Collagenase-2 (MMP-8) and collagenase-3 (MMP-13) in adult periodontitis: molecular forms and levels in gingival crevicular fluid and immunolocalisation in gingival tissue

• Published in: Journal of clinical periodontology 2002-03, Vol.29 (3), p.224-232
• Main Authors: Kiili, M., Cox, S. W., Chen, H. W., Wahlgren, J., Maisi, P., Eley, B. M., Salo, T., Sorsa, T.
• Format: Article
• Language: English
• Subjects: Adult; Blotting, Western; Chronic Disease; collagenase; Collagenases - metabolism; Dentistry; Enzyme Precursors - analysis; Female; Fibroblasts - enzymology; Gingiva - enzymology; gingival crevicular fluid; Gingival Crevicular Fluid - enzymology; Humans; Immunoenzyme Techniques; immunohistochemistry; Macrophages - enzymology; Male; matrix metalloproteinase; Matrix Metalloproteinase 13; Matrix Metalloproteinase 8 - metabolism; Middle Aged; Neutrophils - enzymology; Periodontitis - enzymology; periodontium; Plasma Cells - enzymology; Statistics, Nonparametric; western blotting
• ISSN: 0303-6979; 1600-051X
• DOI: 10.1034/j.1600-051x.2002.290308.x

Aim: To determine the cellular and molecular forms of MMP‐8 (collagenase‐2) and MMP‐13 (collagenase‐3) associated with chronic adult periodontitis by examining the species present in gingival crevicular fluid (GCF) and enzyme distribution in gingival tissue. Methods: 30‐s GCF samples were collected directly from the periodontal pockets of 12 untreated patients using filter paper strips. After elution into buffer, the samples were examined by Western immunoblotting with polyclonal antibodies for MMP‐8 and MMP‐13 and quantification by scanning image analysis. Individual band intensities were expressed as a percentage of total sample absorbance and mean patient values were calculated. Gingival tissue from 6 patients was fixed in formalin and embedded in paraffin wax. MMP‐8 and MMP‐13 were localised using the same antibodies and an avidin‐biotin‐peroxidase detecting system. Double staining was performed with a contrasting substrate reaction. Results: The majority of MMP‐8 staining in pre‐treatment GCF was present in 80, 75 and 60 kD bands corresponding to prepro‐, pro‐ and active forms of PMN‐type enzyme. 43 and 38 kD bands evidently represented active, fibroblast‐type MMP‐8. Immunoreactivities at >100 kD and 30 kD were probably enzyme‐inhibitor complex and degraded fragments, respectively. MMP‐13 was seen mainly as 60 kD proenzyme with some 40 kD active enzyme and a small proportion of >100 kD complex. The percentages of MMP‐8 PMN‐type enzyme and MMP‐13 proenzyme bands correlated significantly with gingival and bleeding indices (p