| 徐颖,胡文涛,周光明.模拟空间辐射的质子照射促进旁观者肺上皮细胞恶性转化[J].中华放射医学与防护杂志,2025,45(4):282-289.Xu Ying,Hu Wentao,Zhou Guangming.The malignant transformation of bystander lung epithelial cells induced by proton irradiation simulating space radiation[J].Chin J Radiol Med Prot,2025,45(4):282-289 |
| 模拟空间辐射的质子照射促进旁观者肺上皮细胞恶性转化 |
| The malignant transformation of bystander lung epithelial cells induced by proton irradiation simulating space radiation |
| 投稿时间：2024-09-18 |
| DOI：10.3760/cma.j.cn112271-20240918-00355 |
| 中文关键词: 空间辐射 转化生长因子β1 质子照射 旁观者细胞 β-arrestin1 恶性转化 |
| 英文关键词:Space radiation TGF-β1 Proton radiation Bystander cell β-arrestin1 Malignant transformation |
| 基金项目:国家自然科学基金(32071243) |
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| 中文摘要: |
| 目的 探讨模拟空间辐射的质子照射后转化生长因子β1(TGF-β1)对旁观者细胞恶性转化的影响及其机制。方法 对人正常支气管上皮细胞BEAS-2B进行质子照射0、0.2、0.5和1.0 Gy,以模拟空间辐射。收集细胞培养基上清作为条件培养基(CM)用于处理旁观者细胞BEAS-2B,应用酶联免疫吸附试验(ELISA)检测上清培养基中TGF-β1的含量,对旁观者细胞进行软琼脂克隆形成实验,检测细胞恶性转化率。细胞免疫荧光和蛋白免疫印迹实验检测CM处理的旁观者细胞及结合TGF-β1受体抑制剂SB525334后β-arrestin1的定位情况。通过软琼脂克隆形成检测CM联合TGF-β1受体抑制剂或敲低β-arrestin1后细胞的恶性转化情况。实时定量PCR和蛋白免疫印迹实验分别检测上皮-间质转化(EMT)相关基因E-cadherin、N-cadherin、Fibronectin1及Vimentin的mRNA水平和蛋白水平。结果 与0 Gy组相比,随照射剂量增加,24 h后细胞分泌的TGF-β1的含量逐渐增多(t=3.38、8.32、10.96,P＜0.05),CM处理的旁观者细胞的克隆形成率也逐渐增多(t=5.04、7.20、10.78,P＜0.05)。通过细胞免疫荧光和蛋白免疫印迹实验发现,CM处理的旁观者细胞随剂量的增加,β-arrestin1入核增多(t=7.57、7.51,P＜0.05),其入核受到TGF-β1刺激和SB525334的抑制。应用SB525334或敲低β-arrestin1能够抑制质子照射导致的旁观者细胞恶性转化和EMT,减少1 Gy质子照射所得CM处理的旁细胞克隆形成率(t=2.84、3.39,P＜0.05),提高E-cadherin基因的mRNA水平(t=7.33、5.38,P<0.05)和蛋白水平,降低N-cadherin、Fibronectin1、Vimentin基因的mRNA(t=4.37、4.10、5.29、10.65、5.15、3.11,P<0.05)和蛋白水平。结论 模拟空间辐射的质子照射能促进肺上皮细胞TGF-β1的分泌,从而诱导旁观者细胞β-arrestin1的入核,促进细胞发生恶性转化,TGF-β1/β-arrestin1通路在其中发挥重要作用。 |
| 英文摘要: |
| Objective To investigate the influence of TGF-β1 on the malignant transformation of bystander cells after proton irradiation simulating space radiation, and its underlying mechanism. Methods Normal human bronchial epithelial cells BEAS-2B were exposed to proton irradiation at 0, 0.2, 0.5, and 1.0 Gy to simulate space radiation. Supernatants from cell culture media were collected as a conditioned medium (CM) for treating bystander BEAS-2B cells. The enzyme-linked immunosorbent assay (ELISA) was employed to detect TGF-β1 levels within the CM. The soft agar colony formation assay was performed to assess the rate of malignant transformation of bystander cells. Immunofluorescence and Western blot techniques were utilized to examine the localization of β-arrestin1 in CM-treated bystander cells, with or without the TGF-β1 receptor inhibitor SB525334. The malignant transformation of bystander cells was assessed via soft agar colony formation assay under CM treatment, combined with either a TGF-β1 receptor inhibitor or β-arrestin1 knockdown. Additionally, mRNA and protein levels of epithelial-mesenchymal transition(EMT)-related genes (e.g., E-cadherin, N-cadherin, Fibronectin1, and Vimentin) were analyzed through qRT-PCR and Western blot, respectively. Results Contrasting with the 0 Gy group, the proton irradiation groups exhibited a dose-dependent increase in TGF-β1 secretion after 24 h (t=3.38, 8.32, 10.96, P<0.05), and a corresponding rise in the soft agar colony formation rate of CM-treated bystander cells (t=5.04, 7.20, 10.78, P<0.05). Immunofluorescence and Western blot results indicated that with escalating doses, CM-treated bystander cells showed increased β-arrestin1 into nuclei (t=7.57, 7.51, P<0.05), being stimulated by TGF-β1 and inhibited by SB525334. The SB525334 application or β-arrestin1 knockdown significantly inhibited the malignant transformation and EMT induced by proton irradiation in bystander cells. This inhibition further reduced the soft agar colony formation rate (t=2.84, 3.39, P<0.05), and increased mRNA and protein levels of the E-cadherin gene in CM-treated bystander cells exposed to 1 Gy proton irradiation (t=7.33, 5.38, P<0.05) while reducing the mRNA and protein levels of N-cadherin, Fibronectin1, and Vimentin genes (t=4.37, 4.10, 5.29, 10.65, 5.15, 3.11, P<0.05). Conclusions Proton irradiation simulating space radiation can enhance TGF-β1 secretion from lung epithelial cells, inducing β-arrestin1 into nuclei in bystander cells, thereby spurring the malignant transformation of cells. The TGF-β1/β-arrestin1 pathway plays a crucial role in this process. |
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