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Fig. 3 | Military Medical Research

Fig. 3

From: Aspartoacylase suppresses prostate cancer progression by blocking LYN activation

Fig. 3

ASPA knockdown promotes PC-3 cell proliferation and migration in vitro. a Western blotting results of ASPA protein expression in PC-3 cells transfected with shRNA or shASPA. Protein expression levels were normalized to β-actin levels. b CCK-8 assay showed that ASPA knockdown promoted PC-3 cell proliferation. c Representative images of EdU-positive PC-3 cells transfected with shRNA or shASPA (scale bar = 100 μm). The graph on the right shows the percentage of EdU-positive nuclei. The data were obtained from 7 fields of 3 independent experiments. d Colony formation assay showed that ASPA knockdown promoted PC-3 cell colony formation ability. The graph on the right shows the colony numbers from 3 independent experiments. e RT-qPCR results of proliferation-related genes in PC-3 cells transfected with shRNA or shASPA. The mRNA expression levels were normalized to ACTB levels. f Western blotting results (left) and quantification results (right) for proliferation-related proteins in PC-3 cells transfected with shRNA or shASPA. Protein expression levels were normalized to β-actin levels. g Transwell assays showed that ASPA knockdown promoted PC-3 cell migration and invasion (scale bar = 100 μm). The graph on the right shows the migrating cells and the invading cells. The data were obtained from 8 fields of 3 independent experiments. h RT-qPCR results of epithelial–mesenchymal transition genes in PC-3 cells transfected with shRNA or shASPA. The mRNA expression was normalized to ACTB levels. i Western blotting results (left) and quantification results (right) of epithelial–mesenchymal transition proteins in PC-3 cells transfected with shRNA or shASPA. The protein expression was normalized to β-actin levels. j Flow chart of RNA-Seq in PC-3 cells transfected with shRNA or shASPA. k Cluster analysis showed the global sample distribution profiles of the shRNA group and shASPA group based on the RNA-Seq dataset in PC-3 cells. l GSEA results showed the significantly altered cancer hallmarks based on the RNA-Seq dataset in PC-3 cells from the shRNA group and shASPA group. The data are presented as the mean ± standard deviation (SD). ASPA aspartoacylase, CCK-8 cell counting kit 8, CCND1 cyclin D1, CDH1 cadherin 1, CDH2 cadherin 2, EdU 5-ethynyl-2′-deoxyuridine, GSEA gene set enrichment analysis, MYC v-Myc myelocytomatosis viral oncogene homolog, MMP9 matrix metallopeptidase 9, NES normalized enrichment score, OD optical density, PCNA proliferating cell nuclear antigen, RNA-Seq RNA sequencing, RT-qPCR real-time quantitative PCR, shRNA small hairpin RNA. **P < 0.01

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