Effects of low-dose rate γ-irradiation combined with simulated microgravity on markers of oxidative stress, DNA methylation potential, and remodeling in the mouse heart

PurposeSpace travel is associated with an exposure to low-dose rate ionizing radiation and the microgravity environment, both of which may lead to impairments in cardiac function. We used a mouse model to determine short- and long-term cardiac effects to simulated microgravity (hindlimb unloading; HU), continuous low-dose rate γ-irradiation, or a combination of HU and low-dose rate γ-irradiation.MethodsCardiac tissue was obtained from female, C57BL/6J mice 7 days, 1 month, 4 months, and 9 months following the completion of a 21 day exposure to HU or a 21 day exposure to low-dose rate γ-irradiation (average dose rate of 0.01 cGy/h to a total of 0.04 Gy), or a 21 day simultaneous exposure to HU and low-dose rate γ-irradiation. Immunoblot analysis, rt-PCR, high-performance liquid chromatography, and histology were used to assess inflammatory cell infiltration, cardiac remodeling, oxidative stress, and the methylation potential of cardiac tissue in 3 to 6 animals per group.ResultsThe combination of HU and γ-irradiation demonstrated the strongest increase in reduced to oxidized glutathione ratios 7 days and 1 month after treatment, but a difference was no longer apparent after 9 months. On the other hand, no significant changes in 4-hydroxynonenal adducts was seen in any of the groups, at the measured endpoints. While manganese superoxide dismutase protein levels decreased 9 months after low-dose γ-radiation, no changes were observed in expression of catalase or Nrf2, a transcription factor that determines the expression of several antioxidant enzymes, at the measured endpoints. Inflammatory marker, CD-2 protein content was significantly decreased in all groups 4 months after treatment. No significant differences were observed in α-smooth muscle cell actin protein content, collagen type III protein content or % total collagen.ConclusionsThis study has provided the first and relatively broad analysis of small molecule and protein markers of oxidative stress, T-lymphocyte infiltration, and cardiac remodeling in response to HU with simultaneous exposure to low-dose rate γ-radiation. Results from the late observation time points suggest that the hearts had mostly recovered from these two experimental conditions. However, further research is needed with larger numbers of animals for a more robust statistical power to fully characterize the early and late effects of simulated microgravity combined with exposure to low-dose rate ionizing radiation on the heart.

【研究背景与目的】太空旅行会使机体暴露于低剂量率电离辐射（low-dose rate ionizing radiation）与微重力环境中，二者均可能导致心功能受损。本研究采用小鼠模型，探究模拟微重力（后肢悬吊法；hindlimb unloading, HU）、持续低剂量率γ射线辐照，以及二者联合暴露对心脏产生的短期与长期影响。
【实验方法】本研究选取雌性C57BL/6J小鼠，分别在21天暴露周期结束后的7天、1个月、4个月及9个月采集心脏组织；暴露方案包括：单独21天后肢悬吊（HU）、单独21天低剂量率γ射线辐照（平均剂量率0.01 cGy/h，总剂量0.04 Gy），以及21天同时联合HU与低剂量率γ射线辐照。每组设置3~6只小鼠，采用免疫印迹分析、逆转录PCR（rt-PCR）、高效液相色谱及组织学技术，检测各组心脏组织的炎症细胞浸润、心脏重构、氧化应激及甲基化潜能水平。
【实验结果】HU与γ射线辐照联合暴露组，在处理后7天及1个月时的还原型/氧化型谷胱甘肽比值升高最为显著，但该差异在9个月后不再显著。另一方面，所有组别在各检测时间点的4-羟基壬烯醛加合物水平均无显著变化。低剂量γ射线辐照组小鼠的锰超氧化物歧化酶蛋白水平在9个月时出现下降，但各检测时间点的过氧化氢酶及核因子E2相关因子2（Nrf2，一种调控多种抗氧化酶表达的转录因子）的表达水平均无明显改变。处理后4个月时，所有组别小鼠的炎症标志物CD-2蛋白含量均显著降低。各组的α平滑肌肌动蛋白、III型胶原蛋白含量及总胶原占比均无显著差异。
【研究结论】本研究首次针对模拟微重力联合低剂量率γ射线辐照暴露后的氧化应激、T淋巴细胞浸润及心脏重构相关小分子与蛋白标志物开展了较为全面的分析。晚期观测时间点的结果显示，心脏在这两种实验暴露条件下大多已恢复正常。不过，后续仍需采用更多实验动物以获得更可靠的统计效力，从而全面阐明模拟微重力联合低剂量率电离辐射暴露对心脏产生的早期与晚期影响。