Tong Louis, Corrales Rosa M, Chen Zhuo, Villarreal Arturo L, De Paiva Cintia S, Beuerman Roger, Li De-Quan, Pflugfelder Stephen C
Ocular Surface Center, Cullen Eye Institute, Baylor College of Medicine, Houston, Texas 77030, USA.
Invest Ophthalmol Vis Sci. 2006 May;47(5):1938-46. doi: 10.1167/iovs.05-1129.
Stratified squamous epithelial cells assemble a specialized protective barrier structure on their periphery, termed the cornified envelope. The purpose of this study was to evaluate the presence and distribution of cornified envelope precursors in human corneal epithelium, their expression in human corneal epithelial cell cultures, and the effect of ultraviolet radiation (UVB) and transglutaminase (TG) inhibition on their expression.
Tissue distribution of small proline-rich proteins (SPRRs) and filaggrin and involucrin was studied in human cornea sections by immunofluorescence staining. Primary human corneal epithelial cells (HCECs) from limbal explants were used in cell culture experiments. A single dose of UVB at 20 mJ/cm2 was used to stimulate these cells, in the presence or absence of mono-dansyl cadaverine (MDC), a TG inhibitor. SPRR2 and involucrin protein levels were studied by immunofluorescence staining and Western blot analysis. Gene expression of 12 proteins was investigated by semiquantitative reverse transcription-polymerase chain reaction.
In human cornea tissue, SPRR1, SPRR2, filaggrin, and involucrin protein expression were detected in the central and peripheral corneal and limbal epithelium. In HCECs, SPRR2 and involucrin proteins were detected in the cytosolic fraction, and involucrin levels increased after UVB. Both SPRR2 and involucrin levels accumulated in the presence of MDC. Nine genes including involucrin, SPRR (types 1A, 1B, 2A, 2B, and 3), late envelope protein (LEP) 1 and 16, and filaggrin were expressed by HCECs. SPRR 4, loricrin, and LEP 6 transcripts were not detected. UVB downregulated SPRR (2A, 2B) and LEP 1 transcripts.
Various envelope precursors are expressed in human corneal epithelium and in HCECs, acute UVB stress differentially alters their expression in HCECs. The expression of envelope precursors and their rapid modulation by UVB supports the role of these proteins in the regulation of ocular surface stress. TG function may be relevant in the regulation of soluble precursors in UVB-stimulated corneal epithelium.
复层鳞状上皮细胞在其周边组装一种特殊的保护性屏障结构，称为角质包膜。本研究的目的是评估角质包膜前体在人角膜上皮中的存在和分布、它们在人角膜上皮细胞培养物中的表达，以及紫外线辐射（UVB）和转谷氨酰胺酶（TG）抑制对其表达的影响。
通过免疫荧光染色研究富含脯氨酸的小蛋白（SPRRs）、丝聚合蛋白和内披蛋白在人角膜切片中的组织分布。来自角膜缘外植体的原代人角膜上皮细胞（HCECs）用于细胞培养实验。在存在或不存在TG抑制剂单丹磺酰尸胺（MDC）的情况下，使用20 mJ/cm²的单剂量UVB刺激这些细胞。通过免疫荧光染色和蛋白质印迹分析研究SPRR2和内披蛋白的蛋白水平。通过半定量逆转录-聚合酶链反应研究12种蛋白质的基因表达。
在人角膜组织中，在角膜中央和周边以及角膜缘上皮中检测到SPRR1、SPRR2、丝聚合蛋白和内披蛋白的蛋白表达。在HCECs中，在细胞溶质部分检测到SPRR2和内披蛋白，UVB照射后内披蛋白水平升高。在MDC存在的情况下，SPRR2和内披蛋白水平均积累。HCECs表达包括内披蛋白、SPRR（1A、1B、2A、2B和3型）、晚期包膜蛋白（LEP）1和16以及丝聚合蛋白在内的9种基因。未检测到SPRR 4、兜甲蛋白和LEP 6转录本。UVB下调SPRR（2A、2B）和LEP 1转录本。
多种包膜前体在人角膜上皮和HCECs中表达，急性UVB应激差异地改变它们在HCECs中的表达。包膜前体的表达及其受UVB的快速调节支持了这些蛋白质在眼表应激调节中的作用。TG功能可能与UVB刺激的角膜上皮中可溶性前体的调节有关。
