A new study published by researchers from Zhejiang University sought to investigate the mechanism underlying the promoting role of CD97 in gastric cancer cell proliferation and invasion. In their study, 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 using LC Sciences’ miRNA microarray service. 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. Researchers saw that 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. The results of LC Sciences’ microarray service 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. Investigators conclude that 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.

Comparison of miRNA profiles of the two exosomes by microarray analysis and validation of differently expressed miRNAs by RT-qPCR analysis. A: Bar graph showed differently expressed miRNAs in kd-exo by microarray as compared to wt-exo; B: Four miRNAs that displayed either an increased or decreased expression were selected for RT-qPCR validation of microarray assay. The same expression trends were observed between microarray and RT-qPCR for all the miRNAs. wt-exo: SGC/wt-derived exosomes; kd-exo: SGC/kd-derived exosomes.

Related Service

miRNA Microarray Service – LC Sciences provides a microRNA (miRNA) expression profiling service using microarrays based on our in-house developed µParaflo® technology platform. We have standard arrays for all mature miRNAs of all species available in the latest version of the miRBase database (Release 21, July 2014). Our service is comprehensive and includes sample labeling, array hybridization, image data processing and in-depth data analysis. Two-three weeks after receiving your total RNA samples, we’ll send you both the raw and fully analyzed data. [Learn more…]

Reference

C. Li, D.R. Liu, G.G. Li, H.H. Wang, X.W. Li, W. Zhang, Y.L. Wu, L. Chen (2015) CD97 promotes gastric cancer cell proliferation and invasion through exosome-mediated MAPK signaling pathway World J Gastroenterol.. 21(20): 6215-6228 doi: 10.3748/wjg.v21.i20.6215 [article]