摘要
采用全基因组表达芯片技术研究反复自然流产(recurrent spontaneous abortion,RSA)患者与正常早孕妇女滋养细胞(trophoblast,TB)中凋亡相关基因表达差异,探寻RSA发病机制。收集正常妊娠和RSA妇女胚胎绒毛,分离滋养层细胞并进行原代培养,提取细胞总RNA,逆转录为双链cDNA后制备生物素标记的cRNA探针,探针与Affymetrix U133 plus2.0基因芯片杂交后,经Affymetrix GeneChip Scanner3000读取荧光图象、GeneChip Operating software(GCOS)软件分析后,将数据导入GeneSpringGX软件分析,获得了差异表达(≥2)凋亡相关基因75个,其中表达上调23个,下调52个。层次聚类结果显示,凋亡相关基因在两类细胞中表现出明显的差异:RSA患者分离的TB细胞中,促凋亡基因多表现为表达上调,而凋亡抑制基因则多表现为表达下调。差异表达的凋亡相关基因涉及两条重要的信号转导通路,即Foxo家族信号通路和IL6介导的信号转导通路。这些研究结果为不明原因反复自然流产的病因研究打下了实验基础。
The whole genome expression chips were used to find differentially expressed apoptotic genes in the primarily cultured trophoblast （TB） cells from pregnant women with recurrent spontaneous abortion （RSA）. The collected TB cells were isolated and primarily cultured from normal pregnant women and the patients with RSA. The total RNA was extracted and reverse transcribed into double-strand cDNA, and then transcribed into biotinlabled cRNA target probes. The probes were hybridized with Affymetrix U133 plus2.0. Subsequently, the signal images were scanned by Affymetrix GeneChip Scanner 3000 and analyzed with GeneChip Operating software （GCOS）. Gene expression profile was analyzed by using Genespring GX. Seventy-five apoptosis related genes expressed differentially were found, among which, 23 genes were up-regulated and 52 genes were down-regulated. Hierarchical cluster analysis showed apoptosis related genes expressed differently between the two groups. The genes promoting apoptosis are almost represented as up-regulated, but the anti-apoptosis genes are represented as down-regulated in the TB cells isolated from patients with RSA. Two signaling pathways, FoxO family signaling and IL6-mediated signaling events pathways, were found related to RSA. The results will contribute to find out the mechanism responsible to unexplained RSA.
出处
《细胞生物学杂志》
CSCD
2009年第2期250-256,共7页
Chinese Journal of Cell Biology
基金
重庆医科大学校办基金(No.XBYB2007111)
重庆市科委自然科学基金(No.CSTC
2007BB5286)资助~~
关键词
反复自然流产
滋养细胞
基因芯片
基因表达谱
凋亡
recurrent spontaneous abortion
trophoblastic cells
gene chip
gene expression profile
apoptosis
