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MMP13 is a critical target gene during the progression of osteoarthritis.

Wang M, Sampson ER, Jin H, Li J, Ke QH, Im HJ, Chen D - Arthritis Res. Ther. (2013)

Bottom Line: Currently there is no effective disease-modifying treatment for OA.Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.Mmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Osteoarthritis (OA) is a degenerative joint disease affecting a large population of people. The mechanism of this highly prevalent disease is not fully understood. Currently there is no effective disease-modifying treatment for OA. The purpose of this study was two-fold: 1) to investigate the role of MMP13 in the development of OA; and 2) to evaluate the efficacy of the MMP13 inhibitor CL82198 as a pharmacologic treatment for preventing OA progression.

Methods: To investigate the role of the endogenous Mmp13 gene in OA development, tamoxifen was administered to two-week-old Col2CreER;Mmp13fx/fx (Mmp13Col2ER) and Cre-negative control mice for five days. OA was induced by meniscal-ligamentous injury (MLI) when the mice were 10 weeks old and MLI or sham-operated joints were harvested 4, 8, 12, or 16 weeks after surgery. To evaluate the efficacy of CL82198, MLI surgery was performed on 10-week-old wild type mice. CL82198 or saline was administered to the mice daily beginning immediately after the surgery for up to 16 weeks. The joint tissues collected from both experiments were evaluated by cartilage grading, histology/histomorphometry, immunohistochemistry (IHC), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The ability of CL82198 to inhibit MMP13 activity in vitro was confirmed by ELISA.

Results: The OA progression was decelerated in Mmp13Col2ER mice 8, 12, and 16 weeks post-surgery. Cartilage grading by blinded observers confirmed decreased articular cartilage degeneration in Mmp13Col2ER mice at 8, 12 and 16 weeks compared to Cre-negative mice. Histomorphometric analysis demonstrated that Mmp13Col2ER mice had a higher articular cartilage area and thickness at 12 and 16 weeks post-surgery compared to the control mice. Results of IHC revealed greater type II collagen and proteoglycan expression in Mmp13Col2ER mice. Chondrocyte apoptosis, as determined by TUNEL staining, was higher in control mice compared to Mmp13Col2ER mice. CL82198 inhibited MMP13 activity in conditioned media from vehicle (>85%) or bone morphogenetic protein 2 (BMP2)-treated (>90%) primary murine sternal chondrocytes. Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.

Conclusions: Mmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.

