Cartilage proteoglycan degradation by a mouse transformed macrophage cell line is mediated by macrophage metalloelastase.
OBJECTIVE
Identify and characterize the matrix metalloproteinase responsible for cartilage proteoglycan degradation mediated by a macrophage cell line in a cell culture model that resembles some aspects of rheumatoid pannus.


METHODS
Supernatants from the transformed mouse macrophage cell line J774A.1 were used to purify the proteoglycan degrading activity.


METHODS
J774A.1 macrophage culture supernatants were purified by sequential column chromatography and proteins were identified by zymography, western blotting and amino acid sequence analysis. Cartilage degradation was measured using 35S labeled bovine nasal cartilage.


RESULTS
The cartilage degrading proteolytic activity in the mouse macrophage supernatants proved to be due to two major proteins with approximate molecular masses of 48 kDa and 22 kDa that were identified as macrophage metalloelastase (MME). Incubation of purified MME at 37 degrees C for up to 16 h resulted in the processing of the 48 kDa protein to several novel bands including a previously undescribed protein of approximately 25 kDa without accumulation of fully processed 22 kDa protein. A number of proteinases increased the rate of this processing. J774A.1 macrophage metalloelastase degraded cartilage proteoglycan with an efficiency approximately equal to human macrophage metalloelastase (MMP-12) and matrilysin (MMP-7) and twice that of stromelysin-1 (MMP-3).


CONCLUSIONS
These data identify the cartilage proteoglycan degrading metalloproteinase secreted by J774A.1 macrophages in this cell culture model as MME, and describes mechanisms of activation and processing of this enzyme that may play an important role in cartilage degradation.
Janusz
M J
MJ
Hoechst Marion Roussel Pharmaceuticals Inc., Cincinnati, OH 45215, USA. janusz.mj@pg.com
Hare
M
M
Durham
S L
SL
Potempa
J
J
McGraw
W
W
Pike
R
R
Travis
J
J
Shapiro
S D
SD
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002356:Cartilage / Q000378:metabolism*; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D003238:Connective Tissue / Q000378:metabolism; D017077:Culture Media, Conditioned; D004549:Elastin / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008264:Macrophages / Q000201:enzymology* / Q000378:metabolism*; D053508:Matrix Metalloproteinase 12; D008666:Metalloendopeptidases / Q000032:analysis / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D011509:Proteoglycans / Q000378:metabolism*
D017077:Culture Media, Conditioned; D011509:Proteoglycans; D004549:Elastin; D008666:Metalloendopeptidases; C507122:MMP12 protein, human; D053508:Matrix Metalloproteinase 12