Comparison of the effects of photon, proton and carbon-ion radiation on the ecto-calreticulin exposure in various tumor cell lines

Accumulating evidence suggested that radiotherapy can activate anti-tumor immune responses by triggering immunogenic cell death (ICD) of tumor cells. Calreticulin is regarded as one of the most important markers of ICD. The cell surface translocation of calreticulin (ecto-CRT) serves as an "eat me" signal for phagocytosis of dying cells, which plays a pivotal role in activating anti-tumor immunity. However, there is limited knowledge describing the effects of proton and carbon-ion radiation on ecto-CRT exposure. Hence, we investigated ecto-CRT exposure in multiple human carcinoma cell lines irradiated by proton and carbon-ion in comparison to photon. This study examined four human cancer cell lines including A549 (lung adenocarcinoma), U251MG (glioma), Tca8113 (tongue squamous carcinoma), and CNE-2 (nasopharyngeal carcinoma). Cell lines were irradiated with photon, proton or carbon-ion at 0, 2, 4, 10 Gy (physical dose). The ecto-CRT exposure level was analyzed by flow cytometry at 12, 24, and 48 h post-irradiation. The median fluorescence intensity was calculated by FlowJo. All three types of radial beam increased ecto-CRT exposure of the 4 tumor cell lines in a time-dependent manner. Ecto-CRT exposure significantly elevated 1.5-2.4 times over 48 h post-irradiation compared with controls (P