Irradiation-Induced Polyploidy Giant Cancer Cells Mediate Tumor Cell Repopulation via Neosis

Background Tumor repopulation generally describes the phenomenon that residual tumor cells surviving therapies tenaciously proliferate and reestablish the tumor, presenting an embarrassing plight for cancer treatment. However, the cellular and molecular mechanisms underlying this process remains poorly understood. In this study, we proposed polyploidy giant cancer cells (PGCCs)-mediated and neosis-based tumor repopulation after radiotherapy.Methods The formation of PGCCs after irradiation was examined in vitro and in vivo. The demise of X-ray irradiated cells was detected by flow cytometry, clonogenic cell survival assay and transmission electron microscopy. Western blot was used to test cell proliferation and death related protein expression level of these irradiated cells. Time lapse microscopy was adopted to observe the destiny of PGCCs. The property of these PGCCs was identified by TUNEL assay, Brdu chasing assay, western blot, immunocytochemical and immunofluorescence staining. The relationship of HMGB1 with PGCCs-derived tumor repopulation was conducted via HMGB1 chemical inhibitors. Finally, animal model was used to verify the formation of PGCCs, and the relevance of HMGB1 in this process was investigated by immunohistochemical staining.Results The majority of PGCCs induced by irradiation move towards cell demise, whereas some of them intriguingly possessed proliferative property. Utilizing time-lapse microscopy and single-cell cloning assay, we observed that neosis derived from those PGCCs with proliferative capacity contributed to tumor cell repopulation after irradiation. Using the conditioned media collected from dying tumor cells to perform single-cell cloning assay, we unexpectedly demonstrated that HMGB1 released from dying tumor cells participated the process of neosis-based tumor repopulation. In irradiation treated animal tumor bearing model, the expression level of HMGB1 increased after irradiation compare with non-irradiated group. Moreover, some PGCCs presented high HMGB1 expression. Interestingly, we also observed that the proliferation potential of PGCCs varied. Some PGCCs proliferated at early stage, while some PGCCs proliferated at late stage.Conclusion X-ray irradiation could induce the formation of PGCCs, which could move towards both cell death and survival; irradiation-generating PGCCs mediated tumor cell repopulation after irradiation via neosis; HMGB1 released from dying cells stimulated the process of neosis and participated in tumor repopulation after irradiation.