In Vitro Regulation of MMP-2 and MMP-9 Expression by Cytokines, Mitogens, Inducers and Inhibitors in Adult Sarcoma Cell Lines

M.W. Roomi, J.C. Monterrey, M. Rath and A. Niedzwiecki
Dr. Rath Research Institute, Santa Clara, CA

Presented at:
49th Annual Meeting of the Society of Toxicology, Salt Lake City, Utah, March 7-11, 2010.

Published in: 
The Toxicologists, Abstract #674, page 143

 

Abstract

Introduction: 
Metastatic sarcoma cells secrete large amounts of MMPs, degrading the ECM and basement membrane and allowing the spread to distal organs. Although many studies have demonstrated the importance of MMPs in invasion and metastasis, little information is currently known regarding the effect of cytokines, mitogens, inducers and inhibitors.

Objective: 
We examined the roles of cytokines, mitogens, inducers and inhibitors in regulating MMPs in adult sarcoma cell lines.

Methods and Materials: 
Human fibrosarcoma (FS) (HT-1080), chondrosarcoma (CS) (SW-1353), liposarcoma (LS) (SW-872) and synovial sarcoma (SS) (SW-982) cell lines (ATCC) were cultured in their recommended media and supplemented with 10% FBS and antibiotics in 24-well cultured plates. At near confluence, the cells were washed with PBS and incubated in serum-free media with: (A) PMA 10, 50, 100 and 200 ng/ml; TNF-a 0.1, 1.0, 10 and 25 ng/ml; Il-1ß 0.1, 1, 10 and 25 ng/ml or LPS 10, 25, 50 and 100 µg/ml; (B) EGCG and doxycycline (Dox), 10, 25, 50 and 100 µg /ml without and with PMA; (C) cyclohexamide, actinomycin D, retinoic acid and dexamethasone; and (D) a nutrient mixture (NM)  containing lysine, proline, ascorbic acid and green tea extract without and with PMA. After 24 hrs the media were removed and analyzed for MMP-2 and MMP-9 by zymography and densitometry.

Results: 
FS, CS and LS exhibited two bands corresponding to MMP–2 and –9, whereas SS exhibited only MMP–2. PMA had a moderate stimulatory effect on MMP–2 in CS and FS, and no effect on LS and SS, whereas it significantly stimulated MMP–9 in all sarcoma cell lines. TNF-a had a stimulatory effect on MMP-2 in CS, and no effect on FS, LS and SS; it had a stimulatory effect on MMP–9 expression in FS, LS, and SS but not in CS. Il-1ß stimulated MMP–2 in CS and FS, but not in LS and SS; however, it had a stimulatory effect on MMP–9 in CS, LS and SS. LPS had a slight stimulatory effect on MMP–2 in CS and FS, but not in LS; in addition, it stimulated MMP–9 in CS, FS and LS. EGCG and Dox without and with PMA down regulated the expression of MMP–2 and –9 in all sarcoma cell lines as did cyclohexamide, actinomycin–D, dexamethasone, and retinoic acid. NM without and with PMA inhibited MMP–2 and –9 expression in all sarcoma cell lines.

Conclusions:
Our results show that cytokines, inducers and inhibitors have an up- and down regulatory effect on MMP expression in all sarcoma cell lines, suggesting their clinical use in management of sarcomas.

Comments

Metastatic sarcoma cells secrete large amounts of MMPs, degrading the ECM and basement membrane and allowing the spread to distal organs. Although many studies have demonstrated the importance of MMPs in invasion and metastasis, little information is currently known regarding the effect of cytokines, mitogens, inducers and inhibitors. We examined the roles of cytokines, mitogens, inducers and inhibitors in regulating MMPs in adult sarcoma cell lines: human fibrosarcoma (HT-1080), chondrosarcoma (SW-1353), liposarcoma (SW-872) and synovial sarcoma (SW-982). Our results showed that cytokines, inducers and inhibitors had an up- and down regulatory effect on MMP expression in all sarcoma cell lines, suggesting their clinical use in management of sarcomas. Of interest, the micronutrient mixture (NM), without and with the inducer PMA, inhibited MMP–2 and –9 expression in all sarcoma cell lines tested. These findings are significant since MMPs, especially MMP-2 and MMP-9 play significant roles in cancer cell invasion and metastasis.

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