Beijing Laboratory of Urban and Rural Ecological Environment, Beijing Municipal Education Commission, Beijing Key Laboratory of Ornamental Plants Germplasm Innovation and Molecular Breeding, National Engineering Research Center for Floriculture, College of Landscape Architecture, Beijing Forestry University, Beijing, 100083, China.
Plant Cell Rep. 2020 Jul;39(7):941-952. doi: 10.1007/s00299-020-02540-0. Epub 2020 Apr 15.
After cryopreservation, the occurrence of apoptosis-like programmed cell death events induced by the accumulation of ROS reduces pollen viability. Cryopreservation, as a biotechnological means for long-term preservation of pollen, has been applied to many species. However, after cryopreservation, the viability of pollen significantly decreases via a mechanism that is not completely clear. In this study, the pollen of Paeonia lactiflora 'Zi Feng Chao Yang', which exhibits significantly reduced viability after liquid nitrogen (LN) storage, was used to study the relationship among pollen viability, programmed cell death (PCD) and reactive oxygen species (ROS). The apoptosis rate was increased significantly in pollen with decreased viability after cryopreservation, and the changes in ROS generation and hydrogen peroxide (HO) were consistent with the apoptosis rate. Correlation analysis results showed that the apoptosis rate is positively correlated with ROS generation and HO content. In addition, ascorbic acid (AsA), glutathione (GSH) and ascorbic acid reductase (APX) levels were significantly correlated with ROS and HO. After LN preservation for 8 months, the exogenous antioxidants AsA and GSH at appropriate concentrations significantly decreased HO content, inhibited PCD indicator levels, and increased cryopreserved pollen viability. These observations suggest that PCD occurred in pollen during LN preservation for 1-8 months and was induced by the accumulation of ROS in pollen after cryopreservation, thus explaining the main reasons for the reduction in pollen viability after cryopreservation in LN.
经过冷冻保存后，ROS 积累引起的凋亡样程序性细胞死亡事件的发生降低了花粉活力。冷冻保存作为花粉长期保存的生物技术手段已应用于许多物种。然而，经过冷冻保存后，花粉的活力通过一种不完全清楚的机制显著降低。在这项研究中，使用经过液氮（LN）储存后活力显著降低的芍药花粉‘紫凤朝阳’来研究花粉活力、程序性细胞死亡（PCD）和活性氧（ROS）之间的关系。冷冻保存后花粉活力降低，凋亡率显著增加，ROS 生成和过氧化氢（HO）的变化与凋亡率一致。相关分析结果表明，凋亡率与 ROS 生成和 HO 含量呈正相关。此外，抗坏血酸（AsA）、谷胱甘肽（GSH）和抗坏血酸过氧化物酶（APX）水平与 ROS 和 HO 显著相关。经过 LN 保存 8 个月后，合适浓度的外源抗氧化剂 AsA 和 GSH 显著降低了 HO 含量，抑制了 PCD 指标水平，并提高了冷冻保存花粉的活力。这些观察结果表明，PCD 发生在 LN 保存的 1-8 个月期间的花粉中，并且是由冷冻保存后花粉中 ROS 的积累诱导的，这解释了 LN 冷冻保存后花粉活力降低的主要原因。
