Abstract

Purpose

Mitochondrial antiviral signaling (MAVS) protein, located in the mitochondrial out-membrane, is necessary for IFN-beta induction and IFN-stimulated gene expression in response to external stress such as viral invasion and ionizing radiation (IR). Although the involvement of radiation induced bystander effect (RIBE) has been investigated for decades for secondary cancer risk related to radiotherapy, the underlying regulatory mechanisms remain largely unclear, especially the roles played by the immune factors such as MAVS.

Material and methods

MAVS gene knockout cells using CRISPR/Cas9 technology were used as donor cells or recipient cells to assess the role of MAVS in RIBE by means of co-cultured system. The micronucleus and γH2AX foci in the recipient cells were counted to demonstrate the degree of RIBE. The reactive oxygen species (ROS) level in the recipient was measured using the fluorescent dye 2′7′-dichlorofluorescein.

Results

Firstly, we found that MAVS expression level was different in A549, BEAS-2B, U937 and HepG2 cells. Cell co-culture experiments showed that MAVS participate in RIBE. Interestingly, the RIBE response was more significant in recipient cells with higher level of MAVS (i.e. A549) than that in recipient cells showing lower level of MAVS (i.e. BEAS-2B). Further, the bystander response was dramatically suppressed in MAVS-silenced A549 and BEAS-2B recipient cells. MAVS-silenced recipient cells exhibited lower level of ROS induced by IR.

Conclusions

Our results indicated that the innate immune signaling molecule MAVS in recipient cells participate in RIBE. ROS is an important factor in RIBE via MAVS pathway and MAVS may be a potential target for the precise radiotherapy and radioprotection.

中文翻译：

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抑制先天免疫信号分子 MAVS 可减少辐射诱导的旁观者效应。

摘要

目的

线粒体抗病毒信号 (MAVS) 蛋白位于线粒体外膜中，对于 IFN-β 诱导和 IFN 刺激基因表达以响应病毒入侵和电离辐射 (IR) 等外部压力是必需的。尽管辐射诱发的旁观者效应 (RIBE) 参与放射治疗的继发性癌症风险已被研究了数十年，但其潜在的调节机制仍很大程度上不清楚，尤其是 MAVS 等免疫因素所起的作用。

材料与方法

采用CRISPR/Cas9技术的MAVS基因敲除细胞作为供体细胞或受体细胞，通过共培养系统评估MAVS在RIBE中的作用。对受体细胞中的微核和 γH2AX 病灶进行计数以证明 RIBE 的程度。使用荧光染料 2'7'-二氯荧光素测量受体中的活性氧 (ROS) 水平。

结果

首先，我们发现MAVS在A549、BEAS-2B、U937和HepG2细胞中的表达水平不同。细胞共培养实验表明，MAVS 参与了 RIBE。有趣的是，在MAVS 水平较高的受体细胞（即A549）中RIBE 反应比在MAVS 水平较低的受体细胞（即BEAS-2B）中更为显着。此外，在 MAVS 沉默的 A549 和 BEAS-2B 受体细胞中，旁观者反应被显着抑制。MAVS 沉默的受体细胞表现出较低水平的 IR 诱导的 ROS。

结论

我们的结果表明受体细胞中的先天免疫信号分子 MAVS 参与了 RIBE。ROS 是通过 MAVS 途径进行 RIBE 的重要因素，而 MAVS 可能是精确放疗和放射防护的潜在靶点。