Background
Results
PSCA was highly expressed in the metastatic DU145 prostate cancer cells
Successful construction of stable knockdown of PSCA DU145 cell lines
Knockdown of PSCA reduced cell migration and invasion in prostate cancer DU145 cells
Knockdown of PSCA induced epithelial-to-mesenchymal transition in vitro
Knockdown of PSCA induced EMT relating genes in DU145
Knockdown of PSCA induced epithelial-to-mesenchymal transition in vivo
Discussion
Conclusions
Methods
Cell lines and cell culture
Antibodies and immunoblotting
Immunofluorescent staining
Generation of the stable knockdown of PSCA cell line DU145
RT-PCR
Clone name | Location | PSCA target sequence |
---|---|---|
PSCA shRNA#1 | 229 | CCTGACCGTCATCAGCAAAGG |
PSCA shRNA#2 | 294 | GCAAGAAGAACATCACGTGCT |
PSCA shRNA#3 | 528 | GAGGCACATCCTAACGCAAGT |
PSCA shRNA#4 | 841 | GGCTGAGATGAAGTGGACTGA |
Scramble | GCTTCGCGCCGTAGTCTTA |
Gene | Sequences | |
---|---|---|
Forward | Reverse | |
Human experiments | ||
PSCA | GCTGCTTGCCCTGTTGATGG | ACATGGTCAGACTTGCGTTAGGA |
GAPDH | CCATCACTGCCACCCAGAAGAC | GTGTCGCTGTTGAAGTCAGAGGAGA |
E-cadherin | TCCTCCCAATACATCTCCCTTCA | TCTCCGCCTCCTTCTTCATCATA |
Vimentin | TTCGCCAACTACATCGACAAGG | TTCAAGGTCAAGACGTGCCAG |
Real-time quantitative PCR
Gene | Sequences | |
---|---|---|
Forward | Reverse | |
Human experiments | ||
PSCA | CCTAACGCAAGTCTGACCATGTATG | TGCAGGCGGATCTGTGTCAATA |
GAPDH | CGCTGAGTACGTCGTGGAGTC | GCTGATGATCTTGAGGCTGTTGTC |
E-cadherin | TCACGCTGTGTCATCCAACGG | TCACGCTGTGTCATCCAACGG |
β-catenin | CAGGAAGGGATGGAAGGTCTC | TACCACCCACTTGGCAGACC |
Vimentin | ATCCAAGTTTGCTGACCTCTCTGA | GACTGCACCTGTCTCCGGTACTC |
Fibronectin | GAGACACCTGGAGCAAGAAGG | GTGAGGCTGCGGTTGGTAAAC |
Snail | CTTCTCCTCTACTTCAGTCTCTTCC | TGAGGTATTCCTTGTTGCAGTATTT |
Slug | AACAGAGCATTTGCAGACAGGTC | GCTACACAGCAGCCAGATTCC |
Twist | CACCATCCTCACACCTCTGCATT | GCTGATTGGCACGACCTCTTGA |