基于NF-κB通路探讨补肾中药在胚胎植入母胎界面免疫耐受中的调节作用

发布时间：2018-05-11 11:50

本文选题：NF-κB信号通路 + 肾气虚证　；参考：《山东中医药大学》2016年硕士论文

【摘要】：目的:研究二至天癸颗粒对肾气虚型反复种植失败患者临床妊娠率及外周血NF-κB信号转导通路中关键因子NF-κB、MCP-1的干预调节作用,探讨肾气虚致反复种植失败在NF-κB信号转导通路水平可能的发生机制以及补肾中药在胚胎植入母胎界面免疫耐受中的调节作用。方法:本研究将符合诊断及辨证标准行体外受精-胚胎移植(IVF-ET)治疗的肾气虚反复种植失败患者70例,按照门诊就诊时间先后顺序编号,奇数号分入治疗组,偶数号分入安慰剂组,另选取因男方因素行IVF的健康妇女35例(非肾气虚组)。治疗组在服用二至天癸颗粒3个周期后联合常规IVF治疗,安慰剂组及非肾气虚组服用安慰剂3个周期并联合常规IVF治疗,观察肾气虚反复种植失败患者的肾虚证候积分变化;受精率;优质胚胎率;生化妊娠率;临床妊娠率。采用酶联免疫吸附方法(ELISA)检测三组抗绒毛膜促性腺激素抗体(Ah CGAb)、抗心磷脂抗体(ACA-Ig G)、抗精子抗体(As Ab)、抗子宫内膜抗体(AEMAb)、抗卵巢抗体(AOVAb)水平,采用实时荧光定量PCR(RT-PCR)技术、蛋白质印记(Western-blot)法分别检测移植日外周血中NF-κB、MCP-1m RNA及NF-κB、MCP-1蛋白的含量;采用ELISA法检测三组患者移植后7天血清MCP-1浓度。结果:1.二至天癸颗粒治疗组患者经治疗后肾气虚症状明显改善,证候积分差异显著(P0.01);两组治疗前后肾气虚证候积分差值有显著统计学差异(P0.01)。2.与安慰剂组相比,治疗组及非肾气虚组受精率、优质胚胎率、生化妊娠率、临床妊娠率均较高,有统计学差异(P0.05);治疗组与非肾气虚组之间无统计学差异(P0.05)。3.与非肾气虚组相比,安慰剂组及治疗组Ah CGAb、ACA-Ig G、As Ab、AEMAb、AOVAb阳性率均较高,有统计学差异(P0.05);安慰剂组与治疗组之间无统计学差异(P0.05)。4.分析三组患者外周血NF-κB信号转导通路中关键效应分子NF-κB及其下游因子MCP-1的表达,发现:与安慰剂组相比,非肾气虚组及治疗组移植日NF-κB、MCP-1蛋白含量均下降,NF-κBm RNA、MCP-1m RNA表达均下调,移植后7天血清MCP-1含量降低,有统计学差异(P0.05);治疗组与非肾气虚组相比,NF-κB、MCP-1蛋白含量、NF-κBm RNA、MCP-1m RNA表达水平、血清MCP-1含量无统计学差异(P0.05)。5.进一步将患者分为妊娠组与未妊娠组,两组相比,发现:妊娠组Ah CGAb、ACA-Ig G、As Ab、AEMAb、AOVAb阳性率及血清MCP-1浓度均低于未妊娠组,有显著统计学差异(P0.01)。结论:1.二至天癸颗粒可明显改善肾气虚型反复种植失败患者的肾虚状态,提高其受精率及优质胚胎率,改善临床妊娠率。2.生殖抗体可能是影响妊娠环境的重要免疫因素。3.NF-κB、MCP-1m RNA及蛋白的高表达可能与反复种植失败的发病机制有关,血清MCP-1对于预测早期胚胎移植状况可能具有重要的临床意义。4.二至天癸颗粒可能是通过抑制NF-κB信号转导通路,下调MCP-1因子表达,降低母胎界面免疫排斥作用,从而提高种植率,改善肾气虚反复种植失败患者的妊娠结局。
[Abstract]:Objective: to study the effect of Erzhi Tiangui granule on the clinical pregnancy rate and the intervention and regulation of the key factor NF- 魏 B MCP-1 in peripheral blood NF- 魏 B signal transduction pathway in patients with repeated implantation failure of kidney-qi deficiency type. To explore the possible mechanism of repeated implantation failure induced by deficiency of kidney qi at the level of NF- 魏 B signal transduction pathway and the regulatory role of traditional Chinese medicine for tonifying the kidney in the immune tolerance of embryo implantation interface. Methods: in this study, 70 patients with kidney qi deficiency were divided into treatment group according to the order of outpatient visit time and odd number, which met the criteria of diagnosis and syndrome differentiation and were treated with IVF-ETS in vitro fertilization and embryo transfer. Even numbers were divided into placebo group and 35 healthy women who were treated with IVF because of male factors (non-kidney qi deficiency group). The patients in the treatment group were treated with routine IVF after taking Erzhi Tiangui granule for 3 cycles, the placebo group and non-kidney-qi deficiency group were treated with placebo for 3 cycles and combined with routine IVF, and the changes of syndrome score of kidney deficiency in the patients with failed implantation of kidney qi deficiency were observed. Fertilization rate; High quality embryo rate; biochemical pregnancy rate; Clinical pregnancy rate. Enzyme linked immunosorbent assay (Elisa) was used to detect the levels of anti-chorionic gonadotropin antibody (ACCH), anti-chorionic gonadotropin antibody (AHG), anti-cardiolipid antibody (ACA -Ig GG), anti-sperm antibody (as Abn), anti-endometrium antibody (AEMAbn), anti-ovarian antibody (AOVAb), and real-time fluorescence quantitative PCRRT-PCR (RT-PCR) technique. Protein imprinting Western-blot method was used to detect the contents of NF- 魏 BmP-1m RNA and NF- 魏 BmP-MCP-1 in peripheral blood of the patients on the day of transplantation, and the serum MCP-1 concentration of the three groups was detected by ELISA method on the 7th day after transplantation. The result is 1: 1. After treatment, the symptoms of deficiency of kidney qi were obviously improved in Erzhi Tiangui granule group, and the difference of syndrome score was significant (P 0.01). Compared with placebo group, the fertilization rate, high quality embryo rate, biochemical pregnancy rate and clinical pregnancy rate were higher in treatment group and non-kidney-qi deficiency group, and there was no statistical difference between treatment group and non-kidney-qi deficiency group. Compared with non-kidney-qi deficiency group, the positive rate of Ah CGAbACA-Ig GG as AbAEMAbAb-AOVAb in placebo group and treatment group was higher than that in non-kidney-qi deficiency group (P 0.05), but there was no significant difference between placebo group and treatment group (P 0.05. 4). The expression of NF- 魏 B and its downstream factor MCP-1 in peripheral blood NF- 魏 B signal transduction pathway was analyzed in three groups. It was found that the expression of NF- 魏 B mRNA MCP-1 protein in non-kidney-qi deficiency group and treatment group was decreased and the expression of NF- 魏 Bm RNA-MCP-1m RNA was down-regulated on the day of transplantation. After 7 days of transplantation, the content of serum MCP-1 decreased with statistical difference (P0.05), and the protein content of NF- 魏 BmRNA-MCP-1 in treatment group was higher than that in non-kidney-qi deficiency group (P < 0.05), and the expression level of NF- 魏 Bm RNA-MCP-1 RNA was not significantly different in treatment group compared with that in non-kidney-qi deficiency group (P < 0.05). The patients were further divided into gestational group and non-pregnant group. It was found that the positive rate of Ah CGAbACA-Ig AEM AbAbAAOVAb and serum MCP-1 concentration in pregnancy group were significantly lower than those in non-pregnant group (P 0.01). Conclusion 1. Erzhi Tiangui granule can obviously improve the kidney deficiency of the patients with kidney qi deficiency, improve the fertilization rate and the high quality embryo rate, and improve the clinical pregnancy rate. 2. The high expression of MCP-1m RNA and protein may be related to the pathogenesis of repeated implantation failure, and serum MCP-1 may have important clinical significance in predicting the status of early embryo transfer. Erzhitiangui granule may be by inhibiting NF- 魏 B signal transduction pathway, down-regulating the expression of MCP-1 factor, reducing the maternal and fetal interface immune rejection, so as to improve the implantation rate and improve the pregnancy outcome of patients with repeated failed implantation of kidney qi deficiency.
【学位授予单位】：山东中医药大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R271.9

【参考文献】