Effect Of Retrogene NANOGP8 On The Biological Characteristics Of Human Gastric Cancer Cells

OBJECTIVE:To construct an eukaryotic expression vector pEGFP-N1-NANOGP8 and recombinant plasmids shRNA-NANOGP8 which could inhibit the expression of NANOGP8 by RNA interference technology, and examine the proliferation, apoptosis, cell cycle, invasion, and migration of human gastric cancer cell line SGC-7901 after transfection.METHODS:(1) Total mRNAs were extracted from human ovarian cell line SKOV-3, then the NANOGP8 cDNAs were obtained by RT-PCR.The pEGFP-NI-NANOGP8 was constructed by cloning the cDNAs into pEGFP-NI and confirmed by enzyme digestion and sequencing analysis. (2)To construct recombinant plasmids named pshRNA-NANOGP8 and pshRNA-negative. The two recombinant plasmids were confirmed by enzyme digestion and sequencing analysis. (3) Then, vector pEGFP-NI-NANOGP8 and recombinant plasmid shRNA-NANOGP8 were transfected into SGC-7901 cells via liposome. The NANOGP8 mRNA and protein were tested by fluorescence microscopy, RT-PCR and Western blotting, respectively. The proliferation of SGC-7901 was tested by CCK8, The apoptosis and the cell cycle were examined by flow cytometry, and NANOGP8 promoted invasion and migration in SGC-7901 were verified by Transwell assay.RESULTS:(1) The NANOGP8 cDNAs were obtained by RT-PCR, The recombinant plasmid PEGEP-NI-NANOGP8 was constructed successfully. (2) The pshKNA-NANOGPS and pshRNA-negative were successful constructed. (3) pEGFP-N1-NANOGP8 transfection group showed high expression of NANOGP8, Conversely, pshRNA-NANOGP8 transfection group showed low expression of NANOGP8. High expression of NANOGP8 significantly promoted the proliferation of tumor cells, arrest cells cycle at the S phase, decreased apoptosis cell and increased invasion and migration obviously. While the low expression group cell inhibit cell proliferation, arrest cell cycle at the G0-G1 phase, promote cell apoptosis and inhibit the migration and invasion.CONCLUSIONS:NANOGP8, a retrogene gene can transcript and express in SGC-7901 cells, promoted the proliferation, cycle, apoptosis, migration and invasion of gastric cancer.