Petroll W M, Jester J V, Bean J J, Cavanagh H D
Department of Ophthalmology, University of Texas Southwestern Medical Center at Dallas, 75235-9057, USA.
Invest Ophthalmol Vis Sci. 1998 Oct;39(11):2018-32.
Under certain pathophysiologic conditions, the corneal endothelium can produce an abnormal posterior collagenous layer (PCL) that reduces light transmission. Previous studies suggest that formation of PCLs can result from transformation of endothelial cells to a proliferative myofibroblast phenotype. The purpose of this study was to determine the potential role of transforming growth factor (TGF)-beta on corneal endothelial transformation.
Three corneal buttons (6-mm diameter) were obtained from each cornea of 28 adult cats. After a 2-mm diameter mechanical scrape injury was made, each button was cultured for 24, 48, or 72 hours in serum-free medium (SFM) or SFM supplemented with 10% fetal calf serum, TGF-gamma1, TGF-beta2, TGF-beta3, basic fibroblast growth factor (bFGF), or TGF-beta1 and bFGF. Buttons were single and double labeled using phalloidin and antibodies to ZO-1, Ki67, fibronectin, alpha-smooth muscle (SM) actin, and vinculin. Counts of Ki67-positive cells were used as a measure of endothelial proliferation.
Organ culture in TGF-beta1, beta2, or beta3 induced myofibroblast transformation of corneal endothelial cells, with formation of stress fibers containing alpha-SM actin, loss of normal pericellular ZO-1 organization, development of extracellular fibronectin fibrils, and formation of focal contacts as indicated by punctate vinculin staining. However, TGF-beta3 did not stimulate endothelial proliferation above that in serum-free control samples. Serum and bFGF each stimulated proliferation significantly, without inducing myofibroblast transformation. A combination of TGF-beta1 and bFGF resulted in both myofibroblast transformation and increased proliferation.
These results suggest that TGF-beta plays a key role in the loss of normal endothelial differentiation, abnormal extracellular matrix synthesis, and myofibroblast transformation, which can induce development of PCLs. However, other factors such as bFGF seem to be required to stimulate concomitant proliferation of corneal endothelium.
在某些病理生理条件下，角膜内皮可产生异常的后胶原层（PCL），从而降低光透射率。先前的研究表明，PCL的形成可能源于内皮细胞向增殖性肌成纤维细胞表型的转变。本研究的目的是确定转化生长因子（TGF）-β在角膜内皮转化中的潜在作用。
从28只成年猫的每只角膜获取三个角膜片（直径6毫米）。在进行直径2毫米的机械刮伤后，每个角膜片在无血清培养基（SFM）或补充有10%胎牛血清、TGF-γ1、TGF-β2、TGF-β3、碱性成纤维细胞生长因子（bFGF）或TGF-β1和bFGF的SFM中培养24、48或72小时。使用鬼笔环肽和针对ZO-1、Ki67、纤连蛋白、α-平滑肌（SM）肌动蛋白和纽蛋白的抗体对角膜片进行单标和双标。Ki67阳性细胞计数用作内皮细胞增殖的指标。
在TGF-β1、β2或β3中进行器官培养可诱导角膜内皮细胞向肌成纤维细胞转化，形成含有α-SM肌动蛋白的应力纤维，正常的细胞周围ZO-1结构丧失，细胞外纤连蛋白原纤维形成，点状纽蛋白染色显示形成粘着斑。然而，TGF-β3并未刺激内皮细胞增殖超过无血清对照样本。血清和bFGF均显著刺激增殖，但未诱导肌成纤维细胞转化。TGF-β1和bFGF的组合导致肌成纤维细胞转化和增殖增加。
这些结果表明，TGF-β在正常内皮分化丧失、异常细胞外基质合成和肌成纤维细胞转化中起关键作用，这可诱导PCL的形成。然而，似乎需要其他因素如bFGF来刺激角膜内皮的伴随增殖。
