降低Fas表达。由此推出，助孕3号方可能通过纠正淋巴细胞亚群比率失调，恢复CD8+T淋巴细胞和CD4+T淋巴细胞在机体免疫应答过程中的免疫协调作用，并通过增加蜕膜组织FasL表达，降低Fas表达，使受异体抗原刺激增生的Fas阳性T淋巴细胞及时凋亡，Pas／FasL介导的AICD机制恢复平衡协调，从而起到安胎的作用。
Objective To research the Prescription of Zhuyun-Ⅲon T lymphocyte subset、NK cell、Fas/FasL of Spontaneous Abortion patient.To observe the effect ofprescription Zhuyun-Ⅲon T lymphocyte subset、NK cell、Fas/FasL ofSpontaneous Abortion rats by set up the animal model of renal-deficiency andluteal insufficiency abortion rats, investigate immunology nosogenesis ofSpontaneous Abortion and tocolysis mechanism of Zhuyun-Ⅲ.
Method
Clinical Research:According to internalize criteria, experiment groupconcluded 30 patients who were diagnosed as spontaneous abortion of both kidneyand spleen deficiency in the first trimester. Control group concluded 15 normalwomen in early pregnancy who requested artificial abortion. To dicover themechanism of action of Zhuyun-Ⅲon spontaneous abortion of both kidney andspleen deficiency, the content of CD3~+、CD4~+、CD8~+、CD4~+/CD8~+、NK cell in bloodof 15 normal pregnancy women were not only detected but also of 30 spontaneoussuffers by before and after two weeks of therapy with Zhuyun-Ⅲ.
Experimental Research: Setting up the animal model of renal-deficiency andluteal-insufficiency abortion rats with Hydrea and Mifepristone and groupingthem at random in 4 teams: model with Zhuyun-Ⅲ、model with flavolutan、modelwithout herbs、controlled group.
Comparing the content of CD3~+、CD4~+、CD8~+、CD4~+/CD8~+ and expression of Fas/FasLin the rats' blood and counting the rate of abortion, to investigate themechanism of spontaneous abortion and the immunology mechanism of action ofPrescription on Zhuyun-Ⅲ.
Result
Clinic study: Comparing with before and of after therapy withPrescription on Zhuyun-Ⅲof experiment group, variation of CD3+ is notsignificantly (P＞0.05). Comparing to control group heighten significantly (P＜0.01). Comparing to control group degrade significantly (P＜0.05).Comparing to control group odds ratio of odds ratio heighten significantly(P＜0.01). Comparing to control group heighten significantly (P＜0.05).Comparing to control groupβ-hCG of quantitative experiment group in blooddegrade significantly (P＜0.01). With Prescription Zhuyun-Ⅲtwo weeks later,there were 29 effective cases in the control group and 1 case which happenedSpontaneous Abortion again with total effective rate 96.7%.Comparing withbefore and of after therapy with Prescription on Zhuyun-Ⅲof experiment groupratio of CD3~+ didn't (P＞0.05).After prescription Zhuyun-Ⅲratio of CD8~+heighten significantly (P＜0.01), while ratio of CD4~+ and CD4~+/CD8~+ and percentcontent of NK cells degrade significantly (P＜0.01).
Experimental study:Comparing with other 3 groups CD3~+ is the highest incontrolled group which isn't significant (P＜0.05), while CD4~+ and CD4~+/CD8~+is the highest in controlled group which is significant(P＜0.01)and CD8~+ don' tdegrade significantly(P＞0.05).Comparing to controlled group CD4~+ and CD4~+/CD8~+degrade significantly in the model with Zhuyun-Ⅲand flavolutan (P＜0.01).Comparing with model without herbs and model with Zhuyun-Ⅲand flavolutanCD4~+ and CD4~+/CD8~+ area' t significantly (P＞0.05).Comparing to model withoutherbs index of average dyeing intensity of expression of index FasL degradesignificantly and Fas heighten significantly in the controlled group(P＜0.05). Comparing to controlled group index ofexpression of FasL heightensignificantly and Fas degrade significantly in the model with flavolutan(P＜0.05). Comparing to model without herbs index of dyeing intensity indexof expression of FasL don' t change significantly in the model with Zhuyun-Ⅲand flavolutan (P＞0.05).
