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. 2020 Aug 2;23(8):101431. doi: 10.1016/j.isci.2020.101431

Figure 5.

Figure 5

Metabolism and DDR Pathway Alterations in sEVs-T

(A, B, and E) GSEA was conducted to search gene sets and KEGG pathways altered by sEVs stimulation. (A) GSEA showing the alteration of gene sets involved in metabolism in sEVs-T. p value <0.05 and q value <0.25 meaning statistically significant. (B) GSEA showing the alteration of KEGG pathways involved in metabolism in sEVs-T. p value <0.05 and q value <0.25 meaning statistically significant. The ratio of count meaning altered gene/total gene of pathway. (E) GSEA showing the alteration of gene sets involved in DNA damage response in sEVs-T.

(C) Western blot showing the upregulation of metabolic regulator p-AMPK, SIRT1, SIRT2, and SIRT6 in sEVs-T (n = 3). Data shown are mean ± standard deviation. ∗p < 0.05; ∗∗p < 0.01 (two-tailed, unpaired Student's t test).

(D) Line chart reflecting the cytotoxic activity of T lymphocyte through BxPC-3 survival curve. sEVs down regulated the cytotoxic activity of T lymphocytes, and it was rescued by AMPK inhibitor Compound C (CC), SIRT1 inhibitor EX527, SIRT2 inhibitor AGK2, and SIRT6 inhibitor OSS_128167 (OSS). Each point in the graph constituting the curve represented the average of 4 biological replicates.

(F) Immunofluorescence showing the increased foci of γ-H2AX, RAD51, RPA2, and 53BP1 in sEVs-T (n = 3). Data shown are mean ± standard deviation. ∗p < 0.05; ∗∗p < 0.01 (two-tailed, unpaired Student's t test).

(G) Western blot showing the upregulated activation of DDR sensor p-ATM in sEVs-T. The ratio of phosphor-ATM/ATM meaning the degree of ATM activation (n = 3). Data shown are mean ± standard deviation. ∗p < 0.05 (two-tailed, unpaired Student's t test).

(H) Line chart reflecting the cytotoxic activity of T lymphocyte through BxPC-3 survival curve. sEVs down-regulated the cytotoxic activity of T lymphocytes, and it was rescued by ATM inhibitor KU-60019. Each point in the graph constituting the curve represented the average of 4 biological replicates.

Graphs of western blots and immunofluorescence are from a single experiment, which is representative of three independent experiments.