Department of Nephrology, Shanghai East Hospital, Tongji University School of Medicine, Shanghai, China.
J Cell Physiol. 2012 May;227(5):2138-44. doi: 10.1002/jcp.22946.
Dedifferentiation, a process by which differentiated cells become mesenchymal-like proliferating cells, is the first step in renal epithelium repair and occurs in vivo after acute kidney injury and in vitro in primary culture. However, the underlying mechanism remains poorly understood. In this report, we studied the signaling events that mediate dedifferentiation of proximal renal tubular cells (RPTC) in primary culture. RPTC dedifferentiation characterized by increased expression of vimentin concurrent with decreased expression of cytokeratin-18 was observed at 24 h after the initial plating of freshly isolated proximal tubules and persisted for 72 h. At 96 h, RPTC started to redifferentiate as revealed by reciprocal expression of cytokeratin-18 and vimentin and completed at 120 h. Phosphorylation levels of Src, epidermal growth factor receptor (EGFR), AKT (a target of phosphoinositide-3-kinase (PI3K)), and ERK1/2 were increased in the early time course of culture (<72 h). Inhibition of Src family kinases (SFKs) with PP1 blocked EGFR, AKT, and ERK1/2 phosphorylation, as well as RPTC dedifferentiation. Inhibition of EGFR with AG1478 also blocked AKT and ERK1/2 phosphorylation and RPTC dedifferentiation. Although inactivation of the PI3K/AKT pathway with LY294002 inhibited RPTC dedifferentiation, blocking the ERK1/2 pathway with U0126 did not show such an effect. Moreover, inhibition of SFKs, EGFR, PI3K/AKT, but not ERK1/2 pathways abrogated RPTC outgrowth and SFK inhibition decreased RPTC proliferation and migration. These findings demonstrate a critical role of SFKs in mediating RPTC dedifferentiation through activation of the EGFR/PI3K signaling pathway.
去分化，即分化细胞变成间充质样增殖细胞的过程，是肾上皮修复的第一步，发生在急性肾损伤后的体内和原代培养中的体外。然而，其潜在机制仍知之甚少。在本报告中，我们研究了介导原代培养中近端肾小管细胞（RPTC）去分化的信号事件。在新鲜分离的近端小管最初接种后 24 小时观察到 RPTC 去分化的特征，即波形蛋白表达增加，同时细胞角蛋白-18 表达减少，并持续 72 小时。96 小时时，RPTC 开始向相反的细胞角蛋白-18 和波形蛋白表达方向重新分化，并在 120 小时时完成。培养早期（<72 小时），Src、表皮生长因子受体（EGFR）、AKT（磷酸肌醇-3-激酶（PI3K）的靶标）和 ERK1/2 的磷酸化水平增加。用 PP1 抑制 Src 家族激酶（SFKs）可阻断 EGFR、AKT 和 ERK1/2 的磷酸化以及 RPTC 去分化。用 AG1478 抑制 EGFR 也可阻断 AKT 和 ERK1/2 的磷酸化和 RPTC 去分化。虽然用 LY294002 使 PI3K/AKT 通路失活可抑制 RPTC 去分化，但用 U0126 阻断 ERK1/2 通路则无此效果。此外，抑制 SFKs、EGFR、PI3K/AKT，但不是 ERK1/2 通路可消除 RPTC 生长，SFK 抑制可减少 RPTC 增殖和迁移。这些发现表明 SFKs 通过激活 EGFR/PI3K 信号通路在介导 RPTC 去分化中起关键作用。
