ABSTRACT
Peroxisome proliferator-activated receptor gamma (PPARgamma) ligands have been shown to inhibit angiogenesis. We showed that treatment with 15d-PGJ(2), a PPARgamma ligand, downregulate the expressions of angiopoietin-1 (Ang-1) in gastric cancer cells MKN45. The medium of MKN45 cells treated with 15d-PGJ(2) significantly inhibited the migration and tube formation of human umbilical vein endothelial cells (HUVECs). Moreover, Matrigel plug assay revealed that 15d-PGJ(2) reduced in vivo angiogenesis induced by MKN45 cells. These modulations were restored by the addition of recombinant Ang-1. Our findings supported that 15d-PGJ(2) suppressed angiogenesis of gastric cancer cells by downregulation of Ang-1.
Subject(s)
Angiopoietin-1/genetics , Neovascularization, Pathologic/prevention & control , Prostaglandin D2/analogs & derivatives , Stomach Neoplasms/blood supply , Angiogenesis Inhibitors/pharmacology , Animals , Cell Line , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Down-Regulation/drug effects , Down-Regulation/genetics , Endothelial Cells/cytology , Endothelial Cells/drug effects , Endothelial Cells/physiology , Female , Gene Expression Profiling , Humans , Mice , Mice, Inbred BALB C , Mice, Nude , Neovascularization, Pathologic/genetics , Neovascularization, Pathologic/metabolism , Neovascularization, Physiologic/drug effects , Oligonucleotide Array Sequence Analysis , Prostaglandin D2/pharmacology , Stomach Neoplasms/pathology , Vascular Endothelial Growth Factor A/metabolismABSTRACT
We characterized the effects of selective Cox-2 inhibition on the angiogenesis gastric cancer cell line SGC7901 by stable transfection of antisense Cox-2 cDNA (Cox-2-AS) or pharmacological inhibition by selective cox-2 inhibitor (NS398). The conditioned media obtained from SGC7901 treated with Cox-2-AS or NS398 suppressed the proliferation, migration and tube formation of human umbilical vein endothelial cells. Moreover, both treatments inhibited in vivo angiogenesis. These inhibitions could be partially reversed by the addition of PGE2. Our findings support that selective inhibition of Cox-2 alone plays an instrumental role on gastric cancer associated angiogenesis.
Subject(s)
DNA, Antisense , Neovascularization, Pathologic , Prostaglandin-Endoperoxide Synthases/genetics , Prostaglandin-Endoperoxide Synthases/metabolism , Stomach Neoplasms/pathology , Cyclooxygenase 2 , Cyclooxygenase 2 Inhibitors , Cyclooxygenase Inhibitors/pharmacology , DNA, Complementary , Humans , Membrane Proteins , Nitrobenzenes/pharmacology , Sulfonamides/pharmacology , Transfection , Tumor Cells, CulturedABSTRACT
AIM: To investigate the apoptosis-inducing effect of Caspases-3 expressed by constructed eukaryotic vector on gastric cancer cell line SGC7901. METHODS: PCR was employed to amplify the sequences of both small and large subunits of Caspases-3. Its products were separately cloned into the Sma I site of pBluescript KS(+) to generate both plasmids pBS/SS and pBS/LS. The small subunit fragment was excised from plasmid pBS/SS with BamH I and then inserted into the BamH I site of plasmid pBS/LS preceding that of the large subunit to yield plasmid pBS/Rev-Caspase-3. Rev-Caspase-3 cDNA was excised with Kpn I+Xba I and then subcloned into plasmid pcDNA3.1 (+) to construct Rev-Caspase-3 eukaryotic expression vector pcDNA/Rev-Caspase-3, which was used to transiently transfect SGC7901 cell line. Cell count, MTT assay and electron microscopy were used to confirm the antiproliferation and apoptosis-inducing effect of Rev-Caspase-3 expression on gastric cancer cells. RESULTS: Plasmid pBS/Rev-Caspase-3 and eukaryotic expression vector pcDNA/Rev-Caspase-3 were successfully constructed. SGC7901 cells were transiently transfected by either pcDNA/Rev-Caspase-3 or pcDNA3.1 (+) for 24, 48, 72, and 96 h respectively. Cell growth was measured by cell count and MTT assay. In cell count assay, the cell numbers were 1.8 X 10(6), 1.55 X 10(6), 2.0 X 10(6), and 3.1 X 10(6) in the experimental group and 2.5 X 10(6), 3.1 X 10(6), 4.0 X 10(6), and 5.7 X 10(6) in the control group at 24, 48, 72 and 96 h respectively. The growth of SGC7901 cells was suppressed by Rev-Caspase-3 in a time-dependent manner (P<0.05). The results of MTT assay were similar to that of cell count (P<0.05). The characteristics of apoptosis such as chromatin condensation, crescent formation and margination were seen and more obvious with time in the given-experimental period in the experimental group, but not easily observed in the control group. CONCLUSION: The expression of Rev-Caspase-3 by the constructed eukaryotic vector can significantly induce apoptosis of gastric cancer cell line SGC7901, which may exhibit a potential way in gastric cancer gene therapy.
