Abstract
BackgroundBone marrow stromal cells (BMSCs) are extensively used in regeneration therapy and cytology experiments simulate how BMSCs respond to radiation. Due to the small number and the heterogeneity of primary isolated BMSCs, extensive in vitro expansion is usually required before application, which affects the cellular characteristics and gene expression of BMSCs. However, whether the radiation response of BMSCs changes during in vitro expansion is unclear.MethodsIn this study, BMSCs were passaged in vitro and irradiated at passage 6 (P6) and passage 10 (P10).Then, apoptosis, the cell cycle, senescence, the cytokine secretion and the gene expression profile were analysed for the P6, P10, and non-irradiated (control) BMSCs at different post-irradiation time points.ResultsThe P6 BMSCs had a lower percentage of apoptotic cells than the P10 BMSCs at 24 and 48 h post-irradiation but not compared to that of the controls at 2 and 8 h post-irradiation.The P6 BMSCs had a lower percentage of cells in S phase and a higher percentage in G1 phase than the P10 BMSCs at 2 and 8 h post-irradiation. The radiation had similar effects on the senescent cell level and impaired immunomodulation capacity of the P6 and P10 BMSCs. Regardless of whether they were irradiated, the P6 and P10 BMSCs always expressed a distinctive set of genes. The upregulated genes were enriched in pathways including the cell cycle, DNA replication and oocyte meiosis.Then, a subset of conserved irradiation response genes across the BMSCs was identified, comprising 12 differentially upregulated genes and 5 differentially downregulated genes.These genes were especially associated with the p53 signaling pathway, DNA damage and DNA repair. Furthermore, validation experiments revealed that the mRNA and protein levels of these conserved genes were different between the P6 and P10 BMSCs after irradiation. Weighted gene co-expression network analysis supported these findings and further revealed the effects of cell passage on the irradiation response in BMSCs.ConclusionThe results indicated that cell passage in vitro affected the irradiation response of BMSCs via molecular mechanisms that mediated differences in apoptosis, the cell cycle, senescence and the cytokine secretion. Thus, accurate cell passage information is not only important for transplantation therapy but also for future studies on the radiation response in BMSCs.
Highlights
Bone marrow stromal cells (BMSCs) are extensively used in regeneration therapy and cytology experiments simulate how Bone marrow mesenchymal stem cell (BMSC) respond to radiation
The results showed the BMSCs-positive markers including the CD73, CD90 and CD105 were highly expressed both in the passage 6 (P6) and passage 10 (P10) BMSCs
The results demonstrated that both the P6 and P10 BMSCs could differentiate into adipocytes, osteoblasts and chondrocytes
Summary
Bone marrow stromal cells (BMSCs) are extensively used in regeneration therapy and cytology experiments simulate how BMSCs respond to radiation. Ionizing radiation can directly affect the cell by DNA oxidation and breakdown of double strands, which induce damage and variation of the chromosome [2]. The water in the cell would be ionized to generate many reactive oxygen species (ROS), which will indirectly affect the cell by oxidizing the cell membrane. These adverse effects will kill the cell or induce cell canceration [2]. Exposure to a relatively high dose of ionizing radiation will induce an acute response, including severe haematopoietic, gastrointestinal and cerebrovascular syndromes [3]. Understanding the molecular mechanisms mediating the radiation response and resistance is urgent
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