Suppression of invasive properties of colorectal carcinoma SW480 cells by 15-hydroxyprostaglandin dehydrogenase gene.
Colorectal carcinoma (CRC) is often lethal when invasion and/or metastasis occur. NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase (15-PGDH), an enzyme involved in prostaglandin (including PGE(2)) bio-inactivation, is down-expressed in several epithelial malignancies including CRC. Although its role in the suppression of colon tumorigenesis has been well learned, little is known about the role of 15-PGDH in the process of tumor metastasis. Here, we tested the hypothesis that 15-PGDH over-expression in CRC cells results in decreased cell motility and invasion. In this study, 15-PGDH was re-expressed in SW480 cells by the use of gene transient transfection with eukaryotic expression vector pcDNA3.1-PGDH. We confirmed the over-expression of 15-PGDH protein by Western blot and enzymatic activity assay. The cell motility was tested by counting the number of cells crossing an 8-micron pore size PET membrane and by measuring cells migration distance through wound healing assay. Furthermore, cell invasive activity was evaluated by counting the number of cells invading through a Matrigel-coated membrane simulating basement membrane. The effects of 15-PGDH on the adhesion were investigated by MTT assay. Ectopic expression of 15-PGDH in SW480 cancer cells significantly inhibited the cell migratory and invasive capacity in vitro by approximately 1.9- and 8.4-fold, respectively. To test the hypothesis that 15-PGDH affects proteases and inactivates extracellular matrix (ECM), Western blot and gelatin zymography were performed by using serum-free conditioned medium. The results showed that re-expression of 15-PGDH suppresed matrix metalloproteinase-2 (MMP2) synthesis and secretion. In addition, the analysis of the MMP2 activity indicated that re-expression of 15-PGDH could inhibit activation of MMP2. Furthermore, we found that 15-PGDH inhibited cell adhesion to ECM and reduced CD44 expression in SW480 cell. Taken together, these results suggest that induced 15-PGDH expression may contribute to the inhibition of the invasive and metastatic capacity of colon cancer cells in vitro.
Li
Meining
M
Department of Biochemistry and Molecular Biology, Shanxi Medical University, Taiyuan City, People's Republic of China.
Xie
Jun
J
Cheng
Le
L
Chang
Bingmei
B
Wang
Ying
Y
Lan
Xiaoqin
X
Zhang
Dong
D
Yin
Yunqin
Y
Cheng
Niuliang
N
D000230:Adenocarcinoma / Q000201:enzymology* / Q000473:pathology*; D002448:Cell Adhesion / Q000235:genetics; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D015179:Colorectal Neoplasms / Q000201:enzymology* / Q000473:pathology*; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis; D006911:Hydroxyprostaglandin Dehydrogenases / Q000096:biosynthesis* / Q000235:genetics; D020778:Matrix Metalloproteinase 2 / Q000096:biosynthesis; D009361:Neoplasm Invasiveness; D014162:Transfection
D018960:Hyaluronan Receptors; D006911:Hydroxyprostaglandin Dehydrogenases; C035859:15-hydroxyprostaglandin dehydrogenase; D020778:Matrix Metalloproteinase 2