ABSTRACT
AIM:To explore the effect of glucose-regulated protein 78 (GRP78) on the gastric carcinogenesis. METHODS:GPR78 expression patterns were examined in 34 specimens from gastric carcinoma patients using the immu-nohistochemistry (IHC) assay, and in 10 specimens using Western blotting analysis .In addition, the expression of GPR78 and cyclin D1 was detected in human gastric cancer cell lines SGC 7901 and SGC7901-H78 (overexpressing GRP78) by Western blotting.RESULTS:By IHC assay, GRP78 was found to be highly expressed in the cytoplasm of gastric carcino-mas as compared with the adjacent non-malignant tissues and corresponding normal tissues .GRP78 expression was positive-ly correlated with gender and histological differentiation (P0.05).Furthermore, we found that with the increased expression of GRP 78 in SGC7901-H78 cells, the expres-sion of cyclin D1 was also elevated .CONCLUSION:GRP78 might be a key player to be involved in the growth of gastric cancer.
ABSTRACT
[ ABSTRACT] AIM:To construct a lentiviral vector for stable delivery of the ER-α36 gene and to detect its effect on SGC7901 cell growth.METHODS: The efficient RNAi targeting sequences identified for the ER-α36 gene were screened.The Oligo DNA was synthesized with target sequences and annealed to form double-stranded DNA.Then it was digested by Xho I and EcoR I and connected with GV307 vector to produce LV-ER-α36-RNAi lentiviral vector.PCR was used to screen the positive clones and sequence.The LV-ER-α36-RNAi, pHelper 1.0 and pHelper 2.0 plasmids were co-transfected into 293T cells for producing lentiviral vector and infecting SGC7901 cell line.Fluorescence microscopy, real-time PCR and Western blotting were used to detect the transfection efficiency and gene silencing effect.17β-estrodial at concentration of 1 ×10 -10 mol/L was used to stimulate the recombinant cell line, and the action on the growth of gastric cancer cells and the expression of Src, ERK1/2 and cyclin D1 were determined.RESULTS: DNA sequencing analysis confirmed the identity of recombinant shRNA expression vectors.Immunofluorescence assay demonstrated that transfection efficiency was above 80%.Transfection of LV-ER-α36-RNAi significantly knocked down the expression of ER-α36 at mR-NA and protein levels with tetracycline ( TeT) simulating as revealed by real-time PCR and Western blotting.Compared with control group, the growth of the recombinant cell line declined and the expression of Src, ERK1/2 and cyclin D1 and the activation of Src decreased (P<0.05).CONCLUSION: Lentiviral vectors that silence ER-α36 expression are con-structed successfully and can be used to study the role of ER-α36 in gastric cancer.The ER-α36 is related with many kinds of cancer cell growth, including gastric cancer cells.