Immunolocalisation of collagenase in rabbit periosteal tissue explants and extraction of the enzyme. The effect of the cytokines IL-1 alpha and EGF.
The effect of interleukin-1 alpha (IL-1 alpha) and murine epidermal growth factor (EGF) on incorporation of endogenously produced collagenase in the extracellular matrix of soft connective tissue was studied in an in vitro model system using periosteal explants obtained from rabbit calvariae. Immunohistochemical analysis indicated the highest level of collagenase in explants cultured for 72 hours with IL-1 alpha in combination with EGF. Most enzyme appeared to be associated with the extracellular matrix, but labeling was also found in numerous fibroblast-like cells. Explants cultured in the presence of IL-1 alpha alone contained less enzyme and in periostea treated without cytokines, or with EGF alone, only a faint label, if any, was seen. Freshly isolated, non-cultured periostea contained no detectable enzyme. Extraction of collagenase from periostea revealed that: (1) non-cultured periosteum did not contain detectable levels of enzyme. (2) The amount of total activatable enzyme synergistically increased (10-fold) under the influence of IL-1 alpha and EGF, whereas IL-1 alpha alone showed a 4-fold enhancement compared to control or EGF-incubated explants. (3) The latent fraction of the enzyme was synergistically increased (up to 100-fold or more) in periostea cultured in the presence of IL-1 alpha + EGF (21.17 mU/explant versus 0.05 mU/explant in controls). (4) Active collagenase, on the other hand, appeared to be present in a relatively high concentration in explants cultured without cytokines (2.45 mU/explant versus 0.36 mU/explant in IL-1 alpha + EGF-treated explants).(ABSTRACT TRUNCATED AT 250 WORDS)
van der Zee
E
E
Department of Periodontology, Academic Centre for Dentistry Amsterdam (ACTA), The Netherlands.
Everts
V
V
Hoeben
K
K
Beertsen
W
W
D000818:Animals; D017364:Collagenases / Q000032:analysis* / Q000302:isolation & purification; D003238:Connective Tissue / Q000187:drug effects / Q000201:enzymology*; D003470:Culture Media / Q000737:chemistry; D004357:Drug Synergism; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005109:Extracellular Matrix / Q000187:drug effects / Q000201:enzymology*; D006909:Hydroxyproline / Q000378:metabolism; D007213:Indomethacin / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology*; D020781:Matrix Metalloproteinase 1; D009924:Organ Culture Techniques; D011817:Rabbits; D012886:Skull
D003470:Culture Media; D007375:Interleukin-1; D004815:Epidermal Growth Factor; D017364:Collagenases; D020781:Matrix Metalloproteinase 1; D006909:Hydroxyproline; D007213:Indomethacin