Conclusion
1. It was conformed that content of CD4~+ and ratio of CD4~+/CD8~+ raiseobviously,with decreased content of CD8~+ and raised percentage of NK cellsspontaneous abortion in the first trimester.The research also demonstratedthat imbalance of proportional T lymphocyte subset and raised percentage ofNK cells might be one reason of spontaneous abortion.
2.1t was disclosed that Comparing to normal rats in early pregnancythere has been not marked change for CD3~+, while content of CD4~+ and ratio ofCD4~+/CD8~+ raise obviously and index of positive staining of expression of FasLheighten significantly in renal-deficiency and luteal-insufficiency abortion rats. It also provide that proportion disturbance of lymphocyte subset andexcess of immunologic function land raised content of CD4~+ and ratio of CD4~+/CD8~+ lead to spontaneous abortion in the first trimester.
3. The research indicated that Prescription Zhuyun-Ⅲcan invigorate thekidney and spleen, so that it can retrieved the proportion disorder oflymphocyte subset by decreasing the content of CD4~+ and ratio of CD4~+/CD8~+tomake CD4~+ and CD8~+ T lymphocyte educe immune coordination in immune responseof body.It also provided the scientific evidence that Prescription Zhuyun-Ⅲcan heighten expression of FasL of decidua tissue in rats which werespontaneous abortion in the first trimester to make Fas lymphocyte apoptosispromptly so that protect nutrient cell from aggression of lymphocyte andcontinue gestation.
引文
[1] 余传霖，主编．现代医学免疫学．第1版．上海：上海医科大学出版社，1998：985．
[2] 张菱，宫美轩，张亚杰，王建业．早期自然流产病人外周血T淋巴细胞亚群的检测．齐鲁医学杂志，2005：20(4)：324—326．
[3] 李昭荣，刘明，吕海侠．等．原因不明性习惯性流产孕早期免疫治疗前后细胞免疫功能的变化 陕西医学杂志，2002：31(5)，490—492．
[4] 崔澄，白小嘉，刘水文，等．妊娠早期自然流产患者蜕膜淋巴细胞功能的变化 白求恩医学院学报，2004；2(2)：70—72．
[5] 崔徵，王惠贤，焦炳忠，等．不明原因早期自然流产患者蜕膜淋巴细胞CD4+／CD8+比值升高．河北医药，1999；21(5)：347—349．
[6] 毛立群，杨鹏，闫燕华，潘春煦．复发性流产患者外周血T淋巴细胞亚群及CD+4辅助性T淋巴细胞亚型的变化．中华妇产科杂志，2000；35(12)：722-723．
[7] 曾品鸿，丘彦，桂文武，刘敬，等．主动免疫治疗复发性自然流产的免疫功能监测．重庆医科大学学报，2003；28(4)：300-302．
[8] 梁义娟，冯丽，王淑梅，等．原因不明性习惯性流产细胞免疫功能变化及妊娠后主动免疫治疗．河北职工医学院学报，1999；16(1)，21-22．
[9] 陈巧儿，秦卫兵，欧汝强，等．免疫治疗原因不明习惯性流产后外周血T淋巴细胞亚群的变化．实用妇产科杂志，2005；21(2)：84-86．
[10] ChristiansenOB, NielsenHS, Pedersen B. Active or passive immunization in unexplained recurrent miscarriage. J ReprodImmunol, 2004; 62:41-52.
[11] Pandey MK, Agrawa IS. Induction of MLR-Bf and protection of fetal loss: a current double blind Randomized trial of paternal lymphocyte immunization for women with recurrent spontaneous abortion. International Immunopharmacology, 2004; 4:289-298.
[12] Gendron RL, Baines MG. Infiltrating deciduas natural killer cells are associated with spontaneous abortion in mice. Cell Immuol, 1988; 113-261.
[13] Ho HN, Chao KH, Chen CK, et al. Activation status of T and NK cells in the endometrium throughout menstrual cycles and normal and abnormal early pregnancy. Human Immuno1,1996; 49:130-6.
[14] Yamamoto T, Takahashi Y, Kase N, Mori H. Role of decidual natural killer cells in patients with missed abortion: differences between cases with normal and abnormal chromosome. Clan Exp Immunol, 1991; 116(3):449-52.