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Decelerated osteoarthritis progression in Mmp13Col2ER mice. Tamoxifen was administered when matrix metalloproteinase (MMP13) conditional knockout (cKO) mice (Mmp13Col2ER) and Cre-negative control mice were two weeks old (1 mg/10 g body weight, i.p., daily for five days). Meniscal-ligamentous injury (MLI)-induced osteoarthritis (OA) surgery was performed on the right hind-limbs when the mice were 10 weeks old. The left hind-limbs were used as sham controls. (A) Knee joint samples were harvested 4, 8, 12, or 16 weeks post-surgery and Alcian blue/Hematoxylin/Orange G staining was performed. Histological results showed decreased articular cartilage degradation in Mmp13Col2ER mice 8, 12 and 16 weeks post-surgery. (B) Histological grading by blinded observers confirmed decreased articular cartilage degradation in Mmp13Col2ER mice at 8, 12, and 16 weeks compared to control Cre-negative mice (*P < 0.05). (C) Tibia articular cartilage area was quantified by tracing the Alcian blue-positive area in the proximal tibia. There was no significant difference 4 and 8 weeks post-surgery in Mmp13Col2ER MLI mice versus control MLI mice. The tibia cartilage area was increased 21% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 31% 16 weeks post-surgery (*P < 0.05). (D) Tibia thickness was quantified by tracing the Alcian blue-positive thickness in the center of the tibial plateau. There was no significant difference in tibial cartilage thickness at 4 or 8 weeks post-surgery. Tibia cartilage thickness was increased 29% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 50% 16 weeks post-surgery (*P < 0.01). (E) Total articular cartilage area was quantified by tracing the Alcian blue-positive area in both the proximal tibia and distal femur. No significant differences were detected 4, 8, and 12 weeks post-surgery. Total cartilage area increased 18% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01). (F) Total cartilage thickness was quantified by tracing the Alcian blue-positive thickness in the center of the proximal tibia and distal femur. No significant differences were detected at 4, 8, and 12 weeks post-surgery. Total cartilage thickness increased 39% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01).
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Figure 1: Decelerated osteoarthritis progression in Mmp13Col2ER mice. Tamoxifen was administered when matrix metalloproteinase (MMP13) conditional knockout (cKO) mice (Mmp13Col2ER) and Cre-negative control mice were two weeks old (1 mg/10 g body weight, i.p., daily for five days). Meniscal-ligamentous injury (MLI)-induced osteoarthritis (OA) surgery was performed on the right hind-limbs when the mice were 10 weeks old. The left hind-limbs were used as sham controls. (A) Knee joint samples were harvested 4, 8, 12, or 16 weeks post-surgery and Alcian blue/Hematoxylin/Orange G staining was performed. Histological results showed decreased articular cartilage degradation in Mmp13Col2ER mice 8, 12 and 16 weeks post-surgery. (B) Histological grading by blinded observers confirmed decreased articular cartilage degradation in Mmp13Col2ER mice at 8, 12, and 16 weeks compared to control Cre-negative mice (*P < 0.05). (C) Tibia articular cartilage area was quantified by tracing the Alcian blue-positive area in the proximal tibia. There was no significant difference 4 and 8 weeks post-surgery in Mmp13Col2ER MLI mice versus control MLI mice. The tibia cartilage area was increased 21% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 31% 16 weeks post-surgery (*P < 0.05). (D) Tibia thickness was quantified by tracing the Alcian blue-positive thickness in the center of the tibial plateau. There was no significant difference in tibial cartilage thickness at 4 or 8 weeks post-surgery. Tibia cartilage thickness was increased 29% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 50% 16 weeks post-surgery (*P < 0.01). (E) Total articular cartilage area was quantified by tracing the Alcian blue-positive area in both the proximal tibia and distal femur. No significant differences were detected 4, 8, and 12 weeks post-surgery. Total cartilage area increased 18% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01). (F) Total cartilage thickness was quantified by tracing the Alcian blue-positive thickness in the center of the proximal tibia and distal femur. No significant differences were detected at 4, 8, and 12 weeks post-surgery. Total cartilage thickness increased 39% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01).

Mentions: To investigate if Mmp13 deletion could prevent or decelerate MLI-induced OA, we crossed Col2CreER transgenic mice [14,16] with Mmp13fx/fx mice [13] to generate Col2CreER;Mmp13fx/fx mice (Mmp13Col2ER) (Table 1). Deletion of the Mmp13 gene in chondrocytes had no significant effect on articular and growth plate chondrocyte morphology [see Additional file 1]. Tamoxifen was administered (i.p., five days) to two-week-old Mmp13Col2ER mice and had no effect on articular and growth plate cartilage [see Additional file 1]. MLI surgery was performed when the mice were 10-weeks-old to induce OA. Knee joints were harvested 4, 8, 12, and 16 weeks post-surgery (n = 5 in each group). Histology showed that four weeks post-MLI surgery, articular cartilage was nearly normal in both Mmp13Col2ER and Cre-negative control groups (Figure 1A). OA-like fibrillation, clefting and cartilage loss down to the tidemark appeared 8 weeks post-surgery and worsened at the 12- and 16-week time points in Cre-negative control mice. In Mmp13Col2ER mice, however, there was markedly less articular cartilage excavation, especially at the 12- and 16-week time points (Figure 1A). OA grading likewise revealed significantly reduced cartilage degeneration at 8, 12 and 16 weeks post-surgery in Mmp13Col2ER mice compared to the control group (Figure 1B). To quantify OA progression, we performed histomorphometry using the OsteoMeasure system. The results showed that articular cartilage area and articular cartilage thickness at proximal tibiae, total cartilage area, and total cartilage thickness progressively decreased at 8, 12, and 16 weeks post-surgery (Figure 1C-F), but this progression was decelerated in Mmp13Col2ER mice compared to the control group. Articular cartilage area and articular cartilage thickness at proximal tibiae were significantly greater at 12 and 16 weeks post-surgery in Mmp13Col2ER groups (Figure 1C-D). Total cartilage area was significantly different at 16 weeks post-surgery (Figure 1E), and total cartilage thickness was significantly different at 8, 12, and 16 weeks post-surgery (Figure 1F). Articular cartilage area and thickness at distal femora had no significant changes at any time point post-surgery.


MMP13 is a critical target gene during the progression of osteoarthritis.

