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. 2017 May 16;6(7):e1327494. doi: 10.1080/2162402X.2017.1327494

Figure 5.

Figure 5.

PD-L2 expression is inducible by IFNγ in colorectal cancer cells. (A) The relationship between gene copy number alterations of PD-L1/PD-L2 and their mRNA expression levels. No significant difference was found between the mRNA levels in tumors bearing different types of copy number alterations (two-sided student t-test). (B) The molecular signatures associated with PD-L2 in CRC as suggested by GSEA analysis. The C2 (curated gene sets), C5 (Gene Ontology gene sets), and C7 (immunologic signatures) gene sets were, respectively, used as reference gene sets, and the top hits are provided. (C) Immunologic (C7) signatures that associated with PD-L2 expression as determined by GSEA analysis. The network plots represent “enrichment maps” generated by Cytoscape program according to the similarity between the gene sets. Relatively more significant results are marked in red, with less significant gene sets in yellow. (D) The GSEA plot for the signature “IFNG_IN_CD8POS_DC_UP,” which represents a set of genes that can be upregulated in dendritic cells by treatment with IFNγ. (E) Western Blot using specific antibody for PD-L2 revealed significant upregulation of PD-L2 by treatment of IFNγ. The human colorectal cancer RKO and Lovo cells were incubated with IFNγ at the indicated concentrations for 12 h, followed by cell lysis and Western Blot analysis. GAPDH was detected as loading control. (F) Correlation between the mRNA expression of PD-L2 and IFNγ (IFNG) according to the microarray data of TCGA colorectal cancer cohort. (G) Immunohistochemistry staining of PD-L2 and IFNγ in the tissue specimen of the same patients, according to the Human Protein Atlas data set. Representative images in each column correspond to the same patient. (H) Statistical results of PD-L2 and IFNγ expression correlation using the Chi-square test. The Human Protein Atlas CRC subjects (with the IHC results of both PD-L2 and IFNγ) were included in the analysis, and the positivity of IFNγ was based on its expression in either the cancer cells or adjacent immune cells (higher expression is considered representative).