Yang Ping, Agapova Olga, Parker Amy, Shannon William, Pecen Paula, Duncan Jill, Salvador-Silva Mercedes, Hernandez M Rosario
Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Physiol Genomics. 2004 Apr 13;17(2):157-69. doi: 10.1152/physiolgenomics.00182.2003.
There is clinical and experimental evidence that elevated intraocular pressure (IOP), a mechanical stress, is involved in the pathogenesis of glaucomatous optic neuropathy. The mechanism by which astrocytes in the optic nerve head (ONH) respond to changes in IOP is under study. Gene transcription by ONH astrocytes exposed either to 60 mmHg hydrostatic pressure (HP) or control ambient pressure (CP) for 6, 24, and 48 h was compared using Affymetrix GeneChip microarrays to identify HP-responsive genes. Data were normalized across arrays within each gene. A linear regression model applied to test effect of time and HP on changes in expression level identified 596 genes affected by HP over time. Using GeneSpring analysis we selected genes whose average expression level increased or decreased more than 1.5-fold at 6, 24, or 48 h. Expression of selected genes was confirmed by real-time RT-PCR; protein levels were detected by Western blot. Among the genes highly responsive to HP were those involved in signal transduction, such as Rho nucleotide exchange factors, Ras p21 protein activator, tyrosine kinases and serine threonine kinases, and genes involved in transcriptional regulation, such as c-Fos, Egr2, and Smad3. Other genes that increased expression included ATP-binding cassettes, solute carriers, and genes associated with lipid metabolism. Among the genes that decreased expression under HP were genes encoding for dual activity phosphatases, transcription factors, and enzymes involved in protein degradation. These HP-responsive genes may be important in the establishment and maintenance of the ONH astrocyte phenotype under conditions of elevated IOP in glaucoma.
有临床和实验证据表明，眼内压（IOP）升高这种机械应力参与了青光眼性视神经病变的发病机制。视神经乳头（ONH）中的星形胶质细胞对IOP变化作出反应的机制正在研究中。使用Affymetrix基因芯片微阵列比较了暴露于60 mmHg静水压力（HP）或对照环境压力（CP）下6、24和48小时的ONH星形胶质细胞的基因转录，以鉴定HP反应性基因。数据在每个基因的阵列之间进行了标准化。应用线性回归模型来测试时间和HP对表达水平变化的影响，确定了596个随时间受HP影响的基因。使用GeneSpring分析，我们选择了在6、24或48小时时平均表达水平增加或减少超过1.5倍的基因。通过实时RT-PCR确认所选基因的表达；通过蛋白质印迹检测蛋白质水平。对HP高度反应的基因包括参与信号转导的基因，如Rho核苷酸交换因子、Ras p21蛋白激活剂、酪氨酸激酶和丝氨酸苏氨酸激酶，以及参与转录调控的基因，如c-Fos、Egr2和Smad3。其他表达增加的基因包括ATP结合盒、溶质载体以及与脂质代谢相关的基因。在HP作用下表达降低的基因包括编码双活性磷酸酶、转录因子和参与蛋白质降解的酶的基因。这些HP反应性基因可能在青光眼IOP升高条件下ONH星形胶质细胞表型的建立和维持中起重要作用。
