Figure 1.

Autophagic flux of TILs, especially exhausted TILs, is significantly impaired. (A, B) Peripheral T cells (PTCs) and tumor-infiltrating lymphocytes (TILs) from donors treated with or without chloroquine (CQ) (50 µM; 12 hours) and LC3-II were detected by western blotting. Representative results from three PTCs and TILs were shown (A). Quantitative analysis was of the LC3-II bands was shown (n=5) (B). (C, D) Representative confocal images captured from PTCs and TILs treated with or without CQ were shown (C). The mean fluorescent intensity (MFI) of each image from different groups was statistically analyzed (n=6) (D). We used the Image J to measure the mean value for each view. To details, we chose six views for each group in ×20 magnification. And at least 12 cells determined by DAPI staining for each view, and then Image J was used to calculate the average intensity of LC3 fluorescence for all the cells in each view. (E, F) Representative flow cytometric results of LC3-II from PTCs and TILs treated with or without CQ were shown (E). The fold increases in LC3-II expression levels in PTCs and TILs after CQ treatment were summarized and shown (n=5) (F). (G, H) Representative flow cytometric results for LC3-II in PD1/TIM3/LAG3+TILs and PD1/TIM3/LAG3- TILs with or without CQ treatment were shown (G). The fold increases in LC3-II expression levels in PD1/TIM3/LAG3+TILs and PD1/TIM3/LAG3- TILs after CQ treatment were summarized and shown (n=6) (H). (I, J) Representative flow cytometric results for LC3-II in Ki67+TILs and Ki67- TILs with or without CQ treatment were shown (I). The fold increases in LC3-II expression levels in Ki67+TILs and Ki67- TILs after CQ treatment were summarized and shown (n=4) (J). (K, L) Representative flow cytometric results for LC3-II in IFN-γ+ TILs and IFN-γ- TILs with or without CQ treatment were shown (K). The fold increases in LC3-II expression levels in IFN-γ+ TILs and IFN-γ- TILs after CQ treatment were summarized and shown (n=4) (L).