Hecquet Christiane, Lefevre Gaëlle, Valtink Monika, Engelmann Katrin, Mascarelli Frederic
Cordeliers Biomedical Institute, National Institute of Health and Medical Research, Unit 450, National Center for Scientific Research, Paris, France.
Invest Ophthalmol Vis Sci. 2003 Mar;44(3):1320-9. doi: 10.1167/iovs.02-0519.
Retinal pigment epithelial (RPE) cell death is an important step in the pathogenesis of ocular diseases. JNK1 and P38 kinase, two stress-activated kinases, play key roles relaying stress signals leading to cell death through cyclin D1 and c-Myc. Recently, stress-activated kinases have been shown to regulate cell proliferation. In the current study, the involvement of the JNK1 and P38 kinase signaling pathways in RPE cell proliferation and death was investigated.
RPE cell proliferation was stimulated with 10% fetal calf serum (FCS). Activation of the JNK1 and P38 kinase cascades and their potential targets was detected by Western blot analysis. Pharmacologic inhibitors and activators, and antisense oligodeoxynucleotides (ODN) directed against the stress kinases were used to analyze the signaling involved in RPE cell death.
P38 and JNK1 and their respective upstream activating kinases, MKK3/6 and -4, were all transiently activated in FCS-stimulated RPE cell cultures. Ras controlled only the activation of JNK1, whereas Rho transmitted the activation of both JNK1 and P38, suggesting parallel signaling pathways and cross talk between the two kinases. Pharmacologic inhibition of JNK1 did not affect cell proliferation in FCS-stimulated cells. Inactivation of P38 kinase and antisense ODN-induced downregulation of P38 kinase also had no affect on cell proliferation. Long-term, high-level activation of JNK1 and P38 kinase occurred during serum depletion-induced RPE cell death. Overactivation of JNK1 and P38 kinase was also observed during pharmacologically induced cell death, suggesting that this process is common to RPE cell-death-signaling pathways induced by various stress stimuli. Cell death mediated by the overactivation of JNK1 and P38 kinase was cyclin D1- and c-Myc-independent.
The inhibition of JNK1 or P38 kinase had no effect on FCS-stimulated proliferation of RPE cells, whereas the overactivation of these two enzymes was involved in RPE cell death in FCS-depleted cultures. Parallel upstream signaling pathways and cross talk between the two kinases suggest that the regulation of signaling in RPE cell death is complex.
视网膜色素上皮（RPE）细胞死亡是眼部疾病发病机制中的重要环节。JNK1和P38激酶这两种应激激活激酶，在通过细胞周期蛋白D1和c-Myc传递导致细胞死亡的应激信号过程中发挥关键作用。近来，应激激活激酶已被证明可调节细胞增殖。在本研究中，对JNK1和P38激酶信号通路在RPE细胞增殖和死亡中的作用进行了研究。
用10%胎牛血清（FCS）刺激RPE细胞增殖。通过蛋白质印迹分析检测JNK1和P38激酶级联反应及其潜在靶点的激活情况。使用药理抑制剂和激活剂以及针对应激激酶的反义寡脱氧核苷酸（ODN）来分析RPE细胞死亡中涉及的信号传导。
在FCS刺激的RPE细胞培养物中，P38和JNK1及其各自的上游激活激酶MKK3/6和-4均被短暂激活。Ras仅控制JNK1的激活，而Rho传递JNK1和P38的激活，提示存在平行信号通路以及两种激酶之间的相互作用。对JNK1的药理抑制不影响FCS刺激细胞中的细胞增殖。P38激酶的失活以及反义ODN诱导的P38激酶下调对细胞增殖也无影响。在血清剥夺诱导的RPE细胞死亡过程中，JNK1和P38激酶出现长期、高水平的激活。在药理诱导的细胞死亡过程中也观察到JNK1和P38激酶的过度激活，提示该过程在由各种应激刺激诱导的RPE细胞死亡信号通路中是常见的。由JNK1和P38激酶过度激活介导的细胞死亡不依赖于细胞周期蛋白D1和c-Myc。
抑制JNK1或P38激酶对FCS刺激的RPE细胞增殖无影响，而这两种酶的过度激活参与了FCS缺乏培养物中RPE细胞的死亡。平行的上游信号通路以及两种激酶之间的相互作用表明，RPE细胞死亡中的信号调节是复杂的。
