Characterization of corneal pannus removed from patients with total limbal stem cell deficiency.
OBJECTIVE
To determine the epithelial lineage of origin in corneal pannus tissue surgically removed from patients with total limbal stem cell (SC) deficiency.


METHODS
The lineage of origin of the entire conjunctivalized pannus removed from eight corneas with a diagnosis of total limbal SC deficiency was characterized by anti-keratin (K)-3 and anti-K19 monoclonal antibodies. The protein and mRNA of epithelial outgrowth from segments of five such pannus specimens were analyzed by Western blot and reverse transcription-polymerase chain reaction, respectively.


RESULTS
Cross sections of all eight specimens showed a stratified epithelium with goblet cells expressing mucin (MUC)-5AC, and a stroma showing blood vessels and inflammatory cell infiltrates. Immunostaining showed full-thickness expression of K19 in the entire pannus of all eight specimens. Expression of K3 was negative in seven patients, but was sporadically positive in a patient with Stevens-Johnson syndrome. In culture, all five pannus specimens generated a compact, small epithelial cell outgrowth, and except for one, reached confluence in 2 to 3 weeks. The K3/K12 pair was expressed by extracts of cell outgrowth from the control limbal epithelial explant, but not in all five pannus specimens. A 60-kDa band of DeltaNp63 was expressed in the control specimen and in all five pannus specimens. Cell outgrowth expressed K3 transcript in three, but none showed K12 transcript.


CONCLUSIONS
The resultant epithelial phenotype of the pannus tissue was not corneal, as evidenced by the negative staining to cornea-specific K12 mRNA and protein, but was conjunctival, as evidenced by the presence of goblet cells, the weak expression of K3, and the strong expression of K19. The abundant expression of DeltaNp63 in such conjunctiva-derived epithelium in eyes with total limbal SC deficiency raises doubts as to its validity as a limbal SC marker.
Espana
Edgar M
EM
Ocular Surface Center, Miami, Florida 33173, USA.
Di Pascuale
Mario A
MA
He
Hua
H
Kawakita
Tetsuya
T
Raju
Vadrevu K
VK
Liu
Chia-Yang
CY
Tseng
Scheffer C G
SC
D000328:Adult; D002648:Child; D016510:Corneal Neovascularization / Q000378:metabolism / Q000473:pathology* / Q000601:surgery*; D004268:DNA-Binding Proteins / Q000378:metabolism; D019573:Epithelium, Corneal / Q000378:metabolism / Q000473:pathology; D005260:Female; D016147:Genes, Tumor Suppressor; D020397:Goblet Cells / Q000473:pathology; D006801:Humans; D053551:Keratin-12; D053558:Keratin-3; D007633:Keratins / Q000235:genetics / Q000378:metabolism; D016850:Limbus Corneae / Q000378:metabolism / Q000473:pathology*; D008297:Male; D008875:Middle Aged; D055257:Mucin-5B; D009077:Mucins / Q000378:metabolism; D010750:Phosphoproteins; D012333:RNA, Messenger / Q000378:metabolism; D013234:Stem Cells / Q000473:pathology*; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins
D004268:DNA-Binding Proteins; C507494:KRT12 protein, human; C479360:KRT3 protein, human; D053551:Keratin-12; D053558:Keratin-3; C524578:MUC5B protein, human; D055257:Mucin-5B; D009077:Mucins; D010750:Phosphoproteins; D012333:RNA, Messenger; C115051:TP63 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D007633:Keratins