分娩启动中NF-κB与AP-1易位及NMB对其的影响

发布时间：2018-06-03 06:54

本文选题：核转录因子kappa + B　；参考：《中南大学》2014年硕士论文

【摘要】：目的： 1、探讨NMB介导的NF-κB及AP-1的表达量变化和相关性与分娩启动的关系。 2、研究子宫下段平滑肌细胞中NMB介导的NF-κB及AP-1发生细胞内易位与分娩启动的关系。方法： 1、组织块贴壁法原代培养人妊娠晚期子宫下段平滑肌细胞,采用免疫细胞化学方法进行鉴定。 2、临产前后子宫下段平滑肌组织采用石蜡包埋固定,采用苏木精-伊红染色方法进行切片质量检测。 3、采用免疫组织化学方法对自然临产前后子宫下段平滑肌细胞中NF-κB及AP-1表达进行测定。 4、采用免疫细胞荧光对NMB诱导前后子宫下段平滑肌细胞中NF-κB(p65)及AP-1(c-Jun)表达进行测定,并通过多重免疫荧光染色进行NF-κB及AP-1的细胞内共定位。 5、采用Image-Pro Plus6.0软件进行NF-κB及AP-1表达的累积光密度值(IOD)测定,采用SPSS19.0软件分析NF-κB及AP-1表达的差异及相关性。结果： 1、组织块贴壁法培养的原代子宫平滑肌细胞生长状态良好,经免疫细胞化学法α-SMA鉴定及NMBR检测,细胞纯度近100%,说明培养的细胞是表达NMBR的子宫平滑肌细胞。 2、组织经石蜡包埋固定,行HE染色检测。子宫下段平滑肌细胞结构清晰,说明组织固定包埋方法可行,组织切片适于进行免疫组织化学染色检测。 3、自然临产前后子宫下段平滑肌细胞中p65、c-Jun定位：未临产组p65与c-Jun主要位于细胞浆,临产组p65与c-Jun主要位于细胞核。并且临产组与未临产组p65、c-Jun表达量存在差异,临产组p65及c-Jun表达量高于未临产组,差异具有统计学意义,P0.05。 4、自然临产前后子宫平滑肌细胞中p65与c-Jun表达量Pearson相关性分析：未临产组及临床组的p65及c-Jun表达均存在正相关,相关系数分别为0.659、0.879,p0.05。 5、NMB诱导前后p65、c-Jun定位：未加NMB的对照组位于细胞浆；NMB4h组主要位于核膜周边区域；NMB8h组主要位于细胞核,细胞浆呈低表达。 6、NMB诱导前后的p65、c-Jun表达：NMB8h组IOD值明显高于对照组及NMB4h组,差异具有统计学意义,P0.05;NMB4h组与对照组之间差异无统计学意义,p0.05。 7、p65与c-Jun共定位：对照组、NMB4h组、NMB8h组,多重免疫荧光染色显示二者在细胞内表达部位一致,NMB诱导前二者主要位于细胞浆,NMB诱导后8小时二者主要位于细胞核,表达部位几乎完全重叠。 8、NMB诱导的p65与c-Jun表达量Pearson相关性分析：对照组、NMB4h组、NMB8h组中,p65与c-Jun均呈正相关,相关系数分别为0.587、0.653、0.874,P0.05。结论： 1.分娩启动前,NF-κB及AP-1定位于细胞浆,分娩启动后NF-κB及AP-1迁移进入细胞核。 2.NMB可同时诱导NF-κB及AP-1易位,二者定位及迁移方向呈同步变化,表达量同时增高,推测NF-κB与AP-1可能存在协同作用。
[Abstract]:Objective: 1. To investigate the relationship between the expression of NF- 魏 B and AP-1 mediated by NMB and the onset of labor. 2. To study the relationship between the intracellular translocation of NF- 魏 B and AP-1 mediated by NMB and the initiation of labor in the lower uterine smooth muscle cells. Methods: 1. The primary culture of human lower uterine smooth muscle cells in late pregnancy was identified by immunocytochemistry. 2. The smooth muscle tissue of the lower segment of uterus was fixed with paraffin and stained with hematoxylin-eosin before and after labor. 3. The expression of NF- 魏 B and AP-1 in the smooth muscle cells of the lower segment of uterus before and after spontaneous labor was determined by immunohistochemical method. (4) the expression of NF- 魏 B p65) and AP-1Tc-Jun) in the smooth muscle cells of the lower uterine segment before and after NMB induction were detected by immunofluorescence, and the intracellular localization of NF- 魏 B and AP-1 were detected by multiple immunofluorescence staining. 5. The cumulative optical density (IOD) of NF- 魏 B and AP-1 expression was determined by Image-Pro Plus6.0 software, and the difference and correlation of NF- 魏 B and AP-1 expression were analyzed by SPSS19.0 software. Results: 1. The primary uterine smooth muscle cells cultured by tissue mass adherence method grew well. The purity of the cells was nearly 100 by immunocytochemistry and NMBR detection, indicating that the cultured cells were uterine smooth muscle cells expressing NMBR. 2. The tissues were fixed with paraffin and stained with HE. The clear structure of smooth muscle cells in the lower segment of uterus showed that the method of tissue fixation and embedding was feasible and the tissue sections were suitable for immunohistochemical staining. 3. The localization of p65 c-Jun in the smooth muscle cells of the lower segment of uterus before and after spontaneous labor: p65 and c-Jun were mainly located in the cytoplasm in the non-parturient group, and p65 and c-Jun in the nucleus in the parturient group. The expression of p65 c-Jun in parturient group was higher than that in non-parturient group, and the difference was statistically significant (P 0.05). 4. Pearson correlation analysis of p65 and c-Jun expression in uterine smooth muscle cells before and after spontaneous labor: there was a positive correlation between p65 and c-Jun expression in unpartum group and clinical group, the correlation coefficient was 0.659 ~ 0.879p 0.05, respectively. Before and after induction of NMB, the position of c-Jun in the control group without NMB was mainly located in the nucleus in the peripheral region of the nuclear membrane, and the cytoplasm of the control group was low in the cytoplasm of NMB4 h group. The expression of p65 c-Jun was significantly higher in the 8h group than that in the control group and NMB4h group before and after induction, and there was no significant difference between the control group and the control group (P 0.05). 7p65 and c-Jun co-localization: in the control group of 4 h after NMB induction, multiple immunofluorescence staining showed that both of them were mainly located in the nucleus 8 hours after induction of NMB in cytoplasm. The site of expression was almost completely overlapped. Analysis of Pearson correlation between p65 and c-Jun expression induced by NMB: there was a positive correlation between p65 and c-Jun in control group (n = 4), and the correlation coefficient was 0.5870.653U 0.874P 0.05, respectively. Conclusion: 1. NF- 魏 B and AP-1 were located in cytoplasm before parturition, and NF- 魏 B and AP-1 migrated into nucleus after parturition. 2.NMB could induce the translocation of NF- 魏 B and AP-1 at the same time. The localization and migration direction of NF- 魏 B and AP-1 were changed synchronously, and the expression of NF- 魏 B was increased simultaneously. It is speculated that NF- 魏 B and AP-1 might have synergistic effect.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714

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