Francesconi C M, Hutcheon A E, Chung E H, Dalbone A C, Joyce N C, Zieske J D
Schepens Eye Research Institute and Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts 02114, USA.
Invest Ophthalmol Vis Sci. 2000 Apr;41(5):1054-62.
To determine the expression patterns of the retinoblastoma protein and the E2F transcription factor families in limbal and corneal epithelia and in corneal keratocytes in situ during corneal development and differentiation.
Retinoblastoma protein (pRb) and its family members p107 and p130; E2F-1, -2, and -4, members of the E2F family of transcription factors; and Ki67, a marker of actively cycling cells, were localized by indirect immunofluorescence microscopy, in corneas of neonatal, juvenile, and adult rats. Presence of mRNA for pRb, p107, p130, and E2F types 1 to 5 in adult corneal epithelium was determined by reverse transcription-polymerase chain reaction.
mRNA for all members of pRb and E2F families was present in adult corneal epithelium. The greatest number of Ki67-positive corneal and limbal epithelial cells were present at days 13 to 19, and Ki67-positive stromal keratocytes at day 2. pRb and E2F-2 were localized to all cells in neonatal, juvenile, and adult corneas. With age, p130 localization became more intense and nuclear in stromal keratocytes and suprabasal cells of corneal and limbal epithelia; p107, initially nuclear in limbal and corneal epithelia, became increasingly cytoplasmic in corneal epithelium. E2F-1 was initially nuclear in keratocytes and diminished after day 10. E2F-1 was localized in the basal cell layer of limbal and corneal epithelia after day 10. E2F4 was always nuclear in limbal epithelium and cytoplasmic in corneal epithelium.
Expression patterns of pRb and E2F family proteins vary with corneal cell differentiation, but are most apparent with p130 and p107. Nuclear localization of p130 appears to correlate with terminal differentiation in epithelium and entrance into a quiescent state by keratocytes. In contrast, p107 is nuclear in the undifferentiated limbal basal cells and is cytoplasmic in the remainder of the corneal epithelial cells.
确定视网膜母细胞瘤蛋白和E2F转录因子家族在角膜发育和分化过程中角膜缘和角膜上皮以及角膜基质细胞原位的表达模式。
通过间接免疫荧光显微镜，在新生、幼年和成年大鼠的角膜中定位视网膜母细胞瘤蛋白（pRb）及其家族成员p107和p130；E2F转录因子家族成员E2F-1、-2和-4；以及活跃增殖细胞的标志物Ki67。通过逆转录-聚合酶链反应确定成年角膜上皮中pRb、p107、p130和E2F 1至5型的mRNA的存在情况。
pRb和E2F家族所有成员的mRNA在成年角膜上皮中均有存在。在第13至19天，Ki67阳性的角膜和角膜缘上皮细胞数量最多，在第2天Ki67阳性的基质角膜细胞数量最多。pRb和E2F-2定位于新生、幼年和成年角膜的所有细胞。随着年龄增长，p130在角膜基质细胞以及角膜和角膜缘上皮的基底上层细胞中的定位变得更加明显且呈核定位；p107最初在角膜缘和角膜上皮中呈核定位，在角膜上皮中逐渐变为胞质定位。E2F-1最初在角膜细胞中呈核定位，在第10天后减少。第10天后，E2F-1定位于角膜缘和角膜上皮的基底细胞层。E2F4在角膜缘上皮中始终呈核定位，在角膜上皮中呈胞质定位。
pRb和E2F家族蛋白的表达模式随角膜细胞分化而变化，但在p130和p107中最为明显。p130的核定位似乎与上皮细胞的终末分化以及角膜细胞进入静止状态相关。相比之下，p107在未分化的角膜缘基底细胞中呈核定位，在角膜上皮的其余细胞中呈胞质定位。
