Inhibition of cartilage degradation and changes in physical properties induced by IL-1beta and retinoic acid using matrix metalloproteinase inhibitors.
Bovine cartilage explants were treated with 100 ng/ml recombinant human interleukin-1beta (IL-1beta) or 1 microM all-trans retinoic acid (RA) and changes in biochemical, biomechanical, and physicochemical properties were assessed. Additionally, samples cultured with IL-1beta or RA were treated with 4 microM recombinant human tissue inhibitor of metalloproteinases-1 (TIMP-1) or a synthetic metalloproteinase inhibitor (L-758,354) to inhibit this degradation. Treatment with IL-1beta or RA each resulted in >90% GAG loss after 8 days in culture. Addition of TIMP or L-758,354 to the culture media inhibited IL-1beta-induced loss of tissue GAG by 40 and 65%, respectively, and inhibited RA-induced GAG loss by 35 and 65%, respectively. Analysis of degradation products in the culture media using a G1 antibody indicated that IL-1beta- and RA-treated plugs released 68-kDa fragments of aggrecan, corresponding to a segment of the aggrecan core protein from the G1 domain to the C-terminus NITEGE, consistent with "aggrecanase" activity. Release of the G1 fragment was inhibited by treatment with L-758,354. Both IL-1beta and RA induced significant loss of hyaluronan from cartilage explants after 8 days of exposure and HA loss was also inhibited by addition of L-756,354 to the culture media. IL-1beta, but not RA, induced a significant increase in swelling ratio (wet weight in 0.01 M NaCl normalized to wet weight in DMEM) after 8 days in culture, consistent with degradation of the collagen network, and the increase in tissue swelling was inhibited by treatment with TIMP-1 or L-758,354. Exposure to IL-1beta or RA resulted in significant changes in cartilage physical properties including streaming potential, equilibrium modulus, hydraulic permeability, and electrokinetic coupling coefficient after 8 days in culture, and these changes were inhibited by 40-90% by exposure to TIMP and by 50-90% by exposure to L-758,354. Measurement of dynamic streaming potential showed that changes due to treatment with IL-1beta alone were highly dependent in compression frequency, with dramatic changes seen at high frequency prior to changes in mechanical properties, and little initial change seen at low frequency. Streaming potential and equilibrium modulus of explants treated with RA decreased to 10% of their initial values after 8 days in culture, but decreased to only 40 and 90%, respectively, when treated with RA plus TIMP-1.
Bonassar
L J
LJ
Department of Electrical Engineering and Computer Science, Center for Biomedical Engineering, Massachusetts Institute of Technology, Cambridge 02139, USA.
Sandy
J D
JD
Lark
M W
MW
Plaas
A H
AH
Frank
E H
EH
Grodzinsky
A J
AJ
D053674:Aggrecans; D000818:Animals; D001696:Biomechanical Phenomena; D002356:Cartilage / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D002417:Cattle; D002809:Chondroitin Sulfates / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D046508:Culture Techniques; D004151:Dipeptides / Q000494:pharmacology; D004594:Electrophysiology; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins / Q000235:genetics / Q000494:pharmacology*; D006025:Glycosaminoglycans / Q000378:metabolism; D006820:Hyaluronic Acid / Q000378:metabolism; D007375:Interleukin-1 / Q000494:pharmacology*; D037181:Lectins, C-Type; D020781:Matrix Metalloproteinase 1; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D019714:Tissue Inhibitor of Metalloproteinases; D014212:Tretinoin / Q000494:pharmacology*
D053674:Aggrecans; D004151:Dipeptides; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006025:Glycosaminoglycans; D007375:Interleukin-1; C105326:L 758354; D037181:Lectins, C-Type; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors; D011509:Proteoglycans; D011994:Recombinant Proteins; D019714:Tissue Inhibitor of Metalloproteinases; D014212:Tretinoin; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D003094:Collagen; D020781:Matrix Metalloproteinase 1