Fig. 5.
Effects of targeting autophagy in GBM on NK cell function and homing. (A) In vitro proliferation of GBM43 WT and BECN1− GBM43 cells (n = 3 or 5). (B) Schematic of the experimental design. (C) Tumor growth was monitored and recorded on the indicated days. IHC staining for (D and E) NK cells (Top), (F and G) CCL5 (Middle), and (H and I) CXCL10 (Bottom) on WT and BECN1- tumors. (Scale bar: 50 μm; 200× magnification.) (J) Viability of GBM43 cells after treatment with various concentrations of CQ for 24 h (n = 4). (K) IC50 value of CQ against GBM43 cells. (L and M) Expression of CCL5 and CXCL10 mRNA in CQ-treated GBM43 cells (n = 4). (N and O) Quantification of CCL5 and CXCL10 in the supernatant of GBM43 cells following treatment with CQ for 24 h in the presence or absence of various inhibitors (n = 3). (P) Schematic diagram illustrating the in vivo CQ treatment program. (Q) Tumor growth of individual mice in each group (n = 4). (R) IHC staining for CCL5 (Left) and CXCL10 (Right) performed on representative tumor sections from each group. (Scale bar: 50 μm; 200× magnification.) (S) IHC scores of CCL5 and CXCL10 in control and CQ-treated tumors. (T) Schematic of in vitro experimental direct contact bilayer blood–brain barrier (BBB) model setup. (U) Transmigration of pNK cells through experimental BBB (n = 3). (V) Correlation between normalized expression of selected genes. Pearson's correlation coefficients are shown with continuous gradient colors. (W) Correlation between normalized expression of the entire NK gene set and individual genes. Correlation expressed as NES. Data shown represent independent samples. Data are shown as mean ± SEM, *P < 0.05, **P < 0.01. P values in N, O, and U were determined using one-way ANOVA analysis and in E, G, I, L, M, Q, and S using the two-tailed Student’s t test.