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RNA干扰对雄激素合成关键酶17α-羟化酶/17,20裂解酶作用
  • ISSN号:1674-1870
  • 期刊名称:《国际妇产科学杂志》
  • 分类:R512.62[医药卫生—临床医学;医药卫生—内科学] R339.21[医药卫生—人体生理学;医药卫生—基础医学]
  • 作者机构:[1]中山大学附属第一医院生殖中心,广州510080
  • 相关基金:国家自然科学基金(编号:30571956,30300372);广东省自然基金(编号:31690,21874);GZST(编号:2004J-C0111).
中文摘要:

目的:探讨细胞色素P45017α-羟化酶/17,20裂解酶(CYP17)小干扰RNA(siRNA)对CYP17基因表达和多囊卵巢综合征(PCOS)卵泡膜细胞雄激素合成的抑制效果。方法:合成4条靶向CYP17基因的特异性siRNA和非特异性siRNA。将0.4μg携带报告基因增强型绿色荧光蛋白和CYP17基因的质粒和siRNA共转染入HeIJa细胞,荧光显微镜观察及流式细胞术检测绿色荧光蛋白的表达反映对CYP17表达的影响,筛选抑制效果最佳的siRNA。卵泡膜细胞取自PCOS患者和非PCOS妇女各5例,将筛选出的最佳CYP17siRNA以最佳浓度转染人卵泡膜细胞,荧光定量实时逆转录多聚酶链反应检测CYP17mRNA的表达,并且检测培养液中CYP17作用产物雄烯二酮和前体孕酮水平。结果:siRNA1221处理后。HeLa细胞中外源性CYP17的表达显著降低,抑制率达50%,卵泡膜细胞中CYP17mRNA水平和雄烯二酮产生降低.但与对照组比较,统计学上无显著差异。结论:RNA干扰技术可能提供一种PCOS高雄激素基因治疗的新方法。

英文摘要:

Objective: To investigate the effects of Cytochrome P450 17α-hydroxylase/17, 20-1yase(CYP17)small interfering RNA (siRNA) on the expression of CYP17 and androgen biosynthesis in theca cells from women with polycystic ovarian syndrome (PCOS). Methods: Four siRNAs targeting the CYP17 gene and one non-specific siRNA were synthesized. 0.4μg of plasmid, in which expressed the enhanced green fluorescent protein (EGFP)reporter gene, the CYP17 gene, and a siRNA were co-transfeeted into HeLa cells, After transfeetion the silencing EGFP was measured by fluorescence microscopy and flow cytometry. Theca cells were isolated from five women with PCOS and five women without PCOS then were cultured in the presence or absence of the most effective siRNA at the most effective concentration. After 72 hours in euhured, the media was collected to evaluate androstenedione and progesterone production. The suppressive effect of the siRNA on CYP17 mRNA was determined by real-time reverse transcriptionpolymerase chain reaction analysis. Results: Following treatment with siRNA 1221, the expression of exogenous CYP17 in HeLa cells was reduced by up to 50%. The CYP17 mRNA expression and androstenedione production of thecal cells were slightly, but not significantly,reduced compared to cell exposed to the non-specific siRNA. Conclusion.RNAi may have applications as gene therapy for reducing hyperandrogenism associated with PCOS.

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期刊信息
  • 《国际妇产科学杂志》
  • 中国科技核心期刊
  • 主管单位:中华人民共和国卫生部
  • 主办单位:天津市医学科学技术信息研究所
  • 主编:郎景和
  • 地址:天津市和平区贵州路94号D座
  • 邮编:300070
  • 邮箱:fuchanfence@163.com
  • 电话:022-23337521
  • 国际标准刊号:ISSN:1674-1870
  • 国内统一刊号:ISSN:12-1399/R
  • 邮发代号:6-1
  • 获奖情况:
  • 国内外数据库收录:
  • 美国化学文摘(网络版),中国中国科技核心期刊
  • 被引量:5391