鉴定白细胞介素-13 受体α2 在人类多形性胶质母细胞瘤中的新作用：白细胞介素-13 通过 AP-1 途径介导信号转导。

BACKGROUND: Previously, we have demonstrated that Interleukin 13 receptor alpha 2 (IL-13Rα2) is overexpressed in approximate 78% Glioblastoma multiforme (GBM) samples. We have also demonstrated that IL-13Rα2 can serve as a target for cancer immunotherapy in several pre-clinical and clinical studies. However, the significance of overexpression of IL-13Rα2 in GBM and astrocytoma and signaling through these receptors is not known. IL-13 can signal through IL-13R via JAK/STAT and AP-1 pathways in certain cell lines including some tumor cell lines. Herein, we have investigated a role of IL-13/IL-13Rα2 axis in signaling through AP-1 transcription factors in human glioma samples in situ. METHODS: We examined the activation of AP-1 family of transcription factors (c-Jun, Fra-1, Jun-D, c-Fos, and Jun-B) after treating U251, A172 (IL-13Rα2 +ve) and T98G (IL-13Rα2 -ve) glioma cell lines with IL-13 by RT-qPCR, and immunocytochemistry (ICC). We also performed colorimetric ELISA based assay to determine AP-1 transcription factor activation in glioma cell lines. Furthermore, we examined the expression of AP-1 transcription factors in situ in GBM and astrocytoma specimens by multiplex-immunohistochemistry (IHC). Student t test and ANOVA were used for statistical analysis of the results. RESULTS: We have demonstrated up-regulation of two AP-1 transcription factors (c-Jun and Fra-1) at mRNA and protein levels upon treatment with IL-13 in IL-13Rα2 positive but not in IL-13Rα2 negative glioma cell lines. Both transcription factors were also overexpressed in patient derived GBM specimens, however, in contrast to GBM cell lines, c-Fos is also overexpressed in patient derived specimens. Astrocytoma specimens showed lesser extent of immunostaining for IL-13Rα2 and three AP-1 factors compared to GBM specimens. By transcription factor activation assay, we demonstrated that AP-1 transcription factors (C-Jun and Fra-1) were activated upon treatment of IL-13Rα2 + GBM cell lines but not IL-13Rα2 - GBM cell line with IL-13. Our results demonstrate functional activity of AP-1 transcription factor in GBM cell lines in response to IL-13. CONCLUSIONS: These results indicate that IL-13/IL-13Rα2 axis can mediate signal transduction in situ via AP-1 pathway in GBM and astrocytoma and may serve as a new target for GBM immunotherapy.

背景：此前，我们已经证明白细胞介素 13 受体 alpha2（IL-13Rα2）在大约 78%的多形性胶质母细胞瘤（GBM）样本中过度表达。我们还证明，IL-13Rα2 可以在几项临床前和临床研究中作为癌症免疫治疗的靶点。然而，GBM 和星形细胞瘤中 IL-13Rα2 的过度表达及其受体信号的意义尚不清楚。在某些细胞系中，包括一些肿瘤细胞系，IL-13 可以通过 JAK/STAT 和 AP-1 途径通过 IL-13R 信号传导。在此，我们研究了 IL-13/IL-13Rα2 轴在人胶质瘤样本原位通过 AP-1 转录因子信号转导中的作用。

方法：我们通过 RT-qPCR 和免疫细胞化学（ICC）检测用 IL-13 处理 U251、A172（IL-13Rα2+ve）和 T98G（IL-13Rα2-ve）神经胶质瘤细胞系后，AP-1 转录因子家族（c-Jun、Fra-1、Jun-D、c-Fos 和 Jun-B）的激活。我们还进行了基于比色 ELISA 的测定，以确定神经胶质瘤细胞系中 AP-1 转录因子的激活。此外，我们通过多重免疫组织化学（IHC）检测了 GBM 和星形细胞瘤标本中 AP-1 转录因子的表达。使用学生 t 检验和 ANOVA 对结果进行统计学分析。

结果：我们已经证明，在 IL-13Rα2 阳性但不是 IL-13Rα2 阴性神经胶质瘤细胞系中，两种 AP-1 转录因子（c-Jun 和 Fra-1）在 mRNA 和蛋白水平上的上调。在患者来源的 GBM 标本中，这两种转录因子也过表达，但与 GBM 细胞系相反，c-Fos 在患者来源的标本中也过表达。与 GBM 标本相比，星形细胞瘤标本中 IL-13Rα2 和三种 AP-1 因子的免疫染色程度较低。通过转录因子激活测定，我们证明了 IL-13Rα2+GBM 细胞系在接受 IL-13 治疗后，AP-1 转录因子（C-Jun 和 Fra-1）被激活，但 IL-13Rα2-GBM 细胞系未被激活。我们的结果表明，AP-1 转录因子在 GBM 细胞系中对 IL-13 的反应中具有功能活性。

结论：这些结果表明，IL-13/IL-13Rα2 轴可以通过原位 AP-1 途径在 GBM 和星形细胞瘤中介导信号转导，并可能成为 GBM 免疫治疗的新靶点。