[15] 李敏，高瑞珍，赵淑霞，王士彦，齐向红，孙春荣，秦英．自然流产患者外周血与蜕 膜组自然杀伤细胞亚群数量变化的研究．中国优生与遗传杂志，2001；9(6)：50-53．
[16] 周龙书，黄立萍，蒋继娟．难免流产者蜕膜内自然杀伤细胞及淋巴细胞的活性。贵阳医学院学报，2002；27(1)：11．
[17] Coulam-CB, Goodman-C, et al. Systemic CD56+cells can predict pregnancy outcome. Am-J-Reprod-Immunol, 1995; 33(1):40-6.
[18] 赵卫秀，林其德．正常妊娠早期孕妇外周血自然杀伤细胞亚群的变化．中华妇产科杂志，1998；33：362．
[19] 陈巧儿，秦卫兵，杨宁，叶嘉玲．免疫治疗原因不明反复自然流产后Th1／Th2型细胞因子和NK细胞的变化．中国妇幼保健，2005：20(20)：2637—2639．
[20] 余进胜，彭来勤，张丙忠．复发性自然流产患者NK细胞活性变化的研究．广州医药，2004：35(3)35—37．
[21] Aoki K et al. Lancet, 1995; 345: 1340-1342.
[22] Kwak JY et al Am J Reprod Immnmol, 1996; 35(4): 363—369.
[23] Medena JP, Scaffidi C, Kischkel ME, et al. FLICE is activated by association with the CD95 death-inducing signaling complex[J]. EMBO J. 1997; 16: 2794.
[24] 刘玉昆，张建平，邝健全，等．Fas／FasL在复发性自然流产患者中的表达．中山大学学报(医学科学版)，2004：25(6)：581-585．
[25] Kauma-SW, Huff-TF, Hayes-N. Placental Fas Ligand expression is a mechanism for meternal immune tolerance to the fetus[J]. J lin ndocrino Metab. 1999; 84(6):2188-2194.
[26] Ejima-g, Koji-T, Tsuruta-D, et al. Induction of apoptosis in placentas of pregnant mice exposed to lipopolysaccharides: possible involvement of Fas/Fas ligand system[J]. Biol Reprod. 2000 Jan; 62(1): 178-185.
[27] Zorzi W, Thellin O, Coumans B, et al. Demonstration of the expression of CD95 ligand transcript and protein in human placenta. Placenta, 1998; 19(4):269-277.
[28] Hammer A, Blaschitz A, Daxbock C, et al. Fas and Fas-ligand are expressed in the uteroplacental unit of first-trimester pregnancy. Am J Reprod Immunol, 1999; 41(1): 41-51.
[29] Roh CR, lxe JW, Kang BH, et al. Differential expressions of Fas and Fas ligand in human placenta. J Korean Med Sci, 2002; 17(2):213-216.
[30] Ohshima K, Nakashinma M, Sonoda K, et al. Expression of RCASI and Fasl in human trophoblasts and uterine glands during pregnancy: the possible role in immune privilege. Clin Exp lmmunol, 2001; 123(3):481-486.
[31] Choi HK, Choi RC, Lee SH, et al. Expression of angiogenesis-and apoptosis-related genes in chorionic villi derived from recurrent pregnancy loss patients. Mol Reprod Dev, 2003: 66(1): 24-31.
