复发性流产的病因分析及其与趋化因子CXCL12、CCL2、RANTES表达异常的研究
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摘要
复发性流产(Recurrent Spontaeous Abortion, RSA)是指连续发生2次或2次以上的自然流产,发病率约占育龄夫妇的5%。RSA的病因复杂,排除临床上可查出的病因,仍有近1/2的RSA患者的病因未明,称为不明原因复发性流产(Unexplained recurrent spontaneous abortion URSA),是临床上难治性的不育症。
第一部分反复性自然流产病因分析
目的：探讨RSA患者的临床病因。
方法：对74对RSA夫妇进行规范化病因筛查,包括夫妇染色体核型分析、生殖道解剖结构检查、内分泌激素检查、生殖道感染因素检查以及自身免疫抗体检测,以分析RSA的病因、建立RSA病因筛查的临床路径。
结果：在74对RSA夫妇中,染色体异常6例,占8.1%；生殖道解剖异常4例,占5.4%；内分泌异常13例,占17.6%；生殖道感染5例,占6.8%；免疫异常46例,占62.1%,其中自身免疫异常14例,占18.9%；同种免疫异常(不明原因)32例,占43.2%。
结论：RSA病因复杂,包括染色体异常,生殖道解剖结构异常,内分泌代谢异常,生殖道感染,免疫异常等因素。免疫因素在RSA的病因中占重要地位,免疫异常中约有2/3原因不明,临床上称为URSA。病因学的探讨以及针对病因进行预防性治疗是防治RSA的关键。
第二部分趋化因子CXCL12、CCL2、RANTES表达异常与URSA发病的关系
目的：探讨URSA患者绒毛、蜕膜基质细胞衍生因子(CXCL12)、单核细胞趋化蛋白-1(CCL2)、调节活化正常T细胞表达和分泌的因子(RANTES)-mRNA表达异常和血清CXCL12、CCL2、RANTES水平变化与URSA发病的关系。
方法：收集URSA患者26例(URSA组)和正常早孕妇女30例(对照组)的绒毛、蜕膜和血清标本,应用实时荧光PCR方法检测两组绒毛、蜕膜CXCL12、CCL2、RANTES-mRNA的表达,应用ELISA方法检测两组血清CXCL12、CCL2、RANTES的蛋白含量。
结果：1)人早孕绒毛、蜕膜组织均有趋化因子CXCL12、CCL2、RANTES-mRNA表达。2)URSA组绒毛、蜕膜组织CXCL12-mRNA的表达量为(0.46±0.15,0.35±0.13),均较对照组(1.79±0.82,0.81±0.21)显著降低(P<0.01)：CCL2-mRNA的表达量为(1.99±0.35,2.77±0.61),均较对照组(1.42±0.28,1.90±0.85)显著升高(P<0.01)；RANTES-mRNA的表达量为(1.40±0.25,2.25±0.33),均较对照组(0.94±0.22,1.45±0.20)显著升高(P<0.01)。3)URSA组血清CXCL12的蛋白含量为(94.52±29.23pg/ml),对照组为(268.13+26.36pg/ml),两组相比,差异有显著性(P<0.01)；CCL2的蛋白含量为(83.34±10.61pg/ml),对照组为(59.41±9.70pg/ml),两组相比,差异有显著性(P<0.01)；RANTES的蛋白含量为(117.29±18.67pg/ml),对照组为(79.88±12.54pg/ml),两组相比,差异有显著性(P<0.01)。4)两组血清CXCL12值与绒毛、蜕膜CXCL12-mRNA表达量呈正相关,相关系数分别为0.723,0.604(P均<0.01)；CCL2值与绒毛、蜕膜CCL2-mRNA表达量呈正相关,相关系数分别为0.595,0.655(P均<0.01)；RANTES值与绒毛、蜕膜RANTES-mRNA表达量呈正相关,相关系数分别为0.482,0.675(P均<0.01)。
结论：CXCL12、CCL2、RANTES表达于人早孕绒毛和蜕膜组织中,参与母胎界面绒毛滋养细胞和蜕膜免疫活性细胞功能的调控；绒毛、蜕膜组织CXCL12低表达,CCL2、RANTES高表达与URSA的发病机制有关；血清CXCL12、CCL2、RANTES在胚胎停育的早期诊断中具有重要的临床意义。
第三部分：米非司酮对早孕绒毛、蜕膜趋化因子CXCL12、CCL2、RANTES表达的影响
目的：探讨孕激素受体拮抗剂米非司酮对早孕绒毛、蜕膜组织趋化因子CXCL12、CCL2、RANTES表达的影响。
方法：收集服用米非司酮48小时后清宫的正常早孕妇女30例(米非司酮组)的绒毛、蜕膜标本,应用实时荧光PCR方法检测米非司酮组绒毛、蜕膜CXCL12、CCL2、RANTES-mRNA的表达,并与对照组比较。
结果：米非司酮组绒毛、蜕膜组织CXCL12-mRNA的表达量为(0.59±0.22,0.42±0.17),均较对照组显著降低(P<0.01)；CCL2-mRNA的表达量值为(1.84±0.35,3.01±0.62),均较对照组显著升高(P<0.01)；RANTES-mRNA的表达量为(1.33±0.22,2.17±0.49),均较对照组显著升高(P<0.01)。
结论：米非司酮通过影响早孕绒毛、蜕膜趋化因子CXCL12、CCL2、RANTES-mRNA的表达,破坏母胎界面免疫耐受机制而发挥其抗早孕作用。
Recurrent spontaneous miscarriage(RSA) is loss of two or more consecutive pregnancies. It is experienced by 5% of couples. The etiology of RSA is complicated, 50% of cases of RSA is unknown, which is called URSA.
