Figure 1

Silencing of Cyr61 inhibited the proliferation of Du145 cells. (A) The protein level of Cyr61 in four PCa cell lines. (B) Du145 cells were infected with lentivirus either expressing Cyr61 siRNA (Du145/Cyr61i) or the control siRNA (Du145/Ctrli), which processed the same A/T composition but different sequences, and not matched with any genes. Three sequences of either Cyr61 siRNA or control siRNA were designed. Lane a: Du145/Ctrli #1 cells; lane b: Du145/Cyr61i #1 cells; lane c: Du145/Ctrli #2 cells; lane d: Du145/Cyr61i #2 cells; lane e: Du145/Ctrli #3 cells; lane f: Du145/Cyr61i #3 cells; and lane g: wild-type Du145 (Du145/WT) cells. Tubulin was the loading control of protein samples. (C) In vitro proliferation of Du145/WT, Du145/Ctrli #1, Du145/Cyr61i #1, and Du145/Cyr61i-Cyr61SM cells was examined by MTT assay. (D) Crystal violet assay of colony formation of Du145/Cyr61i #1 and Du145/Ctrli #1 cells. Cells (3 × 10³) were seeded each well in 12-well plates and photographed 2 weeks later. Number of cells was measured by detecting under OD 600 nm. A typical experiment was shown. (E) Crystal violet assay of PC3/Ctrl, PC3/Cyr61i, PC3/Cyr61i-Cyr61SM and PC3/Cyr61 cells was taken under the same way. (F) Du145 cells were infected with lentivirus expressing both EGFP and control RNAi or Cyr61 RNAi as evidenced by representative light-field or fluorescence photos. (G) LnCap cells were infected with lentivirus either expressing Cyr61 (LnCap/Cyr61) or the control GFP (LnCap/GFP). In vitro proliferation of LnCap/Cyr61 and LnCap/GFP cells was examined by MTT assay. (H) Schematic diagram showing the domain structure of wild-type Cyr61, silent mutation of Cyr61. Six nucleic acid changed Cyr61SM without changing the amino acid of Cyr61 making it resistant to the RNA interference function of 1# siRNA sequence. ^*P<0.05, ^**P<0.01, Student's t-test.