Department of Otolaryngology-Head and Neck Surgery, University of Iowa, Iowa City, Iowa 52242, USA.
Neurosurgery. 2013 Sep;73(3):506-16. doi: 10.1227/01.neu.0000431483.10031.89.
Radiosurgery is increasingly used to treat vestibular schwannomas (VSs). Increasing the sensitivity of VS cells to irradiation (IR) could allow for lower and/or more effective doses of IR, improving safety and efficacy. Persistent c-Jun N-terminal kinase (JNK) activity in VS cells reduces cell death by suppressing the accumulation of reactive oxygen species (ROS), raising the possibility that JNK activity protects against IR-induced VS cell death, which is mediated by ROS.
To determine the extent to which JNK signaling contributes to VS cell radiosensitivity.
Primary human VS cultures, derived from acutely resected tumors, received single doses (5-40 Gy) of gamma irradiation. Histone 2AX phosphorylation, a marker of IR-induced DNA damage, was assayed by Western blot and immunostaining. ROS levels were quantified by measuring 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) fluorescence. Cell apoptosis was determined by terminal deoxynucleotidyl transferase 2'-deoxyuridine, 5'-triphosphate nick end labeling.
The JNK inhibitors SP6000125 and I-JIP reduced histone 2AX phosphorylation after IR. They also increased H2DCFDA fluorescence in nonirradiated cultures and significantly increased IR-induced (5-10 Gy) H2DCFDA fluorescence 72 hours, but not 2 hours, after IR. Finally, I-JIP (50 μmol/L) significantly increased VS cell apoptosis in cultures treated with 20 to 40 Gy. I-JIP (20 μmol/L), SP600125 (20 μmol/L), and JNK1/2 short interfering RNA knockdown each increased VS cell apoptosis in cultures treated with 30 to 40 Gy, but not lower doses, of IR.
Inhibition of JNK signaling decreases histone 2AX phosphorylation and increases ROS and apoptosis in VS cells after gamma irradiation. These results raise the possibility of using JNK inhibitors to increase the effectiveness of radiosurgery for treatment of VSs.
放射外科越来越多地用于治疗前庭神经鞘瘤（VSs）。提高 VS 细胞对辐射（IR）的敏感性，可以降低和/或提高 IR 的有效剂量，从而提高安全性和疗效。VS 细胞中持续的 c-Jun N-末端激酶（JNK）活性通过抑制活性氧（ROS）的积累来减少细胞死亡，这增加了 JNK 活性对 ROS 介导的 IR 诱导的 VS 细胞死亡的保护作用。
确定 JNK 信号传导在多大程度上有助于 VS 细胞的放射敏感性。
从急性切除的肿瘤中获得的原代人 VS 培养物接受单次剂量（5-40Gy）的γ照射。通过 Western blot 和免疫染色测定组蛋白 2AX 磷酸化，这是 IR 诱导的 DNA 损伤的标志物。通过测量 2'，7'-二氯二氢荧光素二乙酸酯（H2DCFDA）荧光来量化 ROS 水平。通过末端脱氧核苷酸转移酶 2'-脱氧尿苷，5'-三磷酸核苷酸缺口末端标记法测定细胞凋亡。
JNK 抑制剂 SP6000125 和 I-JIP 降低了 IR 后的组蛋白 2AX 磷酸化。它们还增加了未照射培养物中的 H2DCFDA 荧光，并在 IR 后 72 小时但不是 2 小时显著增加了 IR 诱导的（5-10Gy）H2DCFDA 荧光。最后，I-JIP（50μmol/L）在 20 至 40Gy 处理的培养物中显著增加了 VS 细胞凋亡。I-JIP（20μmol/L）、SP600125（20μmol/L）和 JNK1/2 短发夹 RNA 敲低均增加了 30 至 40Gy 的 IR 处理培养物中的 VS 细胞凋亡，但不会增加较低剂量的 IR 处理培养物中的 VS 细胞凋亡。
JNK 信号抑制降低了伽马照射后 VS 细胞中的组蛋白 2AX 磷酸化并增加了 ROS 和细胞凋亡。这些结果增加了使用 JNK 抑制剂来提高放射外科治疗 VSs 的有效性的可能性。
