Characterization of corneal pannus removed from patients with total limbal stem cell deficiency
Characterization of corneal pannus removed from patients with total limbal stem cell deficiency
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DOI：
10.1167/iovs.03-1397
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发表时间：
2004-09-01
影响因子：
4.4
通讯作者：
Tseng, SCG
中科院分区：
文献类型：
--
作者：
Espana, EM;Di Pascuale, MA;Tseng, SCG
目的.确定从角膜缘干细胞（SC）完全缺乏患者手术切除的角膜血管翳组织中起源的上皮谱系。方法。通过抗角蛋白（K）-3和抗K19单克隆抗体对诊断为角膜缘SC缺乏的8例角膜中取出的整个结膜血管翳的起源谱系进行了表征。通过Western blot和逆转录-聚合酶链反应分别分析了5例血管翳标本的上皮生长的蛋白质和mRNA。所有8例标本的横截面均显示复层上皮，其中杯状细胞表达粘蛋白（MUC）-5AC，基质显示血管和炎性细胞浸润。免疫组化显示，所有8个标本的整个血管翳中全层表达K19。K3的表达在7例患者中为阴性，但在Stevens-Johnson综合征患者中为零星阳性。在培养中，所有5个血管翳标本均产生了致密的小上皮细胞生长，除1个外，均在2 - 3周内达到融合。K3/K12对由对照角膜缘上皮外植体的细胞生长物提取物表达，但在所有5个血管翳标本中均未表达。对照标本和所有五份血管翳标本中均表达了DeltaNp 63的60 kDa条带。细胞生长产物中有3个表达K3转录本，无1个表达K12转录本。血管翳组织的所得上皮表型不是角膜的，如角膜特异性K12 mRNA和蛋白的阴性染色所证明的，而是结膜的，如杯状细胞的存在、K3的弱表达和K19的强表达所证明的。DeltaNp 63在患有角膜缘SC完全缺乏的眼睛中的这种结膜来源的上皮中的丰富表达引起了对其作为角膜缘SC标记物的有效性的怀疑。
PURPOSE. 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.