Stable knockdown of heparanase expression in gastric cancer cells in vitro

AIM:To develop short hairpin RNA(shRNA)against heparanase,and to determine its effects on heparanase expression and the malignant characteristics of gastric cancer cells. METHODS:Heparanase-specific shRNA was constructed and transferred into cultured the gastric cancer cell line SGC-7901.Stable subclonal cells were screened by G418 selection.Heparanase expression was measured by reverse transcriptase-polymerase chain reaction(RT-PCR),real-time quantitative PCR and Western blotting.Cell proliferation was detected by 2-(4,5-dimethyltriazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT)colorimetry and colony formation assay. The in vitro invasiveness and metastasis of cancer cells were measured by cell adhesion assay,wound healingassay and matrigel invasion assay.The angiogenesis capabilities of cancer cells were measured by tube formation of endothelial cells. RESULTS:Stable transfection of heparanase-specific shRNA,but not of scrambled shRNA and mock vector,resulted in reduced mRNA and protein levels of heparanase.The shRNA-mediated knockdown of heparanase did not affect the cellular proliferation of SGC-7901 cells.However,the in vitro invasiveness and metastasis of cancer cells were decreased after knockdown of heparanase.Moreover,transfection of heparanase-specific shRNA decreased the in vitro angiogenesis capabilities of SGC-7901 cells. CONCLUSION:Stable knockdown of heparanase can efficiently decrease the invasiveness,metastasis and angiogenesis of human gastric cancer cells.In contrast,stable knockdown of heparanase does not affect the cell proliferation.

Effects of resistin-like molecule β over-expression on gastric cancer cells in vitro

AIM: To investigate the effects of resistin-like molecule β (RELMβ) over-expression on the invasion, metastasis and angiogenesis of gastric cancer cells. METHODS: Human RELMβ encoding expression vec tor was constructed and transfected into the RELMβ lowly-expressed gastric cancer cell lines SGC7901 and MKN-45. Gene expression was measured by Western blotting, reverse transcription polymerase chain reaction (PCR) and real-time quantitative PCR. Cell proliferation was measured by 2-(4,5-dimethyltriazol-2-yl)-2,5-diphenyl tetrazolium bromide colorimetry, colony formation and 5-ethynyl-20-deoxyuridine incorporation assays. The in vitro migration, invasion and metastasis of cancer cells were measured by cell adhesion assay, scratch assay and matrigel invasion assay. The angiogenic capabilities of cancer cells were measured by tube formation of endothelial cells. RESULTS: Transfection of RELMβ vector into SGC-7901 and MKN-45 cells resulted in over-expression of RELMβ, which did not infl uence the cellular proliferation. However, over-expression of RELMβ suppressed the in vitro adhesion, invasion and metastasis of cancer cells, accompanied by decreased expression of matrix metalloproteinase-2 (MMP-2) and MMP-9. Moreover, transfection of RELMβ attenuated the expression of vascular endothelial growth factor and in vitro angiogenic capabilities of cancer cells. CONCLUSION: Over-expression of RELMβ abolishes the invasion, metastasis and angiogenesis of gastric cancer cells in vitro, suggesting its potentials as a novel therapeutic target for gastric cancer.

Helicobacter pylori Infection Facilitates the Expression of Resistin-like Molecule Beta in Gastric Carcinoma and Precursor Lesions

Helicobacter pylori(H.pylori)was reported to be associated with gastric carcinogenesis.Resistin-like molecule beta(RELMβ),a recently described goblet cell-specific protein,was demonstrated to aberrantly express in gastric cancer and correlated with its clinicopathological features.This study aimed to examine the association between H.pylori and RELMp expression in gastric carcinoma and precursor lesions.H.pylori infection and RELMβexpression were immunohistochemically evaluated in gastric biopsies from 230 patients.The biopsies consisted of normal gastric mucosa(w=20),mucosa with chronic gastritis(n=41),intestinal metaplasia(n=42),dysplasia(w=31),intestinal-type adenocarcinoma(n=56),and diffuse-type adenocarcinoma(n=40).RELMβ expression was measured in gastric biopsies after H.pylori eradication therapy in a subgroup of 32 patients.Cultured gastric cancer cell line SGC-7901 was infected with H.pylori strains,and RELMp expression was detected by reverse transcription PCR,real-time PCR and Western blotting.Higher RELMP immunoreactivity was observed in H.pyloripositive intestinal metaplasia(P=0.003),dysplasia(P=0.032),intestinal-type(P=0.037)and diffuse-type adenocarcinomas(P=0.001)than in H.pylori-negative specimens.Expression rates of RELMβ in dysplasia(CO.005),intestinal-type adenocarcinoma(P<0.001),and diffuse-type adenocarcinoma(P=0.001)were significantly correlated with the grade of H.pylori density.In addition,H.pylori eradication reduced the RELMβintensity in intestinal metaplasia(P=0.001).Infection of gastric cancer SGC-7901 cells with cag pathogenicity island(PAI)-positive H.pylori TN2,but not with its PAI totally deleted mutant(TN2-APAI)for 4-8 h,resulted in enhanced protein and transcript levels of RELMβ(P<0.05).In summary,our study suggested that H.pylori infection facilitated the expression of RELMβ in gastric garcinoma and precursor lesions.

Gastric carcinoma with osteoclast-like giant cells:a case report and review of the literature

Gastric carcinoma with osteoclast-like giant cells(OGCs) is an extremely rare tumor. So far,only six cases have been reported in the literature. Here we report an additional case of this tumor in a Chinese 78-year-old man presented with abdominal pain,vomiting,and hematemesis. Physical examination and gastroscopy revealed a tumor in the gastric antrum. The biopsy and pathological findings indicated a gastric adenocarcinoma with OGCs,which were present in both the tumor and the metastatic lymph nodes. Further immunohistochemical staining indicated that OGCs were reactive with CD68,CD45,and vimentin protein,but not with pancytokeratin,carcinoembryonic antigen,or epithelial membrane antigen,suggesting the monocytic/histiocytic derivation of these OGCs. In situ hybridization for Epstein-Barr virus showed no nuclear positivity in either adenocarcinoma or OGCs. Postoperative follow-up showed that the patient had survived for at least 6 months without recurrence. Further investigation is warranted to clearly define the prognostic significance of OGCs in gastric carcinoma.