Lee Ji-Eun, Oum Boo Sup, Choi Hee Young, Lee Seung Uk, Lee Jong Soo
The Department of Ophthalmology, College of Medicine, Pusan National University, Pusan, Korea.
Mol Vis. 2009 Nov 28;15:2480-7.
To identify the differentially expressed genes (DEGs) in the human keratocytes in keratoconus.
Total RNA extracted from cultured corneal stromal fibroblasts from normal and keratoconic corneas were used for the synthesis of cDNA. DEGs were screened by an annealing control primer(TM)-based PCR method using GeneFishing() DEG kits. The differentially expressed bands were sequenced and analyzed. The genes identified were further evaluated by reverse transcriptase PCR and quantitative real-time PCR.
Overexpression of bone morphogenetic protein 4 (BMP4), cofilin 1 (CFL1), and JAW1-related protein (MRVI1) and underexpression of actin, alpha 2 (ACTA2), gene rich cluster, and C 10 gene (GRCC10), tissue inhibitor of metalloproteinase 3 (TIMP3), tissue inhibitor of metalloproteinase 1 (TIMP1), and somatostatin receptor 1 (SSTR1) were verified, and these results were confirmed by reverse transcriptase PCR and quantitative real-time PCR.
Eight genes were identified to be differentially expressed in keratoconus and related with apoptosis, the cytoskeleton, wound healing, and nerve fibers. The genes identified may be involved in the mechanism underlying stromal thinning; thus, they could be important and deserve further investigation.
鉴定圆锥角膜患者人角膜细胞中差异表达基因（DEGs）。
从正常角膜和圆锥角膜培养的角膜基质成纤维细胞中提取的总RNA用于合成cDNA。使用GeneFishing() DEG试剂盒，通过基于退火控制引物（TM）的PCR方法筛选DEGs。对差异表达条带进行测序和分析。通过逆转录PCR和定量实时PCR对鉴定出的基因进行进一步评估。
骨形态发生蛋白4（BMP4）、丝切蛋白1（CFL1）和JAW1相关蛋白（MRVI1）的过表达以及肌动蛋白α2（ACTA2）、富含基因簇和C10基因（GRCC10）、金属蛋白酶组织抑制剂3（TIMP3）、金属蛋白酶组织抑制剂1（TIMP1）和生长抑素受体1（SSTR1）的低表达得到验证，这些结果通过逆转录PCR和定量实时PCR得到证实。
鉴定出8个在圆锥角膜中差异表达且与细胞凋亡、细胞骨架、伤口愈合和神经纤维相关的基因。鉴定出的这些基因可能参与基质变薄的潜在机制；因此，它们可能很重要，值得进一步研究。
