Migrating Progenitor Cells Derived From Injured Cartilage Surface Respond to Damage-Associated Molecular Patterns

Objective: To delineate the response of migrating chondrogenic progenitor cells (CPCs) that arose from the surface of mechanically injured articular cartilage to proinflammatory damage-associated-molecular-patterns (DAMPs). Design: Bovine CPCs and non-CPC chondrocytes isolated from either impacted or scratched articular cartilage were studied. Those 2 types of cells were treated with mitochondrial DAMPs (MTDs; 10 nM fMLF and 10 µg/mL CpG DNA), or 10 nM HMGB1, or 10 ng/mL IL-1b for 24 hours. At the end of experiments, conditioned media and cell lysates were collected for analysis of expression levels of matrix metalloproteinases (MMPs), chemokines, and cytokines that are associated with cartilage degeneration with Western blotting and quantitative polymerase chain reaction. The difference of expression levels was compared by Welch’s t-test. Results: Our data indicated that HMGB1 and MTDs remarkably upregulated pro-MMP-13 expression in CPCs. Compared with non-CPCs, CPCs expressed significantly more baseline mRNAs of MMP-13, CXCL12, and IL-6. MTDs greatly increased the expression of MMP-13 and IL-6 in CPCs by over 100-fold ( P < 0.001). MTDs also significantly increased IL-8 expression in CPCs to a similar extent ( P < 0.001). However, when IL-1b was present, CPCs expressed less MMP-3 and active MMP-13 proteins as well as less CCL2 and IL-6 than did non-CPCs. Conclusions: We concluded that CPCs were more sensitive than non-CPCs in response to DAMPs, especially MTDs. The proinflammatory nature of CPCs implied their critical role in the early phase of posttraumatic osteoarthritis development.

Progenitor Cells Derived From Injured Cartilage Surface Respond to Damage Associated Molecular Patterns

Objective: To elucidate how chondrogenic progenitor cells (CPCs) originated from mechanically injured cartilage surface respond to proinflammatory endogenous damage-associated-molecular-patterns (DAMPs). Design: Passage 1 bovine CPCs and non-CPCs isolated from injured articular cartilage either by blunt impaction or by scratches were treated with mitochondrial DAMPs (MTDs) composed of fMLF and CpG DNA, or HMGB1 (a nuclear DAMP), or IL-1b for 24 hrs. At the end of the experiments, the expression levels of matrix metalloproteinases (MMPs), chemokines, and cytokines that are associated with cartilage degeneration was examined with Western blotting and quantitative PCR. Results: Both HMGB1 and MTDs remarkably up-regulated expression level of pro-MMP-13 protein in CPCs while showed weak effect on non-CPCs. Compared to non-CPCs, CPCs expressed significantly higher baseline mRNA levels of MMP-13, CXCL12, and IL-6. MTDs further increased the expression levels of MMP-13 and IL-6 in CPCs while HMGB1 did not show such effect. When treated with MTDs, CPCs also expressed significantly higher levels of IL-8 mRNA than did non-CPCs. However, compared to non-CPCs, CPCs expressed much lower levels of MMP-3 and active MMP-13 proteins as well as lower mRNA levels of CCL2 and IL-6 in response to IL-1b. Conclusions: CPCs were more sensitive than non-CPCs in response to DAMPs, especially MTDs, to up-regulate expression of MMP-13, IL-6 and -8. When IL-1b was present, CPCs were less responsive than non-CPCs in terms of up-regulating MMP-3, CCL2, and IL-6 expression. The proinflammatory nature of CPCs implied their critical role in the early phase of PTOA development.

SMURF1 and SMURF2 in Progenitor Cells from Articular Cartilage and Meniscus during Late-Stage Osteoarthritis

Objective The aim of this study was to investigate the roles of SMURF1 and SMURF2 in progenitor cells from the human knee in late-stage osteoarthritis (OA). Design We applied immunohistochemistry, immunocytochemistry, RNAi, lentiviral transfection, and Western blot analysis. We obtained chondrogenic progenitor cells (CPCs) from the articular cartilage and meniscus progenitor cells (MPCs) from the nonvascularized part of the meniscus. Results SMURF1 and SMURF2 appeared in both osteoarthritic tissues. CPCs and MPCs exhibited comparable amounts of these proteins, which influence the balance between RUNX2 and SOX9. The overexpression of SMURF1 reduced the levels of RUNX2, SOX9, and TGFBR1. The overexpression of SMURF2 also reduced the levels of RUNX2 and TGFBR1, while SOX9 levels were not affected. The knockdown of SMURF1 had no effect on RUNX2, SOX9, or TGFBR1. The knockdown of SMURF2 enhanced RUNX2 and SOX9 levels in CPCs. The respective protein levels in MPCs were not affected. Conclusions This study shows that SMURF1 and SMURF2 are regulatory players for the expression of the major regulator transcription factors RUNX2 and SOX9 in CPCs and MPCs. Our novel findings may help elucidate new treatment strategies for cartilage regeneration.