Ibaraki N, Lin L R, Reddy V N
Eye Research Institute, Oakland University, Rochester, MI 48309, USA.
Exp Eye Res. 1996 Dec;63(6):683-92. doi: 10.1006/exer.1996.0162.
Recent studies have demonstrated that several growth factors enhance fiber differentiation in cultured human lens epithelial (HLE) cells in early passages. However, these effects gradually decrease in cells of later passages. The purpose of this investigation is to test the hypothesis that the decreasing effect of growth factors on fiber differentiation in later passages may be due to a decrease or the inactivation of growth factor receptors as a function of serial subcultures. Specimens of HLE cells were obtained from infants. First through to fourth passage cells were treated with 10 ng ml-1 of epidermal growth factor, basic fibroblast growth factor or insulin-like growth factor-I. Fiber differentiation was determined from spontaneous lentoid formation by phase-contrast and transmission electron microscopy. Growth factor binding to the receptor on the cell surface was determined by transmission electron microscopy using the conjugates of colloidal gold and growth factors, and the number of receptors on the cell surface were also quantified by immunocytochemistry. Spontaneous lentoid formation was enhanced by all of the growth factors studied in the first passage. However, in the second and third passage only double layering of cells without characteristic fiber differentiation was observed while in the fourth passage, growth factors had no effect on differentiation. The number of growth factor bindings as well as the number of growth factor receptors gradually decreased with the number of passages. The loss of effect of growth factors on fiber differentiation with increasing number of passages correlated with the decrease in receptor number.
最近的研究表明，几种生长因子可增强早期传代培养的人晶状体上皮（HLE）细胞中的纤维分化。然而，在后期传代的细胞中，这些作用会逐渐减弱。本研究的目的是检验以下假设：生长因子对后期传代细胞中纤维分化作用的减弱可能是由于生长因子受体随连续传代培养而减少或失活所致。HLE细胞样本取自婴儿。将第一代至第四代的细胞用10 ng/ml的表皮生长因子、碱性成纤维细胞生长因子或胰岛素样生长因子-I进行处理。通过相差显微镜和透射电子显微镜观察自发晶状体样体的形成来确定纤维分化情况。使用胶体金与生长因子的结合物，通过透射电子显微镜测定生长因子与细胞表面受体的结合情况，并用免疫细胞化学方法对细胞表面的受体数量进行定量。在所研究的所有生长因子中，第一代细胞的自发晶状体样体形成均得到增强。然而，在第二代和第三代细胞中，仅观察到细胞的双层排列，而无特征性的纤维分化，在第四代细胞中，生长因子对分化没有影响。随着传代次数的增加，生长因子结合数量以及生长因子受体数量逐渐减少。生长因子对纤维分化的作用随传代次数增加而丧失，这与受体数量的减少相关。
