Differential Network Analysis Reveals the Key Role of the ECM-receptor Pathway in a-Particle-induced Malignant Transformation

Space particle radiation is a major environmental factor in spaceflight, and it is known to cause body damage and even trigger cancer, but with unknown molecular etiologies. To examine these causes, we developed a systems biology analytical approach that included co-expression network analysis of transcriptomics profiles obtained using different a-particle radiation exposures of BEAS-2B human bronchial epithelial cells. These exposures employed either single high-dose irradiation of 0.5 Gy one time, or, multiple low-dose irradiations consisting of 25 exposures to 0.02 Gy once every three days. These exposures were then followed by differential network comparative analysis through the multiscale interactomes. Overall pathway analysis based on the global network and the core modules showed that genes in the multiple low-dose irradiation groups had higher significant enrichment for the extracellular matrix (ECM)-receptor interaction pathway. Then, collagen gene COL1A1 was screened as an important ECM-related gene in the multiple low-dose irradiation group assessed by network robustness parameters, and an expression study of lung adenocarcinoma samples from The Cancer Genome Atlas (TCGA) database and additional biological experiments was performed. COL1A1 was found to promote the emergence of the neoplastic characteristics of BEAS-2B cells by both in vitro experimental analyses and in vivo immunohistochemical staining. These findings suggested that the degree of malignant transformation of cells exposed to multiple low-dose irradiations was greater than that of a single high-dose irradiation, which may be caused by the dysregulation of the ECM-receptor pathway. This study provides information on the molecular mechanisms underlying multiple low-dose irradiations used to mimic spaceflight, with a combination of high-throughput RNA-seq-based transcriptomics and systems biology approaches. Overall design: The a-irradiator that we used in this study consisted of an Am source, a rotating radiation source holder, a sample holder and other necessary parts. The Am source emitted a-particles at a dose rate of 0.14 Gy/min. The BEAS-2B cells were irradiated either with a single dose of 0.5 Gy one time or were irradiated with 0.02 Gy (9 seconds of irradiation time) once every three days for 25 exposures. Meanwhile, a blank control was set up, which was sub-cultured together with the irradiated group. Thus, the three groups of cells were labeled as the Ctrl (blank control) group, the single exposure (SE) group (0.5 Gy) and the multiple exposures (ME) group (25 single-dose exposures of 0.02 Gy). After all exposures were completed, the cells were continuously sub-cultured to the 10th passage (approximately 3 weeks), 30th passage (approximately 9 weeks) or 50th passage (approximately 15 weeks), and harvested for RNA-seq at various passages. Thus, the SE groups included R1_05_10, R1_05_30 and R1_05_50, and the ME group included R25_05_10, R25_05_30 and R25_05_50. In summary, this project includes a total of 21 sets of samples.