Part 1 The etiological factors of RSA.
Objective To analyze etiological factors of RSA.
Methods 74 RSA couples were selected and scanned systematically for analyzing etiological factors.
Results 74 RSA couples,6 cases with abnormal chromosomes(8.1%),4 cases with uterine malformation(5.4%); 13 cases with abnormal hormones(17.6%); 5 cases with reproductive infection(6.8%); 46 cases with immunological factors(62.1%), including 14 cases with positive self-antibodies(18.9%) and 32 cases with unknown aetiology(43.2%).
Conclusion The etiology of RSA is complicated, including genetic factors, endocrine factors, uterine malformation, infective and immunological factors. RSA is mainly related to immunological factors.
Part 2:Roles of CXCL12, CCL2, RANTES in pathogenesis of URSA
Objective To investigate the abnormal expression of CXCL12、CCL2、RANTES mRNA in villus and decidua and serum levels of CXCL12, CCL2, RANTES in URSA patients in order to find out its association with the pathogenesis of URSA.
Methods The expression of CXCL12, CCL2, RANTESmRNA in villus and decidua tissues of 26 URSA patients(URSA group)and 30 normal early pregnancy women(control group) were determined by real-time fluorescence quantitative PCR. the maternal serum levels of CXCL12, CCL2, RANTES were detected by ELISA.
Results 1) CXCL12、CCL2、RANTESmRNA expression were detected in human first-trimester placental villus and decidua tissues; 2) The expression of CXCL12mRNA in villus and decidua in URSA group were (0.46±0.15,0.35±0.13), which was lower than that in control group(1.79±0.82,0.81±0.21)(P<0.01); CCL2mRNA were (1.99±0.35,2.77±0.61), which was higher than that in control group(1.42±0.28,1.90±0.85) (P<0.01); RANTESmRNA were (1.40±0.25,2.25±0.33), which was higher than that in control group(0.94±0.22,1.45±0.20)(P<0.01).3) The level of serum CXCL12 in URSA group was (194.52±29.23pg/ml), which was lower than control group(268.13±26.36pg/ml)(P<0.01); CCL2 was (83.34±10.61pg/ml), which was higher than control group (59.41±9.70pg/ml)(P<0.01); RANTES was (117.29±18.67pg/ml), which was higher than that in control group(79.88±12.54pg/ml) (P<0.01).4)The serum CXCL12 levels were positively correlated with CXCL12mRNA levels in villus and decidua (r=0.723,0.604 respectively)(P<0.01); CCL2 levels were positively correlated with CCL2 mRNA levels (r=0.595,0.655 respectively)(P<0.01); RANTES levels were positively correlated with RANTES mRNA levels in both groups(r=0.482,0.675 respectively) (P<0.01).
Conclusion CXCL12、CCL2、RANTES were co-expressed in human villus and decidua, they contribute to the trophoblast invasion and deciduous cell constitution at matemo-fetal interface; the lower CXCL12 expression and higher CCL2, RANTES expression play important roles in the pathogenesis of URSA; the levels of serum CXCL12, CCL2, RANTES can be used as a tool for diagnosing embryonic death.
Part 3:Effect of mifepristone on the expression of chemokines CXCL12, CCL2, RANTES in villus and decidua of early pregnancy
Objective To investigate mifepristone on the expression of CXCL12、CCL2、RANTES in villus and decidua of early pregnancy.
Methods The expression of CXCL12, CCL2, RANTESmRNA in villous and deciduous tissues of 30 normal early pregnancy women who volunteered to take mifepristone (mifepristone group) were determined by real-time PCR.
Results The expression of CXCL12 in mifepristone group were (0.59±0.22, 0.42±0.17), which was lower than control group(P<0.01); CCL2 were(1.84±0.35, 3.01±0.62), which was higher than control group(P<0.01); RANTES were(1.33±0.22; 2.17±0.49), which was higher than that in control group(P<0.01).
