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. 2002 Jul;61(7):591–597. doi: 10.1136/ard.61.7.591

Differential expression patterns of matrix metalloproteinases and their inhibitors during development of osteoarthritis in a transgenic mouse model

H Salminen 1, A Saamanen 1, M Vankemmelbeke 1, P Auho 1, M Perala 1, E Vuorio 1
PMCID: PMC1754156  PMID: 12079898

Abstract

Objective: To characterise the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) during degeneration of articular cartilage in a transgenic Del1 mouse model for osteoarthritis.

Methods: Northern analysis was used to measure mRNA levels of MMP-2, -3, -8, -9, -13, and -14, and TIMP-1, -2, and -3 in total RNA extracted from knee joints of transgenic Del1 mice, harbouring a 15 amino acid deletion in the triple helical domain of the α1(II) collagen chain, using their non-transgenic littermates as controls. Immunohistochemistry was used to study the presence of cleavage products (neoepitopes) of type II collagen, and the distribution of MMP-13 and TIMP-1 in degenerating cartilage.

Results: Each of the MMP and TIMP mRNAs analysed exhibited distinct expression patterns during development and osteoarthritic degeneration of the knee joint. The most striking change was up regulation of MMP-13 mRNA expression in the knee joints of Del1 mice at the onset of cartilage degeneration. However, the strongest immunostaining for MMP-13 and its inhibitor TIMP-1 was not seen in the degenerating articular cartilage but in synovial tissue, deep calcified cartilage, and subchondral bone. The localisation of type II collagen neoepitopes in chondrocytes and their pericellular matrix followed a similar pattern; they were not seen in cartilage fibrillations, but in adjacent unaffected cartilage.

Conclusion: The primary localisation of MMP-13 and TIMP-1 in hyperplastic synovial tissue, subchondral bone, and calcified cartilage suggests that up regulation of MMP-13 expression during early degeneration of articular cartilage is a secondary response to cartilage erosion. This interpretation is supported by the distribution of type II collagen neoepitopes. Synovial production of MMP-13 may be related to removal of tissue debris released from articular cartilage. In the deep calcified cartilage and adjacent subchondral bone, MMP-13 probably participates in tissue remodelling.

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Figure 1 .

Figure 1

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Compiled results of Northern analyses of knee joints for MMP-2 (A), MMP-3 (B), MMP-8 (C), MMP-9 (D), MMP-13 (E), MMP-14 (F), TIMP-1 (G), TIMP-2 (H), and TIMP-3 (I) mRNA levels. Total RNAs isolated from knee joints of transgenic Del1 mice and non-transgenic controls at different ages shown below the columns (NB, newborn; d, day; m, month) were analysed by Northern hybridisation. After hybridisation with cDNA probes which detected specific mRNAs of various MMPs and TIMP-1, -2, and -3, the hybridisation intensities were quantified by phosphor imaging and normalised per amount of 28S rRNA determined by hybridisation. The results are shown as relative units. Open columns denote control samples (n=6 at each time/age group), and black columns Del1 samples (n=6 at each time/age group). Statistical significance of the difference in mRNA levels between the control and Del1 mice is shown above the columns (Student's t test; *p<0.05; **p<0.01).

Figure 2 .

Figure 2

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Immunolocalisation of type II collagen neoepitopes (A, B), MMP-13 (C, D), and TIMP-1 (E, F) during early stages of cartilage degeneration. Tissue sections from 3 month old Del1 (B, D, F) and control (A, C, E) mice. In control mice, type II collagen neoepitopes (A, arrows) and MMP-13 (C, arrows) were detected intracellularly and pericellularly in the uncalcified cartilage. In Del1 mice, MMP-13 was mainly localised in deep calcified cartilage and in subchondral bone (D, arrows). Also, TIMP-1 immunostaining was seen in the matrix of subchondral bone in Del1 mice (F). The distribution of TIMP-1 in control mice (E, arrows) resembled that of the type II collagen neoepitopes and MMP-13. Paraffin sections; avidin-biotin complex with horseradish peroxidase label and DAP detection (A, B, E, F) or alkaline phosphatase label and Fast Red detection (C, D), counterstaining with haematoxylin. Staining controls using normal rabbit serum (G) and without secondary antibody (H). The bar in panel A=25 µm.

Figure 3 .

Figure 3

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Appearance of the synovial membrane in control (A, C) and Del1 mice (B, D) at the age of 4 months. In Del1 mice, the synovial membrane of the knee joint was thickened and the immunostaining for MMP-13 (B, arrows) and TIMP-1 was increased (D, arrows) compared with the control knee joint (A, C, respectively). Paraffin sections; avidin-biotin complex with horseradish peroxidase label and DAP detection (C, D) or alkaline phosphatase label and Fast Red detection (A, B), counterstaining with haematoxylin. The bar in panel A=50 µm.

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