【摘要】 目的 研究12 Gy⁶⁰Coγ射线对睾丸支持细胞（sertoli cells, SCs）铁死亡的影响,并探究益气解毒方干预对支持细胞铁死亡的防护机制。方法 支持细胞分为空白（NC）组,模型（IR）组、铁死亡激动剂（Erastin）组、铁死亡抑制剂（Lip-1）组、益气解毒方高剂量（YQJD-H）组、益气解毒方低剂量（YQJD-L）组,分别按照相应的条件培养,除NC组和Erastin组外,其余各组细胞均使用12 Gy⁶⁰Coγ射线进行一次性照射,建立支持细胞辐射损伤模型。照射后24 h,用细胞计数试剂（cell counting kit-8, CCK-8）检测细胞活性;DCFH-DA荧光探针检测细胞内活性氧（reactive oxygen species, ROS）水平;铁离子比色法检测细胞内Fe²⁺水平;ELISA法检测细胞上清液高迁移率族蛋白B1 （high mobility group protein box-1, HMGB1）水平;JC-1荧光探针检测细胞线粒体膜电位改变;实时荧光定量聚合酶链式反应（real time quantitative polymerase chain reaction,RT-qPCR）检测细胞GPX4、ACSL4、LPCAT3 m RNA的表达。结果 照射后24 h,与NC组比较,Erastin组和IR组支持细胞活性及线粒体膜电位均显著下降（P<0.01）,ROS、Fe²⁺、HMGB1水平均明显升高（P<0.01）,GPX4 m RNA表达水平下降（P<0.05）;IR组ACSL4、LPCAT3 mRNA表达水平升高（P<0.05）。与IR组比较,YQJD-H组、YQJD-L组、Lip-1组的细胞活性均显著升高（P<0.01）,ROS、Fe²⁺、HMGB1水平均明显降低（P<0.01）;YQJD-H组GPX4 m RNA表达水平升高（P<0.05）,ACSL4、LPCAT3 m RNA表达水平下降（P<0.05）。结论 12 Gy⁶⁰Coγ射线诱导支持细胞发生铁死亡,而益气解毒方能减轻支持细胞铁死亡,起到辐射防护作用,其作用机制可能与GPX4/ACSL4/LPCAT3信号通路有关。更多还原

【Abstract】 Objective To study whether 12 Gy⁶⁰Coγ-ray induces ferroptosis of sertoli cells（SCs）, and to investigate the mechanism of Yiqi Jiedu decoction against irradiation-induced ferroptosis of Sertoli cells. Methods Sertoli cells were divided into the negative control（NC） group, ionizing radiation（IR） group, ferroptosis activator（Erastin） group,ferroptosis inhibitor（Lip-1） group, Yiqi Jiedu decoction high dose（YQJD-H）group, Yiqi Jiedu decoction low dose（YQJD-L）group. Sertoli cells in each group were cultured according to the corresponding conditions. Except for NC group and Erastin group, all other groups received a one-time, single dose of 12 Gy⁶⁰Coγ irradiation. 24 h after irradiation, cell viability was detected by CCK8 method. The level of reactive oxygen species（ROS） was detected by DCFH-DA fluorescent probe. The level of Fe2+ was detected by intracellular iron colorimetric method. ELISA was used to detect the level of high mobility group protein box-1（HMGB1） in cell supernatant. JC-1 fluorescent probe was used to detect cellular mitochondrial membrane potential changes. The GPX4, ACSL4 and LPCAT3 gene expression were detected by real time quantitative polymerase chain reaction（RT-qPCR）.Results 24 h after irradiation, compared with NC group, both Erastin and IR groups showed a significant decrease in sertoli cell viability and mitochondrial membrane potential（P<0. 01）, a significant increase in the levels of ROS, Fe2+, and HMGB1（P<0. 01）, and a decrease in the expression of GPX4 m RNA（P<0. 05）; yet the expression of ACSL4 and LPCAT3 m RNA in the IR group went up（P<0. 05）. Compared with the IR group, cell viability was significantly higher in the YQJD-H, YQJD-L, and Lip-1 groups（P<0. 01）, and ROS, Fe2+, and HMGB1 levels were significantly lower（P<0. 01）. GPX4 m RNA expression increased in the YQJD-H group（P<0. 05）,and ACSL4 and LPCAT3 m RNA expression decreased（ P< 0. 05）. Conclusion 12 Gy⁶⁰Coγ-ray induced ferroptosis in sertoli cells. Yiqi Jiedu decoction could alleviate ferroptosis in sertoli cells to reduce the radiation injury on sertoli cells. Its mechanism of action may be related to the GPX4/ACSL4/LPCAT3 signaling pathway.更多还原