Conclusion Mifepristone affect the expression of chemokines and induce the disorder of micro-enviromnenl, which might be the mechanism of medical abortion.
第一部分反复性自然流产病因分析
目的：探讨RSA患者的临床病因。
方法：对74对RSA夫妇进行规范化病因筛查,包括夫妇染色体核型分析、生殖道解剖结构检查、内分泌激素检查、生殖道感染因素检查以及自身免疫抗体检测,以分析RSA的病因、建立RSA病因筛查的临床路径。
结果：在74对RSA夫妇中,染色体异常6例,占8.1%；生殖道解剖异常4例,占5.4%；内分泌异常13例,占17.6%；生殖道感染5例,占6.8%；免疫异常46例,占62.1%,其中自身免疫异常14例,占18.9%；同种免疫异常(不明原因)32例,占43.2%。
结论：RSA病因复杂,包括染色体异常,生殖道解剖结构异常,内分泌代谢异常,生殖道感染,免疫异常等因素。免疫因素在RSA的病因中占重要地位,免疫异常中约有2/3原因不明,临床上称为URSA。病因学的探讨以及针对病因进行预防性治疗是防治RSA的关键。
第二部分趋化因子CXCL12、CCL2、RANTES表达异常与URSA发病的关系
目的：探讨URSA患者绒毛、蜕膜基质细胞衍生因子(CXCL12)、单核细胞趋化蛋白-1(CCL2)、调节活化正常T细胞表达和分泌的因子(RANTES)-mRNA表达异常和血清CXCL12、CCL2、RANTES水平变化与URSA发病的关系。
方法：收集URSA患者26例(URSA组)和正常早孕妇女30例(对照组)的绒毛、蜕膜和血清标本,应用实时荧光PCR方法检测两组绒毛、蜕膜CXCL12、CCL2、RANTES-mRNA的表达,应用ELISA方法检测两组血清CXCL12、CCL2、RANTES的蛋白含量。
结果：1)人早孕绒毛、蜕膜组织均有趋化因子CXCL12、CCL2、RANTES-mRNA表达。2)URSA组绒毛、蜕膜组织CXCL12-mRNA的表达量为(0.46±0.15,0.35±0.13),均较对照组(1.79±0.82,0.81±0.21)显著降低(P<0.01)：CCL2-mRNA的表达量为(1.99±0.35,2.77±0.61),均较对照组(1.42±0.28,1.90±0.85)显著升高(P<0.01)；RANTES-mRNA的表达量为(1.40±0.25,2.25±0.33),均较对照组(0.94±0.22,1.45±0.20)显著升高(P<0.01)。3)URSA组血清CXCL12的蛋白含量为(94.52±29.23pg/ml),对照组为(268.13+26.36pg/ml),两组相比,差异有显著性(P<0.01)；CCL2的蛋白含量为(83.34±10.61pg/ml),对照组为(59.41±9.70pg/ml),两组相比,差异有显著性(P<0.01)；RANTES的蛋白含量为(117.29±18.67pg/ml),对照组为(79.88±12.54pg/ml),两组相比,差异有显著性(P<0.01)。4)两组血清CXCL12值与绒毛、蜕膜CXCL12-mRNA表达量呈正相关,相关系数分别为0.723,0.604(P均<0.01)；CCL2值与绒毛、蜕膜CCL2-mRNA表达量呈正相关,相关系数分别为0.595,0.655(P均<0.01)；RANTES值与绒毛、蜕膜RANTES-mRNA表达量呈正相关,相关系数分别为0.482,0.675(P均<0.01)。
结论：CXCL12、CCL2、RANTES表达于人早孕绒毛和蜕膜组织中,参与母胎界面绒毛滋养细胞和蜕膜免疫活性细胞功能的调控；绒毛、蜕膜组织CXCL12低表达,CCL2、RANTES高表达与URSA的发病机制有关；血清CXCL12、CCL2、RANTES在胚胎停育的早期诊断中具有重要的临床意义。
第三部分：米非司酮对早孕绒毛、蜕膜趋化因子CXCL12、CCL2、RANTES表达的影响
目的：探讨孕激素受体拮抗剂米非司酮对早孕绒毛、蜕膜组织趋化因子CXCL12、CCL2、RANTES表达的影响。
方法：收集服用米非司酮48小时后清宫的正常早孕妇女30例(米非司酮组)的绒毛、蜕膜标本,应用实时荧光PCR方法检测米非司酮组绒毛、蜕膜CXCL12、CCL2、RANTES-mRNA的表达,并与对照组比较。
结果：米非司酮组绒毛、蜕膜组织CXCL12-mRNA的表达量为(0.59±0.22,0.42±0.17),均较对照组显著降低(P<0.01)；CCL2-mRNA的表达量值为(1.84±0.35,3.01±0.62),均较对照组显著升高(P<0.01)；RANTES-mRNA的表达量为(1.33±0.22,2.17±0.49),均较对照组显著升高(P<0.01)。
结论：米非司酮通过影响早孕绒毛、蜕膜趋化因子CXCL12、CCL2、RANTES-mRNA的表达,破坏母胎界面免疫耐受机制而发挥其抗早孕作用。
Recurrent spontaneous miscarriage(RSA) is loss of two or more consecutive pregnancies. It is experienced by 5% of couples. The etiology of RSA is complicated, 50% of cases of RSA is unknown, which is called URSA.
