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Knee loading reduces MMP13 activity in the mouse cartilage.

Hamamura K, Zhang P, Zhao L, Shim JW, Chen A, Dodge TR, Wan Q, Shih H, Na S, Lin CC, Sun HB, Yokota H - BMC Musculoskelet Disord (2013)

Bottom Line: Load-driven reduction in MMP13 was associated with a decrease in the phosphorylation level of p38 MAPK (p-p38) and NFκB (p-NFκB).Silencing Rac1 GTPase significantly reduced MMP13 expression and p-p38 but not p-NFκB.Knee loading reduces MMP13 activity at least in part through Rac1-mediated p38 MAPK signaling.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biomedical Engineering, Indiana University Purdue University Indianapolis, SL220C, 723 West Michigan Street, Indianapolis, IN 46202, USA. hyokota@iupui.edu.

ABSTRACT

Background: Moderate loads with knee loading enhance bone formation, but its effects on the maintenance of the knee are not well understood. In this study, we examined the effects of knee loading on the activity of matrix metalloproteinase13 (MMP13) and evaluated the role of p38 MAPK and Rac1 GTPase in the regulation of MMP13.

Methods: Knee loading (0.5-3 N for 5 min) was applied to the right knee of surgically-induced osteoarthritis (OA) mice as well as normal (non-OA) mice, and MMP13 activity in the femoral cartilage was examined. The sham-loaded knee was used as a non-loading control. We also employed primary non-OA and OA human chondrocytes as well as C28/I2 chondrocyte cells, and examined MMP13 activity and molecular signaling in response to shear at 2-20 dyn/cm².

Results: Daily knee loading at 1 N for 2 weeks suppressed cartilage destruction in the knee of OA mice. Induction of OA elevated MMP13 activity and knee loading at 1 N suppressed this elevation. MMP13 activity was also increased in primary OA chondrocytes, and this increase was attenuated by applying shear at 10 dyn/cm². Load-driven reduction in MMP13 was associated with a decrease in the phosphorylation level of p38 MAPK (p-p38) and NFκB (p-NFκB). Molecular imaging using a fluorescence resonance energy transfer (FRET) technique showed that Rac1 activity was reduced by shear at 10 dyn/cm² and elevated by it at 20 dyn/cm². Silencing Rac1 GTPase significantly reduced MMP13 expression and p-p38 but not p-NFκB. Transfection of a constitutively active Rac1 GTPase mutant increased MMP13 activity, while a dominant negative mutant decreased it.

Conclusions: Knee loading reduces MMP13 activity at least in part through Rac1-mediated p38 MAPK signaling. This study suggests the possibility of knee loading as a therapy not only for strengthening bone but also preventing tissue degradation of the femoral cartilage.

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Knee loading and histological observation of the mouse knee. (A) Force profiles for knee loading. A 5 Hz sinusoidal load was applied to the right knee at a magnitude of 0.5 N, 1.0 N, or 3.0 N (peak-to-peak). Sham-loaded controls received a static load of 0.5 N. (B) Force-voltage relationship. To verify peak forces applied to the knee, the loading device was calibrated using a load cell and a linear relationship was defined relating actuator voltage and peak compressive force. (C) Application of knee loading. A custom-made piezoelectric loader was utilized to load the mouse knee.
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Figure 1: Knee loading and histological observation of the mouse knee. (A) Force profiles for knee loading. A 5 Hz sinusoidal load was applied to the right knee at a magnitude of 0.5 N, 1.0 N, or 3.0 N (peak-to-peak). Sham-loaded controls received a static load of 0.5 N. (B) Force-voltage relationship. To verify peak forces applied to the knee, the loading device was calibrated using a load cell and a linear relationship was defined relating actuator voltage and peak compressive force. (C) Application of knee loading. A custom-made piezoelectric loader was utilized to load the mouse knee.

Mentions: Animal procedures were approved by the Indiana University IACUC. Fifty-seven C57/BL/6 female mice (~12 weeks, Harlan Laboratories) were used. OA in the right knee was surgically induced by transecting the medial collateral ligament and removing the medial meniscus [17]. Using the procedure previously described [6-8], knee loading was applied to the right limb using a piezoelectric loader (Figures 1A-C). The loading device was calibrated using a load cell (Model LLB130, FUTEK Advanced Sensor Technology, Irvine, CA) to determine peak compressive force for increasing actuator voltage. Pre-load was established at 0.5 N, and peak forces during three trials at each of six input voltages were averaged using manufacturer-provided software to establish a linear equation relating actuator voltage and peak compressive force. The mouse was anesthetized using 2% isoflurane and lateral dynamic loads to the knee were applied for 5 min. Loads were sinusoidal at 0.5, 1, or 3 N (peak-to-peak) and the frequency was 5 Hz. The anesthetized mice placed on the loading device were used as the sham-loaded control. The femoral cartilage was harvested 1 h after the loading bout.


