不同区域晶状体上皮细胞基因表达差异分析

Different Gene Expression Between Central and Peripheral Epithelial Cells in Porcine Lens

【目的】比较不同区域猪晶状体上皮细胞的基因表达在转录组水平上的差异。【方法】解剖显微镜下分离晶状体前囊膜,将附着于其上的上皮细胞分为中央(直径为10.56mm)和周边两部分。分别提取两个样本的总RNAs并经PCR扩增,以Cy3和Cy5分别标记扩增的中央与周边部分的cDNA,与含7548个基因的表达谱芯片杂交,经图像分析,生物信息学处理获得基因表达在转录水平差异的相关信息。【结果】中央与周边区域的猪晶状体上皮细胞在转录组水平共鉴定出952个有效表达的基因点,其中差异表达基因261个,以中央区域为参照,周边上皮细胞mRNA上调137个,下调124个。差异表达基因主要涉及的功能有:细胞周期与凋亡、细胞骨架蛋白及细胞外基质、转录、细胞信号分子等。【结论】中央与周边区域猪晶状体上皮细胞基因表达在转录组水平上差异明显。这类差异呈明显的功能聚类。

[Objective] To compare the gene expression between the central and the peripheral epithelial cells in the porcine lens.[Methods] The anterior sub-capsular epithelial cells adherent to the porcine lens capsule were dissected into the central （6 mm, diameter） and the peripheral regions. Total RNAs were extracted from epithelia of beth parts and amplified by PCR. The products were incorporated with fluorescent markers to prepare the hybridization probes. The mixed probes were hybridized to a microarray containing 7548 ESTs. After high stringent washing, the chip was scanned for the fluorescent signals and the differences between two tissues were analyzed by software （Imagequant 5.0; Array Vision 6.0）.[Results] Totally 952 transcripts with efficient hybridization were found on the gene microarray. The ratio of fluorescent intensities between two samples displayed obvious alteration in 261 genes, including 137 up-regulated and 124 down-regulated in peripheral area as compared with central parts. According to the functional classification, the genes differendy expressed mainly include signal transduction, cell cycle, apoptosis, transcription factor, extracellular matrix and cytoskeleton. [Conclusion] There were different genome activities during lens epithelial differentiation from central area to periphery. The results of regional alteration of gene expression provide clues for further study.