Invasion of melanoma cells into dermal connective tissue in vitro: evidence for an important role of cysteine proteases.
Invasion of melanoma cells into the dermal connective tissue is a major characteristic in the complex process of metastasis. Proteases play an important role in tumor cell invasion as these enzymes are able to degrade most components of the extracellular matrix (ECM), and thus enable cells to penetrate interstitial connective tissues and basement membranes. We developed an improved culture model that allows the detailed study of melanoma cell invasion in vitro. In this model, high (BLM) or low (530) invasive melanoma cells were seeded on the dermal side of dead deepidermized dermis (DDD) and cultured for 14 days at the air/liquid interface. The high invasive cells invaded the tissue, leading to dermal tumor formation, whereas the low invasive cells did not. Analysis of the enzymatic activity of gelatinases by in situ gelatin zymography at neutral pH revealed proteolysis only in those composites cultured with high invasive melanoma cells. Interestingly, in situ zymograms performed at more acidic conditions, favoring the activity of cysteine proteases, exhibited markedly enhanced and widespread gelatinolysis compared to neutral pH. Cysteine protease inhibitors (E-64 and leupeptin) significantly reduced invasion of melanoma cells into these composites. These results indicate an important role of cysteine proteases for tumor invasion.
Dennhöfer
Ralf
R
Department of Dermatology, University of Cologne, Cologne, Germany.
Kurschat
Peter
P
Zigrino
Paola
P
Klose
Anke
A
Bosserhoff
Anja
A
van Muijen
Goos
G
Krieg
Thomas
T
Mauch
Cornelia
C
Hunzelmann
Nicolas
N
D002403:Cathepsins / Q000037:antagonists & inhibitors; D003238:Connective Tissue / Q000473:pathology*; D003546:Cysteine Endopeptidases / Q000502:physiology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D020405:Dermis / Q000473:pathology; D005109:Extracellular Matrix / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007930:Leucine / Q000031:analogs & derivatives* / Q000494:pharmacology; D007976:Leupeptins / Q000494:pharmacology; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism / Q000494:pharmacology; D061965:Matrix Metalloproteinase Inhibitors; D008545:Melanoma / Q000201:enzymology / Q000473:pathology*; D009361:Neoplasm Invasiveness; D011480:Protease Inhibitors / Q000494:pharmacology; D012878:Skin Neoplasms / Q000201:enzymology / Q000473:pathology*; D014407:Tumor Cells, Cultured
D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors; D002403:Cathepsins; D003546:Cysteine Endopeptidases; D020778:Matrix Metalloproteinase 2; D007930:Leucine; C024974:E 64