What a study of pterygia teaches us about the cornea? Molecular mechanisms of formation.

Experiments were carried out in the early 1990s to investigate the cell types involved in a pterygium and to determine a possible mechanism of formation. Our first experiments used monoclonal antibodies to keratins and an associated protein (vimentin), to look at the cells that compose a pterygium. These experiments demonstrated that a pterygium is the result of an abnormal limbal basal epithelial stem cell that moves onto Bowman's layer and brings about the dissolution of this layer. More importantly, these data showed that the clear corneal epithelial cells in front of the pterygium also contained these abnormal limbal cells, which we named the pterygium cell. This demonstrated that when a pterygium is removed, a wide area of what appears to be normal epithelium must be removed to inhibit reoccurrence of the growth. Later experiments using expressed sequence tag analysis of an un-normalized unamplified complementary DNA library from surgically removed pterygia were compared with normal cornea and confirmed the role of the epithelial cells in this growth. The gene expression studies also showed that genes involved in cellular migration are stimulated, and this led to studies on polyamine analogs as inhibitors of pterygial migration. Immunohistochemical studies with antibodies to matrix metalloproteinases (MMPs) showed that it is the pterygium cell that produces the MMPs that dissolve Bowman's layer resulting in the growth stimulation of stromal fibroblasts. This led to experiments on the use of MMP inhibitors to inhibit the growth of pterygia.

MLH1 and MSH2 expression in pterygia.

PURPOSE: Pterygia have been reported to share some of the genetic defects seen in cancers, including microsatellite instability (MSI). We examined pterygia for the presence of proteins typically missing or defective in adenocarcinomas with MSI. We also performed microsatellite analysis on DNA from pterygia to test for instability in the size of the microsatellites, using markers conventionally used to characterize MSI in tumors (Bethesda convention markers). METHODS: We examined 13 pterygia by immunohistochemistry for MLH1 and MSH2, 2 proteins involved in DNA mismatch repair. In addition, we amplified the pterygial DNA with primers specific for 5 Bethesda markers (BAT25, BAT26, D2S123, D5S346, and D17S250). RESULTS: MLH1 staining was present at low levels in the basal cells of the cornea and migrating limbal cells of the pterygia. MSH2 staining was present in basal and in maturing epithelial cells of the cornea and migrating limbal cells of pterygia. We observed no reproducible examples of MSI or loss of heterozygosity (LOH). CONCLUSIONS: We were unable to confirm the presence of MSI and LOH by using the markers we examined. MSH2 staining appeared to be normal in pterygia. MLH1 staining was present but in reduced amounts compared with that seen in the conjunctiva.

Microarray and protein analysis of human pterygium.

PURPOSE: Pterygium is a sunlight-related, ocular-surface lesion that can obscure vision. In order to identify specific genes that may play a role in pterygium pathogenesis, we analyzed the global gene expression profile of pterygium in relation to autologous conjunctiva. METHODS: Oligonucleotide microarray hybridization was used to compare the gene expression profile between human whole pterygium and autologous conjunctiva. Selected genes were further characterized by RT-PCR, western blot, and immunohistochemistry, and comparisons were made with limbal and corneal tissues. RESULTS: Thirty-four genes exhibited a 2 fold or greater difference in expression between human whole pterygium and autologous conjunctiva. Twenty-nine transcripts were increased and five transcripts were decreased in pterygium. Fibronectin, macrophage-inflammatory protein-4 (MIP-4), and lipocalin 2 (oncogene 24p3; NGAL) were increased 9, 5, and 2.4 fold, respectively, while Per1 and Ephrin-A1 were decreased 2 fold in pterygium. Western blots showed that fibronectin and MIP-4 were increased in pterygium compared to limbus, cornea, and conjunctiva. Immunohistochemical analysis showed fibronectin in the stroma; lipocalin 2 in the basal epithelial cells, basement membrane, and extracellular stroma; and MIP-4 in all areas of the pterygium. CONCLUSIONS: These data show both novel and previously identified extracellular-matrix-related, proinflammatory, angiogenic, fibrogenic, and oncogenic genes expressed in human pterygium. Comparisons of selected genes with limbal and corneal tissues gave results similar to comparisons between pterygium and normal conjunctiva. The increased expression of lipocalin 2, which activates matrix metalloproteinases (MMP), is consistent with our previous findings that MMP-9 and other MMPs are highly expressed in pterygium basal epithelium.