Figure 4

CD8⁺ and CD4⁺ T cell infiltration and accumulation in tumors. (A) APJ expression by qRT-PCR. CD8⁺ and CD4⁺ T cells were isolated from WT and Apelin-KO mice bearing tumors at day19 by flow cytometry. APJ expression levels were normalized to GAPDH. (B) Flow cytometric analysis of total infiltrated CD8⁺ and CD4⁺ T cells. Cell numbers are presented as percentages of total living cells (n = 6 for each group). (C) Immunofluorescence staining of CD8 (green) and CD4 (red) in the frozen sections of MC38 tumors from WT or Apelin-KO mice. Tumor samples were collected at day 19. Scale bar = 100 µm. (D) Quantitative evaluation of the number of infiltrated CD8⁺ and CD4⁺ T cells in MC38 tumors (6 random fields of 5 independent tumor sections). (E) Immunofluorescence staining of CD8 (green) and CD4 (red) in the frozen sections of MC38 tumors from WT mice infused with [Pyr¹]Apelin-13 or saline using osmotic pumps. Tumor samples were collected at day 19. Scale bar = 100 µm. (F) Quantitative evaluation of the number of infiltrated CD8⁺ and CD4⁺ T cells in MC38 tumors (6 random fields of 5 independent tumor sections). The error bars indicate mean ± SD and all data were analyzed by two-sided Student's t-test. *p < 0.05, **p < 0.01.