Fig. 3

Akt activation is involved in spermine-mediated PD-L1 expression in HCC. A Effects of spermine (200 µM) on the level of Akt Ser473 phosphorylation in SNU-368 cells. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA, n = 5 independent experiments per group. B Effects of pretreatment with 10 µM NPS2134 or 10 µM BAPTA-AM on spermine-induced increased phosphorylation levels of Akt at Ser473 in SNU-368 cells after 12 h of coincubation. **p < 0.01, ***p < 0.001, one-way ANOVA, n = 5 independent experiments per group. C, D Effects of ectopic expression of myr-Akt1 isoform on endogenous CD274 mRNA (C) and PD-L1 protein (D) in SNU-368 cells. **p < 0.01, ***p < 0.001, two-tailed unpaired t test, n = 5 independent experiments per group. E, F Effects of pretreatment with Akt inhibitor MK2206 (10 µM, “MK”) on spermine-induced the expression of CD274 mRNA (E) and PD-L1 protein (F) in SNU-368 cells after incubation with spermine for 24 h. **p < 0.01, ***p < 0.001, one-way ANOVA, n = 5 independent experiments per group. G Immunohistochemical staining of phosphor-Akt and PD-L1 in 24 human HCC specimens. Scale bar, 50 μm. H Spearman correlation test between blood spermine concentration and tumor phosphor-Akt IHC scores. I Spearman correlation analysis of phosphor-Akt and PD-L1. Note that the scores of some samples overlap