SOCS3在子痫前期胎盘组织中的表达及其对滋养细胞增殖和迁移能力的影响

发布时间：2018-05-06 10:07

本文选题：子痫前期 + 细胞因子信号转导蛋白抑制因子　；参考：《南京医科大学》2014年硕士论文

【摘要】：目的：探究细胞因子信号转导抑制因子3(suppressor of cytokine signaling3, SOCS3)基因在人子痫前期胎盘组织中的表达情况及其对HTR-8/SVneo细胞增殖与迁移能力的影响。方法： 1.选取2011年10月至2012年10月在南京医科大学第一附属医院住院分娩的15例重度子痫前期孕妇为子痫前期组,15例正常孕妇作为正常妊娠组。 2.采用实时荧光定量逆转录-聚合酶链反应和Western印迹技术检测两组孕妇胎盘组织中SOCS3mRNA和蛋白表达水平。 3.体外培养的HTR-8/SVneo细胞分别转染SOCS3小分子干扰RNA(实验组)和无意义的阴性对照小分子干扰RNA(阴性对照组)。4.采用实时荧光定量逆转录-聚合酶链反应和Western印迹技术检测体外培养的HTR-8/SVneo细胞中SOCS3mRNA和蛋白表达水平。 5.采用克隆形成实验和3-(4,5)-二甲基噻唑-(2,5)-二苯基溴化四氮唑蓝法(MTT)检测转染后两个实验组中细胞的增殖能力。 6.应用流式细胞仪对转染后两个实验组中细胞进行细胞周期分析。 7.采用transwell小室实验检测转染后两个实验组中细胞迁移能力。结果： 1.子痫前期组的胎盘组织中SOCS3mRNA和SOCS3蛋白水平显著低于正常组胎盘组织,分别为0.254±0.03和0.21±0.05,均低于正常妊娠组0.71±0.08和0.75±0.12。差异均有统计学意义(p0.05)。 2.小分子干扰RNA转染24h后,实验组SOCS3mRNA水平低于阴性对照组0.39±0.02与1.0±0.04,蛋白水平也较低(0.0037+0.0014与1.5149±0.0357)。实验组中SOCS3mRNA和SOCS3蛋白水平表达均降低,差异均有统计学意义(P0.05),证明干扰SOCS3表达成功。 3.MTT法检测证实实验组细胞增殖能力降低,转染后48、72和96h实验组增殖能力分别为0.23±0.01,0.32±0.02和0.3±0.02,均低于阴性对照组(分别为0.39±0.02,0.55±0.04和0.86±0.04),差异有统计学意义(P0.05)。 4.克隆形成实验证实转染10d时实验组克隆形成细胞个数显著低于阴性对照组,实验组细胞数为116±15,低于阴性对照组312±24,差异有统计学意义(P0.05)。 5.应用流式细胞仪分析细胞周期发现转染48h后,实验组G1/Go期细胞比例为(55.75±2.21)%,高于阴性对照组[(47.88±1.87)%](t＝45.43,P0.05)；S期细胞比例为(31.59±0.83)%,低于阴性对照组[(37.38±1.34)%](t=20.06,P0.05)。两组分别比较,差异均有统计学意义(P0.05)。 6.transwell小室实验证实转染48h后,实验组穿膜细胞数为(93±11)个,低于阴性对照组(167±17)个,两组相比较,差异有统计学意义(P0.05)。结论： 1.子痫前期患者胎盘组织中SOCS3水平与正常妊娠孕妇相比较表达下降,推测SOCS3可能参与了子痫前期的发生发展过程。 2. SOCS3可能通过降低表达水平来抑制滋养细胞的增殖和迁移能力,从而在子痫前期的发病过程中起作用。
[Abstract]:Objective: To investigate the expression of cytokine signal transduction inhibitor 3(suppressor of cytokine signaling3 (SOCS3) gene in human preeclampsia placenta and its effect on the proliferation and migration of HTR-8/SVneo cells. Methods: 1. From October 2011 to October 2012, 15 pregnant women with severe preeclampsia were selected as normal pregnancy group and 15 normal pregnant women in the first affiliated Hospital of Nanjing Medical University. 2. The expression of SOCS3mRNA and protein in placenta of pregnant women was detected by real-time fluorescence quantitative reverse transcription-polymerase chain reaction and Western blotting. 3. HTR-8/SVneo cells were transfected with SOCS3 small molecule interference RNAs (experimental group) and meaningless negative control group (negative control group) respectively. The expression of SOCS3mRNA and protein in cultured HTR-8/SVneo cells was detected by real-time fluorescence quantitative reverse transcription-polymerase chain reaction and Western blotting. 5. Clone formation assay and MTT assay were used to detect the proliferation of the cells in the two experimental groups after transfection. 6. Flow cytometry was used to analyze the cell cycle in the two experimental groups after transfection. 7. The ability of cell migration in the two experimental groups after transfection was detected by transwell chamber experiment. Results: 1. The levels of SOCS3mRNA and SOCS3 protein in placenta of preeclampsia group were significantly lower than those of normal group (0.254 卤0.03 and 0.21 卤0.05, respectively), and were lower than those of normal pregnancy group (0.71 卤0.08) and 0.75 卤0.12 (P < 0.05). The difference was statistically significant (P 0.05). 2. 24 hours after transfection with small interfering RNA, the level of SOCS3mRNA in the experimental group was lower than that in the negative control group (0.39 卤0.02 vs 1.0 卤0.04), and the protein level was also lower than that in the negative control group (0.0037 0.0014 and 1.5149 卤0.0357). The expression of SOCS3mRNA and SOCS3 protein decreased in the experimental group, and the difference was statistically significant (P 0.05), which proved the interference of SOCS3 expression was successful. The proliferative ability of the experimental group was 0.23 卤0.01 卤0.032 卤0.02 and 0.3 卤0.02, respectively, which was significantly lower than that of the negative control group (0.39 卤0.02 卤0.55 卤0.04 and 0.86 卤0.04) at 48 ~ 72 and 96 h after transfection, respectively (P 0.05). 4. The number of Clone forming cells in the experimental group was significantly lower than that in the negative control group at 10 days after transfection, and the number of the cells in the experimental group was 116 卤15, which was lower than that in the negative control group (312 卤24). The difference was statistically significant (P 0.05). 5. Flow cytometry analysis showed that the percentage of G1/Go phase cells in the experimental group was 55.75 卤2.21 after 48 h transfection, which was higher than that in the negative control group [47.88 卤1.87%] TX 45.43 P 0.05%, and was lower than that in the negative control group (37.38 卤1.34%), and the ratio of the cells in the S phase was 31.59 卤0.83%, which was lower than that in the negative control group (37.38 卤1.34%). The difference between the two groups was statistically significant (P 0.05). 6.transwell chamber experiment confirmed that the number of perforated cells in the experimental group was 93 卤11, which was lower than that in the negative control group (167 卤17) after 48 hours of transfection. The difference between the two groups was statistically significant (P 0.05). Conclusion: 1. The expression of SOCS3 in placenta of preeclampsia patients was lower than that of normal pregnant women. It was speculated that SOCS3 might be involved in the occurrence and development of preeclampsia. 2. SOCS3 may play a role in the pathogenesis of preeclampsia by inhibiting the proliferation and migration of trophoblast.
【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714.2

【共引文献】