Wang M, Sampson ER, Jin H, Li J, Ke QH, Im HJ, Chen D - Arthritis Res. Ther. (2013)

Decelerated osteoarthritis progression in Mmp13Col2ER mice. Tamoxifen was administered when matrix metalloproteinase (MMP13) conditional knockout (cKO) mice (Mmp13Col2ER) and Cre-negative control mice were two weeks old (1 mg/10 g body weight, i.p., daily for five days). Meniscal-ligamentous injury (MLI)-induced osteoarthritis (OA) surgery was performed on the right hind-limbs when the mice were 10 weeks old. The left hind-limbs were used as sham controls. (A) Knee joint samples were harvested 4, 8, 12, or 16 weeks post-surgery and Alcian blue/Hematoxylin/Orange G staining was performed. Histological results showed decreased articular cartilage degradation in Mmp13Col2ER mice 8, 12 and 16 weeks post-surgery. (B) Histological grading by blinded observers confirmed decreased articular cartilage degradation in Mmp13Col2ER mice at 8, 12, and 16 weeks compared to control Cre-negative mice (*P < 0.05). (C) Tibia articular cartilage area was quantified by tracing the Alcian blue-positive area in the proximal tibia. There was no significant difference 4 and 8 weeks post-surgery in Mmp13Col2ER MLI mice versus control MLI mice. The tibia cartilage area was increased 21% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 31% 16 weeks post-surgery (*P < 0.05). (D) Tibia thickness was quantified by tracing the Alcian blue-positive thickness in the center of the tibial plateau. There was no significant difference in tibial cartilage thickness at 4 or 8 weeks post-surgery. Tibia cartilage thickness was increased 29% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 50% 16 weeks post-surgery (*P < 0.01). (E) Total articular cartilage area was quantified by tracing the Alcian blue-positive area in both the proximal tibia and distal femur. No significant differences were detected 4, 8, and 12 weeks post-surgery. Total cartilage area increased 18% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01). (F) Total cartilage thickness was quantified by tracing the Alcian blue-positive thickness in the center of the proximal tibia and distal femur. No significant differences were detected at 4, 8, and 12 weeks post-surgery. Total cartilage thickness increased 39% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01).
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Figure 1: Decelerated osteoarthritis progression in Mmp13Col2ER mice. Tamoxifen was administered when matrix metalloproteinase (MMP13) conditional knockout (cKO) mice (Mmp13Col2ER) and Cre-negative control mice were two weeks old (1 mg/10 g body weight, i.p., daily for five days). Meniscal-ligamentous injury (MLI)-induced osteoarthritis (OA) surgery was performed on the right hind-limbs when the mice were 10 weeks old. The left hind-limbs were used as sham controls. (A) Knee joint samples were harvested 4, 8, 12, or 16 weeks post-surgery and Alcian blue/Hematoxylin/Orange G staining was performed. Histological results showed decreased articular cartilage degradation in Mmp13Col2ER mice 8, 12 and 16 weeks post-surgery. (B) Histological grading by blinded observers confirmed decreased articular cartilage degradation in Mmp13Col2ER mice at 8, 12, and 16 weeks compared to control Cre-negative mice (*P < 0.05). (C) Tibia articular cartilage area was quantified by tracing the Alcian blue-positive area in the proximal tibia. There was no significant difference 4 and 8 weeks post-surgery in Mmp13Col2ER MLI mice versus control MLI mice. The tibia cartilage area was increased 21% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 31% 16 weeks post-surgery (*P < 0.05). (D) Tibia thickness was quantified by tracing the Alcian blue-positive thickness in the center of the tibial plateau. There was no significant difference in tibial cartilage thickness at 4 or 8 weeks post-surgery. Tibia cartilage thickness was increased 29% in Mmp13Col2ER MLI mice compared to control MLI mice 12 weeks post-surgery (*P < 0.05) and increased 50% 16 weeks post-surgery (*P < 0.01). (E) Total articular cartilage area was quantified by tracing the Alcian blue-positive area in both the proximal tibia and distal femur. No significant differences were detected 4, 8, and 12 weeks post-surgery. Total cartilage area increased 18% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01). (F) Total cartilage thickness was quantified by tracing the Alcian blue-positive thickness in the center of the proximal tibia and distal femur. No significant differences were detected at 4, 8, and 12 weeks post-surgery. Total cartilage thickness increased 39% in Mmp13Col2ER MLI mice compared to control MLI mice 16 weeks post-surgery (*P < 0.01).
Mentions: To investigate if Mmp13 deletion could prevent or decelerate MLI-induced OA, we crossed Col2CreER transgenic mice [14,16] with Mmp13fx/fx mice [13] to generate Col2CreER;Mmp13fx/fx mice (Mmp13Col2ER) (Table 1). Deletion of the Mmp13 gene in chondrocytes had no significant effect on articular and growth plate chondrocyte morphology [see Additional file 1]. Tamoxifen was administered (i.p., five days) to two-week-old Mmp13Col2ER mice and had no effect on articular and growth plate cartilage [see Additional file 1]. MLI surgery was performed when the mice were 10-weeks-old to induce OA. Knee joints were harvested 4, 8, 12, and 16 weeks post-surgery (n = 5 in each group). Histology showed that four weeks post-MLI surgery, articular cartilage was nearly normal in both Mmp13Col2ER and Cre-negative control groups (Figure 1A). OA-like fibrillation, clefting and cartilage loss down to the tidemark appeared 8 weeks post-surgery and worsened at the 12- and 16-week time points in Cre-negative control mice. In Mmp13Col2ER mice, however, there was markedly less articular cartilage excavation, especially at the 12- and 16-week time points (Figure 1A). OA grading likewise revealed significantly reduced cartilage degeneration at 8, 12 and 16 weeks post-surgery in Mmp13Col2ER mice compared to the control group (Figure 1B). To quantify OA progression, we performed histomorphometry using the OsteoMeasure system. The results showed that articular cartilage area and articular cartilage thickness at proximal tibiae, total cartilage area, and total cartilage thickness progressively decreased at 8, 12, and 16 weeks post-surgery (Figure 1C-F), but this progression was decelerated in Mmp13Col2ER mice compared to the control group. Articular cartilage area and articular cartilage thickness at proximal tibiae were significantly greater at 12 and 16 weeks post-surgery in Mmp13Col2ER groups (Figure 1C-D). Total cartilage area was significantly different at 16 weeks post-surgery (Figure 1E), and total cartilage thickness was significantly different at 8, 12, and 16 weeks post-surgery (Figure 1F). Articular cartilage area and thickness at distal femora had no significant changes at any time point post-surgery.