Part 1 The etiological factors of RSA.
Objective To analyze etiological factors of RSA.
Methods 74 RSA couples were selected and scanned systematically for analyzing etiological factors.
Results 74 RSA couples,6 cases with abnormal chromosomes(8.1%),4 cases with uterine malformation(5.4%); 13 cases with abnormal hormones(17.6%); 5 cases with reproductive infection(6.8%); 46 cases with immunological factors(62.1%), including 14 cases with positive self-antibodies(18.9%) and 32 cases with unknown aetiology(43.2%).
Conclusion The etiology of RSA is complicated, including genetic factors, endocrine factors, uterine malformation, infective and immunological factors. RSA is mainly related to immunological factors.
Part 2:Roles of CXCL12, CCL2, RANTES in pathogenesis of URSA
Objective To investigate the abnormal expression of CXCL12、CCL2、RANTES mRNA in villus and decidua and serum levels of CXCL12, CCL2, RANTES in URSA patients in order to find out its association with the pathogenesis of URSA.
Methods The expression of CXCL12, CCL2, RANTESmRNA in villus and decidua tissues of 26 URSA patients(URSA group)and 30 normal early pregnancy women(control group) were determined by real-time fluorescence quantitative PCR. the maternal serum levels of CXCL12, CCL2, RANTES were detected by ELISA.
Results 1) CXCL12、CCL2、RANTESmRNA expression were detected in human first-trimester placental villus and decidua tissues; 2) The expression of CXCL12mRNA in villus and decidua in URSA group were (0.46±0.15,0.35±0.13), which was lower than that in control group(1.79±0.82,0.81±0.21)(P<0.01); CCL2mRNA were (1.99±0.35,2.77±0.61), which was higher than that in control group(1.42±0.28,1.90±0.85) (P<0.01); RANTESmRNA were (1.40±0.25,2.25±0.33), which was higher than that in control group(0.94±0.22,1.45±0.20)(P<0.01).3) The level of serum CXCL12 in URSA group was (194.52±29.23pg/ml), which was lower than control group(268.13±26.36pg/ml)(P<0.01); CCL2 was (83.34±10.61pg/ml), which was higher than control group (59.41±9.70pg/ml)(P<0.01); RANTES was (117.29±18.67pg/ml), which was higher than that in control group(79.88±12.54pg/ml) (P<0.01).4)The serum CXCL12 levels were positively correlated with CXCL12mRNA levels in villus and decidua (r=0.723,0.604 respectively)(P<0.01); CCL2 levels were positively correlated with CCL2 mRNA levels (r=0.595,0.655 respectively)(P<0.01); RANTES levels were positively correlated with RANTES mRNA levels in both groups(r=0.482,0.675 respectively) (P<0.01).
Conclusion CXCL12、CCL2、RANTES were co-expressed in human villus and decidua, they contribute to the trophoblast invasion and deciduous cell constitution at matemo-fetal interface; the lower CXCL12 expression and higher CCL2, RANTES expression play important roles in the pathogenesis of URSA; the levels of serum CXCL12, CCL2, RANTES can be used as a tool for diagnosing embryonic death.
Part 3:Effect of mifepristone on the expression of chemokines CXCL12, CCL2, RANTES in villus and decidua of early pregnancy
Objective To investigate mifepristone on the expression of CXCL12、CCL2、RANTES in villus and decidua of early pregnancy.
Methods The expression of CXCL12, CCL2, RANTESmRNA in villous and deciduous tissues of 30 normal early pregnancy women who volunteered to take mifepristone (mifepristone group) were determined by real-time PCR.
Results The expression of CXCL12 in mifepristone group were (0.59±0.22, 0.42±0.17), which was lower than control group(P<0.01); CCL2 were(1.84±0.35, 3.01±0.62), which was higher than control group(P<0.01); RANTES were(1.33±0.22; 2.17±0.49), which was higher than that in control group(P<0.01).
Conclusion Mifepristone affect the expression of chemokines and induce the disorder of micro-enviromnenl, which might be the mechanism of medical abortion.
引文
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