Jaworski C J, Aryankalayil-John M, Campos M M, Fariss R N, Rowsey J, Agarwalla N, Reid T W, Dushku N, Cox C A, Carper D, Wistow G
Section on Molecular Therapeutics, NEI/NIH, Bethesda, MD 20892, USA.
Mol Vis. 2009 Nov 20;15:2421-34.
Pterygium is a vision-impairing fibrovascular lesion that grows across the corneal surface and is associated with sunlight exposure. To increase our understanding of the cells types involved in pterygium, we have used expressed sequence tag analysis to examine the transcriptional repertoire of isolated pterygium and to identify marker genes for tissue origin and cell migration.
An unnormalized unamplified cDNA library was prepared from 15 pooled specimens of surgically removed pterygia as part of the NEIBank project. Gene expression patterns were compared with existing data for human cornea, limbus, and conjunctiva, and expression of selected genes was verified by immunofluorescence localization in normal eye ocular surface and in pterygium.
Sequence analysis of 2,976 randomly selected clones produced over 1,800 unique clusters, potentially representing single genes. The most abundant complementary DNAs from pterygium include clusterin, keratins 13 (Krt13) and 4 (Krt4), S100A9/calgranulin B, and spermidine/spermine N1-acetyltransferase (SAT1). Markers for both conjunctiva (such as keratin 13/4 and AQP3) and corneal epithelium (such as keratin 12/3 and AQP5) were present. Immunofluorescence of Krt12 and 13 in the normal ocular surface showed specificity of Krt12 in cornea and Krt13 in conjunctival and limbal epithelia, with a fairly sharp boundary at the limbal-corneal border. In the pterygium there was a patchy distribution of both Krt12 and 13 up to a normal corneal epithelial region specific for Krt12. Immunoglobulins were also among the prominently expressed transcripts. Several of the genes expressed most abundantly in excised pterygium, particularly S100A9 and SAT1, have roles in cell migration. SAT1 exerts its effects through control of polyamine levels. IPENSpm, a polyamine analogue, showed a significant ability to reduce migration in primary cultures of pterygium. A number of genes highly expressed in cornea were not found in pterygium (several small leucine-rich proteoglycan family members) or were expressed at considerably lower levels (ALDH3A1 and decorin).
The expression pattern of keratins and other markers in pterygium most closely resemble those of conjunctival and limbal cells; some corneal markers are present, notably Krt12, but at lower levels than equivalent conjunctival markers. Our data are consistent with the model of pterygium developing from the migration of conjunctival- and limbal-like cells into corneal epithelium. Identification of genes with roles in cell migration suggests potential therapeutic targets. In particular, the ability of polyamine analogues to reduce migration in primary cultures of pterygium presents a possible approach to slowing pterygium growth.
翼状胬肉是一种可损害视力的纤维血管性病变，生长于角膜表面，与阳光照射相关。为增进我们对翼状胬肉中所涉及细胞类型的了解，我们利用表达序列标签分析来检测分离出的翼状胬肉的转录组，并确定组织起源和细胞迁移的标记基因。
作为国家眼科研究所生物银行项目的一部分，从15例手术切除的翼状胬肉合并标本中制备了一个未标准化、未扩增的cDNA文库。将基因表达模式与人类角膜、角膜缘和结膜的现有数据进行比较，并通过免疫荧光定位在正常眼表和翼状胬肉中验证所选基因的表达。
对2976个随机选择的克隆进行序列分析产生了1800多个独特的簇，可能代表单个基因。翼状胬肉中最丰富的互补DNA包括簇集蛋白、角蛋白13（Krt13）和4（Krt4）、S100A9/钙粒蛋白B以及亚精胺/精胺N1-乙酰转移酶（SAT1）。结膜（如角蛋白13/4和水通道蛋白3）和角膜上皮（如角蛋白12/3和水通道蛋白5）的标记物均存在。正常眼表中Krt12和13的免疫荧光显示Krt12在角膜中具有特异性，而Krt13在结膜和角膜缘上皮中具有特异性，在角膜缘-角膜边界处有相当清晰的界限。在翼状胬肉中，Krt12和13均呈斑片状分布，直至Krt12特异性的正常角膜上皮区域。免疫球蛋白也在显著表达的转录本中。在切除的翼状胬肉中表达最丰富的几个基因，特别是S100A9和SAT1，在细胞迁移中起作用。SAT1通过控制多胺水平发挥作用。多胺类似物IPENSpm在翼状胬肉原代培养物中显示出显著的减少迁移的能力。在翼状胬肉中未发现一些在角膜中高表达的基因（几个富含亮氨酸的小分子蛋白聚糖家族成员），或者其表达水平显著较低（醛脱氢酶3A1和核心蛋白聚糖）。
翼状胬肉中角蛋白和其他标记物的表达模式与结膜和角膜缘细胞的表达模式最为相似；存在一些角膜标记物，尤其是Krt12，但水平低于相应的结膜标记物。我们的数据与翼状胬肉由结膜样和角膜缘样细胞迁移至角膜上皮而形成的模型一致。鉴定在细胞迁移中起作用的基因提示了潜在的治疗靶点。特别是，多胺类似物在翼状胬肉原代培养物中减少迁移的能力为减缓翼状胬肉生长提供了一种可能的方法。