[32] 赵延楼．逐瘀法治疗滑胎21例．辽宁中医杂志，1986, (9)：26．
[33] 宋文武．宋鸿之老中医治疗习惯性流产经验，河南中医1995：15(4)：228-229．
[34] 哈荔田著．哈荔田妇科医案医话选．天津：天津科学技术出版社，1982：17．
[35] 罗元恺，夏桂成等．习惯性流产的防治．中医杂，1998：(4)：5．
[36] 姚宣芬．中西医结合治疗习惯性流产22例．湖南中医学院学报，1998：8(2)：24—25。
[37] 马宝璋．试论安胎必补肾兼予调养气血．中医药学报，1998；(1)：34-35．
[38] 苏华林．治疗先兆流产50例临床分析．四川中医．1995；(10)：38．
[39] 赵广安，宋惠珍，张立华．玉屏风散治疗习惯性流产36例．实用中西医结合杂志．1993，(7)：438．
[40] 程航，李祥云．助黄保胎方治疗先兆流产115例．陕西中医．1998：(6)：241．
[41] 何瑞华．泰山磐石散加减治疗胎漏36例．浙江中医学院学报，1996；(1)：25．
[42] 张永霖．加味泰山磐石散治疗习惯性流产27例．江苏中医，1997：18(10)：17．
[43] 段世彪．安奠二天汤加减治疗先兆流产50例．湖北中医杂志，1987；(1)：13．
[44] 沈宗华．自拟补肾益气汤治疗先兆流产的体会．江西中医药，1987：(3)：24．
[45] 叶宝贵，陈伟民．自拟保胎汤治疗先兆流产60例．浙江中医杂志，1986：(2)：60．
[46] 刘家磊．羊肾固冲饮治疗滑胎181例．浙江中医杂志，1991；(12)：558．
[47] 张传芳，舒沪英，黄光英．补肾益气养血法治疗先兆流产53例临床观察．同济医科大学学报，1990(3)：210．
[48] 蒋俭，李光荣，藏美佳，活血化瘀法治疗血瘀型胎漏胎动不安滑胎41例临床观察．中医杂志，1992；(11)：30—32．
[49] 归绥琪．封闭抗体缺乏性自然流产者的中药治疗．上海医科大学学报，1997；24，(3)：217—219．
[50] 吴宁．中药复方优生宁对先兆流产病人NK细胞活性的影响．中医药信息2002：19(2)：59-60．
[51] 景苏玉，归绥棋．“保胎Ⅰ冲剂”提高反复自然流产患者免疫功能的临床观察 上海中医药杂志，2002：9：23-24．
[52] 索玉平．海马生髓丸对复发性流产的免疫调节作用研究．中医药学刊．2003；21(9)：1532-1533．
[53] 胡昌东，归绥琪，陆利民．中药治疗溴隐亭致大鼠流产模型中内分泌对蜕膜TNF-α、Fas的影响．中华中医药杂志，2005；20(1)：36-39．
[54] 许钧，归绥琪．自然流产病因分类与中医证型相关性研究．上海中医药杂志，2003； 37(3)：10-11．
[55] 赵颖，张玉珍．助孕3号丸治疗脾肾虚证胎漏、胎动不安、滑胎疗效观察。新中医，1999：31(12)：20—21。
[56] 韩惠，罗颂平．助孕3号方对大鼠子宫收缩活动的影响．广州中医药大学学报，2002；19(1)；33-35．
[57] 李小球，罗颂平．助孕3号丸对妊娠大鼠孕激素及受体的影响．暨南大学学报(医学版)，1999：20(6)：111-113．
[58] 罗颂平，梁国珍，张玉珍，等．助孕3号方防治大鼠自然流产的机理研究．中国中西医结合杂志，2003：23(7)：522-525．
[59] 宁艳，罗颂平，梁国珍，补肾健脾中药复方对肾虚黄体抑制妊娠大鼠流产模型的实验研究．中国医药学报，2000；15(6)：428-430．
[60] 曾诚，岳明明，试论先兆流产的中医证型分布规律．中国中医药信息杂志，2002；9(8)：5-7．
[2] 张菱，宫美轩，张亚杰，王建业．早期自然流产病人外周血T淋巴细胞亚群的检测．齐鲁医学杂志，2005：20(4)：324—326．
[3] 李昭荣，刘明，吕海侠．等．原因不明性习惯性流产孕早期免疫治疗前后细胞免疫功能的变化 陕西医学杂志，2002：31(5)，490—492．
[4] 崔澄，白小嘉，刘水文，等．妊娠早期自然流产患者蜕膜淋巴细胞功能的变化 白求恩医学院学报，2004；2(2)：70—72．
[5] 崔徵，王惠贤，焦炳忠，等．不明原因早期自然流产患者蜕膜淋巴细胞CD4+／CD8+比值升高．河北医药，1999；21(5)：347—349．
[6] 毛立群，杨鹏，闫燕华，潘春煦．复发性流产患者外周血T淋巴细胞亚群及CD+4辅助性T淋巴细胞亚型的变化．中华妇产科杂志，2000；35(12)：722-723．
[7] 曾品鸿，丘彦，桂文武，刘敬，等．主动免疫治疗复发性自然流产的免疫功能监测．重庆医科大学学报，2003；28(4)：300-302．
[8] 梁义娟，冯丽，王淑梅，等．原因不明性习惯性流产细胞免疫功能变化及妊娠后主动免疫治疗．河北职工医学院学报，1999；16(1)，21-22．
[9] 陈巧儿，秦卫兵，欧汝强，等．免疫治疗原因不明习惯性流产后外周血T淋巴细胞亚群的变化．实用妇产科杂志，2005；21(2)：84-86．
[10] ChristiansenOB, NielsenHS, Pedersen B. Active or passive immunization in unexplained recurrent miscarriage. J ReprodImmunol, 2004; 62:41-52.
