Carloni Vinicio, Defranco Raffaella M S, Caligiuri Alessandra, Gentilini Alessandra, Sciammetta Silvia Cappadona, Baldi Elisabetta, Lottini Benedetta, Gentilini Paolo, Pinzani Massimo
Dipartimento di Medicina Interna and Dipartimento di Fisiopatologia Clinica-Unità di Andrologia, Università degli Studi di Firenze, Florence, Italy.
Hepatology. 2002 Sep;36(3):582-91. doi: 10.1053/jhep.2002.35277.
The biologic effects of growth factors are dependent on cell adhesion, and a cross talk occurs between growth factors and adhesion complexes. The aim of the present study was to evaluate the influence of cell adhesion on the major intracellular signaling pathways elicited by platelet-derived growth factor (PDGF) in hepatic stellate cells (HSC). PDGF signaling was investigated in an experimental condition characterized by lack of cell adhesion for different intervals of time. Basal and PDGF-induced focal adhesion kinase (FAK) tyrosine phosphorylation was maintained in a condition of cell suspension for 2, 4, and 6 hours, whereas it was completely lost after 12 and 24 hours. We examined MAP kinase activity at 2 and 24 hours, corresponding to the higher and lower levels of FAK phosphorylation. In these experiments, MAP kinase activity correlated with FAK phosphorylation. Stimulation with PDGF was able to cause Ras-GTP loading only in adherent cells. The ability of PDGF to induce phosphatidylinositol 3-kinase (PI 3-K) activity was abrogated in cells maintained in suspension. The Ser473 phosphorylation of Akt was only marginally affected by the lack of cell adhesion. We then evaluated the association of FAK with c-Src. This association was found to be cell adhesion dependent, and it did not appear to be dependent from phosphorylated FAK. These changes in PDGF-induced intracellular signaling were associated with a remarkable reduction of PDGF-proliferative potential in nonadherent cells, although no marked differences in the apoptotic rate were observed. In conclusion, these results suggest that cell adhesion differentially regulates major signaling pathways activated by PDGF in HSC.
生长因子的生物学效应取决于细胞黏附，并且生长因子与黏附复合物之间会发生相互作用。本研究的目的是评估细胞黏附对肝星状细胞（HSC）中血小板衍生生长因子（PDGF）引发的主要细胞内信号通路的影响。在缺乏细胞黏附的不同时间段的实验条件下研究了PDGF信号传导。在细胞悬浮状态下2、4和6小时，基础和PDGF诱导的粘着斑激酶（FAK）酪氨酸磷酸化得以维持，而在12和24小时后则完全丧失。我们在2小时和24小时检测了MAP激酶活性，分别对应于FAK磷酸化的较高和较低水平。在这些实验中，MAP激酶活性与FAK磷酸化相关。仅在贴壁细胞中，PDGF刺激能够导致Ras-GTP负载。在悬浮培养的细胞中，PDGF诱导磷脂酰肌醇3激酶（PI 3-K）活性的能力被消除。细胞黏附缺乏对Akt的Ser473磷酸化仅有轻微影响。然后我们评估了FAK与c-Src的结合。发现这种结合依赖于细胞黏附，并且似乎不依赖于磷酸化的FAK。PDGF诱导的细胞内信号传导的这些变化与非贴壁细胞中PDGF增殖潜能的显著降低相关，尽管未观察到凋亡率有明显差异。总之，这些结果表明细胞黏附以不同方式调节HSC中由PDGF激活的主要信号通路。