Bottom Line: Currently there is no effective disease-modifying treatment for OA.Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.Mmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Osteoarthritis (OA) is a degenerative joint disease affecting a large population of people. The mechanism of this highly prevalent disease is not fully understood. Currently there is no effective disease-modifying treatment for OA. The purpose of this study was two-fold: 1) to investigate the role of MMP13 in the development of OA; and 2) to evaluate the efficacy of the MMP13 inhibitor CL82198 as a pharmacologic treatment for preventing OA progression.

Methods: To investigate the role of the endogenous Mmp13 gene in OA development, tamoxifen was administered to two-week-old Col2CreER;Mmp13fx/fx (Mmp13Col2ER) and Cre-negative control mice for five days. OA was induced by meniscal-ligamentous injury (MLI) when the mice were 10 weeks old and MLI or sham-operated joints were harvested 4, 8, 12, or 16 weeks after surgery. To evaluate the efficacy of CL82198, MLI surgery was performed on 10-week-old wild type mice. CL82198 or saline was administered to the mice daily beginning immediately after the surgery for up to 16 weeks. The joint tissues collected from both experiments were evaluated by cartilage grading, histology/histomorphometry, immunohistochemistry (IHC), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The ability of CL82198 to inhibit MMP13 activity in vitro was confirmed by ELISA.

Results: The OA progression was decelerated in Mmp13Col2ER mice 8, 12, and 16 weeks post-surgery. Cartilage grading by blinded observers confirmed decreased articular cartilage degeneration in Mmp13Col2ER mice at 8, 12 and 16 weeks compared to Cre-negative mice. Histomorphometric analysis demonstrated that Mmp13Col2ER mice had a higher articular cartilage area and thickness at 12 and 16 weeks post-surgery compared to the control mice. Results of IHC revealed greater type II collagen and proteoglycan expression in Mmp13Col2ER mice. Chondrocyte apoptosis, as determined by TUNEL staining, was higher in control mice compared to Mmp13Col2ER mice. CL82198 inhibited MMP13 activity in conditioned media from vehicle (>85%) or bone morphogenetic protein 2 (BMP2)-treated (>90%) primary murine sternal chondrocytes. Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.

Conclusions: Mmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.

Show MeSH
Related in: MedlinePlus