Published online May 28, 2015. doi: 10.3748/wjg.v21.i20.6215
Peer-review started: November 28, 2014
First decision: December 26, 2014
Revised: January 14, 2015
Accepted: February 12, 2015
Article in press: February 13, 2015
Published online: May 28, 2015
AIM: To investigate the mechanism underlying the promoting role of CD97 in gastric cancer cell proliferation and invasion.
METHODS: Two types of exosomes released by gastric cancer cells with high (SGC/wt) or low (SGC/kd) CD97 expression were isolated by ultracentrifugation and identified by electron microscopy and western blot analysis. The influences of the two exosomes on gastric cancer cell proliferation and invasion were investigated by proliferation and Matrigel invasion assays. Exosomal miRNAs were subsequently isolated from the two samples and their miRNA profiles were compared via microarray assay analysis. Reverse transcription-quantitative real-time polymerase chain reaction was used to validate the microarray assay. Target genes of the differently expressed microRNAs were predicted based on five independent algorithms and were then subjected to gene oncology enrichment and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis. After identifying the pathway that was the most likely altered, tumor cells were treated with the two exosomes at different concentrations, and the pathway activation was identified through western blot analysis.
RESULTS: Exosomes isolated from SGC/wt cells significantly promoted tumor cell proliferation in a dose-dependent manner in vitro. SGC/wt exosomes also significantly elevated the invasiveness of both SGC/wt (129.67 ± 8.327 vs 76.00 ± 5.292, P < 0.001) and SGC/kd (114.52 ± 9.814 vs 45.73 ± 4.835, P < 0.001) cells as compared to the exosomes released by SGC/kd cells. Microarray assay of the two exosomes revealed that 62 miRNAs were differently regulated with a signal intensity of > 500 and a false discovery rate < 0.05. The following KEGG analysis defined the MAPK signaling pathway as the most likely candidate pathway that regulated tumor cell proliferation and invasion. Through western blot analysis, significant up-regulations of phosphorylated MAPKs, including extracellular signal-regulated kinase, Jun NH2-terminal kinase, and p38 mitogen-activated protein kinase, were detected in a dose-dependent manner in the SGC/wt exosomes treated groups, confirming activation of the MAPK signaling pathway stimulated by SGC/wt exosomes.
CONCLUSION: CD97 promotes gastric cancer cell proliferation and invasion in vitro through exosome-mediated MAPK signaling pathway, and exosomal miRNAs are probably involved in activation of the CD97-associated pathway.
Core tip: CD97, belongs to the epidermal growth factor-seven-transmembrane subfamily, and has been found to promote proliferation and invasion of gastric cancer cells. However, the underlying mechanism is poorly understood. In this study, we found that exosomes isolated from gastric cancer cells with high CD97 expression promoted tumor cell proliferation and invasion. Furthermore, through microarray and western blot analyses, MAPK signaling pathway activation was observed when cells were treated with those exosomes. These results indicated that CD97 promotes gastric cancer cell proliferation and invasion in vitro, at least in part, through the exosome-mediated MAPK signaling pathway, and exosomal miRNAs are probably involved in activation of the CD97-associated pathway.