[11] Pandey MK, Agrawa IS. Induction of MLR-Bf and protection of fetal loss: a current double blind Randomized trial of paternal lymphocyte immunization for women with recurrent spontaneous abortion. International Immunopharmacology, 2004; 4:289-298.
[12] Gendron RL, Baines MG. Infiltrating deciduas natural killer cells are associated with spontaneous abortion in mice. Cell Immuol, 1988; 113-261.
[13] Ho HN, Chao KH, Chen CK, et al. Activation status of T and NK cells in the endometrium throughout menstrual cycles and normal and abnormal early pregnancy. Human Immuno1,1996; 49:130-6.
[14] Yamamoto T, Takahashi Y, Kase N, Mori H. Role of decidual natural killer cells in patients with missed abortion: differences between cases with normal and abnormal chromosome. Clan Exp Immunol, 1991; 116(3):449-52.
[15] 李敏，高瑞珍，赵淑霞，王士彦，齐向红，孙春荣，秦英．自然流产患者外周血与蜕 膜组自然杀伤细胞亚群数量变化的研究．中国优生与遗传杂志，2001；9(6)：50-53．
[16] 周龙书，黄立萍，蒋继娟．难免流产者蜕膜内自然杀伤细胞及淋巴细胞的活性。贵阳医学院学报，2002；27(1)：11．
[17] Coulam-CB, Goodman-C, et al. Systemic CD56+cells can predict pregnancy outcome. Am-J-Reprod-Immunol, 1995; 33(1):40-6.
[18] 赵卫秀，林其德．正常妊娠早期孕妇外周血自然杀伤细胞亚群的变化．中华妇产科杂志，1998；33：362．
[19] 陈巧儿，秦卫兵，杨宁，叶嘉玲．免疫治疗原因不明反复自然流产后Th1／Th2型细胞因子和NK细胞的变化．中国妇幼保健，2005：20(20)：2637—2639．
[20] 余进胜，彭来勤，张丙忠．复发性自然流产患者NK细胞活性变化的研究．广州医药，2004：35(3)35—37．
[21] Aoki K et al. Lancet, 1995; 345: 1340-1342.
[22] Kwak JY et al Am J Reprod Immnmol, 1996; 35(4): 363—369.
[23] Medena JP, Scaffidi C, Kischkel ME, et al. FLICE is activated by association with the CD95 death-inducing signaling complex[J]. EMBO J. 1997; 16: 2794.
[24] 刘玉昆，张建平，邝健全，等．Fas／FasL在复发性自然流产患者中的表达．中山大学学报(医学科学版)，2004：25(6)：581-585．
[25] Kauma-SW, Huff-TF, Hayes-N. Placental Fas Ligand expression is a mechanism for meternal immune tolerance to the fetus[J]. J lin ndocrino Metab. 1999; 84(6):2188-2194.
[26] Ejima-g, Koji-T, Tsuruta-D, et al. Induction of apoptosis in placentas of pregnant mice exposed to lipopolysaccharides: possible involvement of Fas/Fas ligand system[J]. Biol Reprod. 2000 Jan; 62(1): 178-185.
[27] Zorzi W, Thellin O, Coumans B, et al. Demonstration of the expression of CD95 ligand transcript and protein in human placenta. Placenta, 1998; 19(4):269-277.
[28] Hammer A, Blaschitz A, Daxbock C, et al. Fas and Fas-ligand are expressed in the uteroplacental unit of first-trimester pregnancy. Am J Reprod Immunol, 1999; 41(1): 41-51.
[29] Roh CR, lxe JW, Kang BH, et al. Differential expressions of Fas and Fas ligand in human placenta. J Korean Med Sci, 2002; 17(2):213-216.
[30] Ohshima K, Nakashinma M, Sonoda K, et al. Expression of RCASI and Fasl in human trophoblasts and uterine glands during pregnancy: the possible role in immune privilege. Clin Exp lmmunol, 2001; 123(3):481-486.