Knee loading reduces MMP13 activity in the mouse cartilage.

Hamamura K, Zhang P, Zhao L, Shim JW, Chen A, Dodge TR, Wan Q, Shih H, Na S, Lin CC, Sun HB, Yokota H - BMC Musculoskelet Disord (2013)

Knee loading and histological observation of the mouse knee. (A) Force profiles for knee loading. A 5 Hz sinusoidal load was applied to the right knee at a magnitude of 0.5 N, 1.0 N, or 3.0 N (peak-to-peak). Sham-loaded controls received a static load of 0.5 N. (B) Force-voltage relationship. To verify peak forces applied to the knee, the loading device was calibrated using a load cell and a linear relationship was defined relating actuator voltage and peak compressive force. (C) Application of knee loading. A custom-made piezoelectric loader was utilized to load the mouse knee.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3924329&req=5

Figure 1: Knee loading and histological observation of the mouse knee. (A) Force profiles for knee loading. A 5 Hz sinusoidal load was applied to the right knee at a magnitude of 0.5 N, 1.0 N, or 3.0 N (peak-to-peak). Sham-loaded controls received a static load of 0.5 N. (B) Force-voltage relationship. To verify peak forces applied to the knee, the loading device was calibrated using a load cell and a linear relationship was defined relating actuator voltage and peak compressive force. (C) Application of knee loading. A custom-made piezoelectric loader was utilized to load the mouse knee.
Mentions: Animal procedures were approved by the Indiana University IACUC. Fifty-seven C57/BL/6 female mice (~12 weeks, Harlan Laboratories) were used. OA in the right knee was surgically induced by transecting the medial collateral ligament and removing the medial meniscus [17]. Using the procedure previously described [6-8], knee loading was applied to the right limb using a piezoelectric loader (Figures 1A-C). The loading device was calibrated using a load cell (Model LLB130, FUTEK Advanced Sensor Technology, Irvine, CA) to determine peak compressive force for increasing actuator voltage. Pre-load was established at 0.5 N, and peak forces during three trials at each of six input voltages were averaged using manufacturer-provided software to establish a linear equation relating actuator voltage and peak compressive force. The mouse was anesthetized using 2% isoflurane and lateral dynamic loads to the knee were applied for 5 min. Loads were sinusoidal at 0.5, 1, or 3 N (peak-to-peak) and the frequency was 5 Hz. The anesthetized mice placed on the loading device were used as the sham-loaded control. The femoral cartilage was harvested 1 h after the loading bout.

Bottom Line: Load-driven reduction in MMP13 was associated with a decrease in the phosphorylation level of p38 MAPK (p-p38) and NFκB (p-NFκB).Silencing Rac1 GTPase significantly reduced MMP13 expression and p-p38 but not p-NFκB.Knee loading reduces MMP13 activity at least in part through Rac1-mediated p38 MAPK signaling.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biomedical Engineering, Indiana University Purdue University Indianapolis, SL220C, 723 West Michigan Street, Indianapolis, IN 46202, USA. hyokota@iupui.edu.

ABSTRACT

Background: Moderate loads with knee loading enhance bone formation, but its effects on the maintenance of the knee are not well understood. In this study, we examined the effects of knee loading on the activity of matrix metalloproteinase13 (MMP13) and evaluated the role of p38 MAPK and Rac1 GTPase in the regulation of MMP13.

Methods: Knee loading (0.5-3 N for 5 min) was applied to the right knee of surgically-induced osteoarthritis (OA) mice as well as normal (non-OA) mice, and MMP13 activity in the femoral cartilage was examined. The sham-loaded knee was used as a non-loading control. We also employed primary non-OA and OA human chondrocytes as well as C28/I2 chondrocyte cells, and examined MMP13 activity and molecular signaling in response to shear at 2-20 dyn/cm².

Results: Daily knee loading at 1 N for 2 weeks suppressed cartilage destruction in the knee of OA mice. Induction of OA elevated MMP13 activity and knee loading at 1 N suppressed this elevation. MMP13 activity was also increased in primary OA chondrocytes, and this increase was attenuated by applying shear at 10 dyn/cm². Load-driven reduction in MMP13 was associated with a decrease in the phosphorylation level of p38 MAPK (p-p38) and NFκB (p-NFκB). Molecular imaging using a fluorescence resonance energy transfer (FRET) technique showed that Rac1 activity was reduced by shear at 10 dyn/cm² and elevated by it at 20 dyn/cm². Silencing Rac1 GTPase significantly reduced MMP13 expression and p-p38 but not p-NFκB. Transfection of a constitutively active Rac1 GTPase mutant increased MMP13 activity, while a dominant negative mutant decreased it.

Conclusions: Knee loading reduces MMP13 activity at least in part through Rac1-mediated p38 MAPK signaling. This study suggests the possibility of knee loading as a therapy not only for strengthening bone but also preventing tissue degradation of the femoral cartilage.

Show MeSH
Related in: MedlinePlus