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- Author or Editor: Clelia Domenzain x
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Abstract
Objective—To identify extracellular proteoglycans produced by canine melanoma cell lines and analyze the effect of transforming growth factor-β1 (TGF-β1), insulin-like growth factor-I (IGF-I), and hepatocyte growth factor (HGF) on these proteoglycans.
Sample Population—3 canine melanoma cell lines (ie, CML-1, CML-6M, and CML-10c2).
Procedure—Extracellular proteoglycans were analyzed by use of metabolic labeling and western immunoblot analysis. The effect of TGF-β1 on cell proliferation was determined by incorporation of 5- bromo-2'-deoxyuridine.
Results—The CML-1 and CML-6M melanoma cell lines produced 2 main extracellular proteoglycans. One of them was identified as versican, a proteoglycan found in undifferentiated human melanoma cell lines. The CML-10c2 cells produced a small amount of extracellular proteoglycans. Addition of TGF-β1 (1.25 to 6.25 ng/ml) increased the release of sulfated proteoglycans into the medium. The TGF-β1 had mainly a posttranslational effect, because it increased the molecular mass of the sulfated bands. Addition of IGF-I (50 ng/ml) slightly increased production of proteoglycans in the CML-6M cell line, whereas HGF (50 ng/ml) did not have any effect on proteoglycan production.
Conclusions and Clinical Relevance—The proteoglycan content and response to TGF-β1 treatment for CML-1 and CML-6M canine melanoma cell lines are similar to that for undifferentiated human melanoma cell lines. In contrast, CML-10c2 cells produced a low amount of proteoglycans with high molecular weight. Because these extracellular proteoglycans are involved in the control of cell adhesion, proliferation, and migration, they may play an important role in the progression of melanomas in dogs. (Am J Vet Res 2002;63:1151–1158)
Abstract
Objective—To analyze the expression of versican and hyaluronan in melanocytomas and malignant melanomas of dogs, to correlate their expression with expression of the hyaluronan receptor CD44, and to identify enzymes responsible for the synthesis and degradation of hyaluronan in canine dermal fibroblasts and canine melanoma cell lines.
Sample Population—35 biopsy specimens from melanocytic tumors of dogs, canine primary dermal fibroblasts, and 3 canine melanoma cell lines.
Procedures—Versican, hyaluronan, and CD44 were detected in tumor samples by use of histochemical or immunohistochemical methods. Expression of hyaluronan-metabolizing enzymes was analyzed with a reverse transcriptase–PCR assay.
Results—Versican was found only in some hair follicles and around some blood vessels in normal canine skin, whereas hyaluronan was primarily found within the dermis. Hyaluronan was found in connective tissue of the oral mucosa. Versican and, to a lesser extent, hyaluronan were significantly overexpressed in malignant melanomas, compared with expression in melanocytomas. No significant difference was found between malignant tumors from oral or cutaneous origin. The expression of both molecules was correlated, but hyaluronan had a more extensive distribution than versican. Versican and hyaluronan were mainly associated with tumor stroma. Canine fibroblasts and melanoma cell lines expressed hyaluronan synthase 2 and 3 (but not 1) and hyaluronidase 1 and 2.
Conclusions and Clinical Relevance—Versican may be useful as a diagnostic marker for melanocytic tumors in dogs. Knowledge of the enzymes involved in hyaluronan metabolism could reveal new potential therapeutic targets.
