Sperm associated antigen 9 promotes oncogenic KSHV-encoded interferon regulatory factor-induced cellular transformation and angiogenesis by activating the JNK/VEGFA pathway
Fig 3
Knockdown of SPAG9 inhibits vIRF1-induced angiogenesis, cell proliferation and migration.
(A). Western-blotting analysis of SPAG9 expression in vIRF1-expressing HUVECs transduced with lentivirus-mediated No.1 (shSPAG9-1) and No. 2 (shSPAG9-2) shRNAs targeting SPAG9. (B). Lentiviral vIRF1- or its control pHAGE-infected endothelial cell line were transduced with lentivirus-mediated No.1 (shSPAG9-1) and No. 2 (shSPAG9-2) shRNAs targeting SPAG9, and then were subjected to chicken chorioallantoic membranes (CAMs) assay. Representative images are shown. Magnification, ×100. Scar bars, 40 μm. (C). Quantification of CAMs assay described in (B). (D). Cells treated as in (B) were mixed with the high concentration Matrigel, and then were injected into the right flanks of nude mice. Plugs were harvested 10 days after the injection and photographed using stereomicroscope. Representative images of Matrigel plug assay in mice are displayed. (E). Quantification of Matrigel plug assay in mice described in (D). (F). CCK-8 assay of HUVECs treated as in (A). (G). Transwell migration analysis of HUVECs treated as in (A). The migrated and invaded HUVECs were counted at 6 h and 12 h post seeding. (H). Quantification of Transwell migration assay described in (G). (I). Western-blotting analysis of SPAG9 expression in KSHV-infected HUVECs transduced with lentivirus-mediated No.1 (shSPAG9-1) and No. 2 (shSPAG9-2) shRNAs targeting SPAG9. (J). CCK-8 assay of HUVECs treated as in (I). (K). Transwell migration analysis of HUVECs treated as in (I). The migrated and invaded HUVECs were counted at 6 h and 12 h post seeding. Data were shown as mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001, Student's t-test.