Inhibition of hyaluronan synthases decreases matrix metalloproteinase-7 (MMP-7) expression and activity.
BACKGROUND
Hyaluronan (HA) and its biosynthetic enzymes hyaluronan synthases (HAS2 and HAS3) mediate Matrigel invasion by SW620 colon carcinoma cells. Because matrix metalloproteinases (MMPs) have been implicated in cancer invasion, we hypothesized that changes in HAS expression would alter MMP expression and activity in these cells.


METHODS
To determine whether an MMP was involved in invasion, Matrigel invasion assays with SW620 cells were performed in the presence or absence of the MMP inhibitors GM6001 or TIMP2. HAS isozymes were inhibited by stably transfecting SW620 cells with vectors that contained antisense HAS2 and/or -3 cDNA; transfection with an empty vector served as a control. MMP-7 transcription was assessed by quantitative reverse transcription polymerase chain reaction (RT-PCR). MMP-7 protein was detected by enzyme-linked immunosorbent assay (ELISA) and enzymatic activity compared using zymography.


RESULTS
GM6001 and TIMP2 decreased Matrigel invasion, which confirms that an MMP played a key role in this process. MMP-7 expression was then detected in SW620 cells. Finally, MMP-7 expression, protein, and enzymatic activity were significantly lower in antisense HAS tranfectants than in SW620 or vector control cells.


CONCLUSIONS
We have demonstrated previously that inhibition of HAS expression and HA production in SW620 colon carcinoma cells inhibits Matrigel invasion. In the studies presented here, we have demonstrated that SW620 cells express high levels of MMP-7 and that inhibition of HAS isozymes dramatically decreases MMP-7 expression, protein, and enzymatic activity. Taken together, these findings suggest that HAS and HA may mediate cellular invasion via changes in MMP-7 expression.
Dunn
Kelli M Bullard
KM
Department of Surgery and Surgical Oncology, Roswell Park Cancer Institute, University at Buffalo/State University of New York, Buffalo, NY 14263, USA. kelli.bullarddunn@roswellpark.org
Lee
Peter K
PK
Wilson
Christopher M
CM
Iida
Joji
J
Wasiluk
Karen R
KR
Hugger
Marie
M
McCarthy
James B
JB
D002448:Cell Adhesion / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003094:Collagen; D015179:Colorectal Neoplasms / Q000378:metabolism* / Q000473:pathology*; D016373:DNA, Antisense / Q000235:genetics; D004151:Dipeptides / Q000494:pharmacology; D018450:Disease Progression; D004338:Drug Combinations; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000378:metabolism*; D007797:Laminin; D020783:Matrix Metalloproteinase 7 / Q000378:metabolism*; D011509:Proteoglycans; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000494:pharmacology; D014162:Transfection
D016373:DNA, Antisense; D004151:Dipeptides; D004338:Drug Combinations; D007797:Laminin; C078131:N-(2(R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl)-L-tryptophan methylamide; D011509:Proteoglycans; C059485:matrigel; D019716:Tissue Inhibitor of Metalloproteinase-2; D006820:Hyaluronic Acid; D003094:Collagen; D014453:Glucuronosyltransferase; C101754:HAS2 protein, human; C105458:HAS3 protein, human; D000076002:Hyaluronan Synthases; D020783:Matrix Metalloproteinase 7