Guillonneau X, Tassin J, Berrou E, Bryckaert M, Courtois Y, Mascarelli F
Unité de Recherches Gérontologiques INSERM U. 118, Affiliée CNRS, Association Claude-Bernard, Paris, France.
J Cell Physiol. 1996 Jan;166(1):170-87. doi: 10.1002/(SICI)1097-4652(199601)166:1<170::AID-JCP19>3.0.CO;2-J.
Fibroblast growth factor 1 (FGF1) and 2 (FGF2) bind to two classes of receptors: the high affinity receptors, a family of four known transmembrane tyrosine kinases (FGF R1-R4), and the low affinity receptors, cell surface and basement membrane heparan sulfate proteoglycan (HSPG). During early (first and second) passages of retinal pigmented epithelial (RPE) cells, both FGF1 and FGF2 exhibited low mitogenic activity, while in later (fifth to ninth) passages the activity of FGF1 remained constant but FGF2 activity increased two- to threefold. We have investigated aspects of FGF receptor interactions and the role of heparin/heparan sulfate which modulates FGF activity on RPE cells during in vitro senescence. Northern blot analysis demonstrated that FGF receptor type 1 (FGF R1) is the major high affinity receptor expressed in RPE cells and that its level of expression did not change during serially passage. Both the FGF R1 and the FGF low affinity receptors' binding characteristics (i.e., Kd and number of sites per cell) for FGF1 were unaffected by passage number, whereas the capacity of FGF2 binding to FGF R1 and to the low affinity receptors increased by two- and fivefold, respectively, in late passages, although the affinities were unchanged. This change in the capacity of FGF2 to bind to FGF R1 and to HSPG was not due to a switch of the IIIc splice form of FGF R1 to the IIIb splice form since the exon IIIc was the most predominant splice form of FGF R1 during RPE cell cultures. Furthermore the ratio of the IIIb to the IIIc splice form was not modified during cell subcultures. In parallel in the older RPE cell passages, expression of perlecan, the major FGF low affinity binding site localized on the extracellular matrix of RPE cells, was much elevated compared to early RPE cell passages. Moreover, the cell surface of late passage RPE cells had 79% more HSPG than early passage cells. Therefore, it is suggested that the increase in the number of FGF low affinity receptors present on the cell surface or basement membrane could account for a part of the greater proliferative response of aged RPE cells to FGF2.
成纤维细胞生长因子1（FGF1）和2（FGF2）可与两类受体结合：高亲和力受体，即一个由四种已知跨膜酪氨酸激酶组成的家族（FGF R1 - R4），以及低亲和力受体，即细胞表面和基底膜硫酸乙酰肝素蛋白聚糖（HSPG）。在视网膜色素上皮（RPE）细胞的早期传代（第一代和第二代）过程中，FGF1和FGF2均表现出较低的促有丝分裂活性，而在后期传代（第五代至第九代）中，FGF1的活性保持恒定，FGF2的活性则增加了两到三倍。我们研究了FGF受体相互作用的相关方面以及肝素/硫酸乙酰肝素在体外衰老过程中对RPE细胞FGF活性的调节作用。Northern印迹分析表明，FGF受体1型（FGF R1）是RPE细胞中表达的主要高亲和力受体，其表达水平在连续传代过程中没有变化。FGF R1和FGF低亲和力受体对FGF1的结合特性（即解离常数Kd和每个细胞的结合位点数量）不受传代次数的影响，而在后期传代中，FGF2与FGF R1和低亲和力受体的结合能力分别增加了两倍和五倍，尽管亲和力未变。FGF2与FGF R1和HSPG结合能力的这种变化并非由于FGF R1的IIIc剪接形式转换为IIIb剪接形式，因为外显子IIIc是RPE细胞培养过程中FGF R1最主要的剪接形式。此外，在细胞传代培养过程中，IIIb与IIIc剪接形式的比例没有改变。同时，在较老的RPE细胞传代中，与早期RPE细胞传代相比，位于RPE细胞外基质上的主要FGF低亲和力结合位点核心蛋白聚糖的表达显著升高。此外，后期传代RPE细胞表面的HSPG比早期传代细胞多79%。因此，有人认为细胞表面或基底膜上FGF低亲和力受体数量的增加可能是衰老RPE细胞对FGF2产生更大增殖反应的部分原因。
