Fig. 4: O + C induced ICD relies on eIF2α phosphorylation.

A LLC cells were treated with OGD for 7.5 hours, followed by 5 nM of ISRIB for 30 minutes before reperfusion, during reperfusion, 5 nM of ISRIB together with 100 μM of CDDP were added for another 30 min, then cells were harvested, and proteins were detected by western blot. B LLC cells were treated with OGD for 7.5 hours, followed by 5 nM of ISRIB for 30 minutes before reperfusion, during reperfusion, 5 nM of ISRIB together with 100 μM of CDDP were added for 6 h, then cells were harvested and the CRT exposure were tested by flow cytometry, n = 6 biological replicates, the quantification of combines data from three independent experiments. C Western blot of EIF2AK3 knockdown LLC cells. D EIF2AK3 knockdown LLC cells were treated with CDDP, OGD 5 h/R or O + C at the indicated dose for 6 hours, followed by the cytofluorometric detection of ecto-CRT, n = 4 biological replicates, three independent experiments were repeated. E PBS (n = 12) and LLC cells treated with O + C (n = 13) and O + C + I (n = 12) for 20 h and incubated for 4 h without the chemotherapeutic drugs were inoculated into immunocompetent C57BL/6 mice, which were rechallenged 7 days later with live LLC cells. The tumor growth curves and tumor free survival of C57BL/6 mice after injected with different tumor vaccines are shown. Data are reported as the mean ± SEM, and statistical analyses were performed with two-tailed unpaired Student’s t test D and one-way ANOVA followed by the Dunnett post hoc test B. The tumor incidence in the vaccination experiments was analyzed by means of the log-rank test. *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001.