[31] Choi HK, Choi RC, Lee SH, et al. Expression of angiogenesis-and apoptosis-related genes in chorionic villi derived from recurrent pregnancy loss patients. Mol Reprod Dev, 2003: 66(1): 24-31.
[32] 赵延楼．逐瘀法治疗滑胎21例．辽宁中医杂志，1986, (9)：26．
[33] 宋文武．宋鸿之老中医治疗习惯性流产经验，河南中医1995：15(4)：228-229．
[34] 哈荔田著．哈荔田妇科医案医话选．天津：天津科学技术出版社，1982：17．
[35] 罗元恺，夏桂成等．习惯性流产的防治．中医杂，1998：(4)：5．
[36] 姚宣芬．中西医结合治疗习惯性流产22例．湖南中医学院学报，1998：8(2)：24—25。
[37] 马宝璋．试论安胎必补肾兼予调养气血．中医药学报，1998；(1)：34-35．
[38] 苏华林．治疗先兆流产50例临床分析．四川中医．1995；(10)：38．
[39] 赵广安，宋惠珍，张立华．玉屏风散治疗习惯性流产36例．实用中西医结合杂志．1993，(7)：438．
[40] 程航，李祥云．助黄保胎方治疗先兆流产115例．陕西中医．1998：(6)：241．
[41] 何瑞华．泰山磐石散加减治疗胎漏36例．浙江中医学院学报，1996；(1)：25．
[42] 张永霖．加味泰山磐石散治疗习惯性流产27例．江苏中医，1997：18(10)：17．
[43] 段世彪．安奠二天汤加减治疗先兆流产50例．湖北中医杂志，1987；(1)：13．
[44] 沈宗华．自拟补肾益气汤治疗先兆流产的体会．江西中医药，1987：(3)：24．
[45] 叶宝贵，陈伟民．自拟保胎汤治疗先兆流产60例．浙江中医杂志，1986：(2)：60．
[46] 刘家磊．羊肾固冲饮治疗滑胎181例．浙江中医杂志，1991；(12)：558．
[47] 张传芳，舒沪英，黄光英．补肾益气养血法治疗先兆流产53例临床观察．同济医科大学学报，1990(3)：210．
[48] 蒋俭，李光荣，藏美佳，活血化瘀法治疗血瘀型胎漏胎动不安滑胎41例临床观察．中医杂志，1992；(11)：30—32．
[49] 归绥琪．封闭抗体缺乏性自然流产者的中药治疗．上海医科大学学报，1997；24，(3)：217—219．
[50] 吴宁．中药复方优生宁对先兆流产病人NK细胞活性的影响．中医药信息2002：19(2)：59-60．
[51] 景苏玉，归绥棋．“保胎Ⅰ冲剂”提高反复自然流产患者免疫功能的临床观察 上海中医药杂志，2002：9：23-24．
[52] 索玉平．海马生髓丸对复发性流产的免疫调节作用研究．中医药学刊．2003；21(9)：1532-1533．
[53] 胡昌东，归绥琪，陆利民．中药治疗溴隐亭致大鼠流产模型中内分泌对蜕膜TNF-α、Fas的影响．中华中医药杂志，2005；20(1)：36-39．
[54] 许钧，归绥琪．自然流产病因分类与中医证型相关性研究．上海中医药杂志，2003； 37(3)：10-11．
[55] 赵颖，张玉珍．助孕3号丸治疗脾肾虚证胎漏、胎动不安、滑胎疗效观察。新中医，1999：31(12)：20—21。
[56] 韩惠，罗颂平．助孕3号方对大鼠子宫收缩活动的影响．广州中医药大学学报，2002；19(1)；33-35．
[57] 李小球，罗颂平．助孕3号丸对妊娠大鼠孕激素及受体的影响．暨南大学学报(医学版)，1999：20(6)：111-113．
[58] 罗颂平，梁国珍，张玉珍，等．助孕3号方防治大鼠自然流产的机理研究．中国中西医结合杂志，2003：23(7)：522-525．
[59] 宁艳，罗颂平，梁国珍，补肾健脾中药复方对肾虚黄体抑制妊娠大鼠流产模型的实验研究．中国医药学报，2000；15(6)：428-430．
[60] 曾诚，岳明明，试论先兆流产的中医证型分布规律．中国中医药信息杂志，2002；9(8)：5-7．