Objective To examine by immunohistochemistry the relative distributions of 6 matrix metalloproteinases (MMPs 1, 2, 3, 8, 9, and 13) and the 2 proinflammatory cytokines interleukin‐1β (IL‐1β) and tumor necrosis factor α (TNFα) in osteoarthritic (OA) cartilage compared with normal, age‐matched articular cartilage. Methods Articular cartilage samples were obtained from the tibial plateau of OA knees removed at arthroplasty and from normal, nonarthritic, knees obtained at autopsy. Specimens were promptly fixed in Carnoy's fixative, processed, embedded in paraffin, sectioned, and examined by immunohistochemistry for MMP and cytokine production. In addition, human articular chondrocytes (HAC) were treated in vitro with either IL‐1β, TNFα, or phorbol myristate acetate (PMA) to assess their potential to produce each of the MMPs, as determined by Western blotting and gelatin zymography. Results Immunodetection of the collagenases (MMPs 1, 8, and 13) and stromelysin 1 (MMP‐3) was demonstrated in a proportion of chondrocytes in the superficial zone of almost all of the OA specimens that had degenerative matrix changes. The gelatinases (MMPs 2 and 9) were also demonstrated by immunohistochemistry but were not so prominent. IL‐1β– and TNFα‐positive chondrocytes were also observed in a proportion of cells in the superficial zones of OA specimens. Much less immunostaining for MMPs and cytokines was observed in the deep zone of all OA specimens, where the cartilage matrix and chondrocyte morphology appeared normal. In contrast, full‐thickness normal cartilage specimens showed virtually no immunostaining for these MMPs or cytokines. Confirmation that chondrocytes can produce these 6 MMPs was obtained from HAC cultures treated with either IL‐1β, TNFα, or PMA; conditioned medium from activated HAC contained all the MMPs demonstrated by immunohistochemistry. Dual immunolocalization studies of OA cartilage specimens demonstrated the coexpression of IL‐1 with MMP‐8 by individual chondrocytes in situ. Conclusion These results indicate that the superficial zone of OA cartilage specimens, which is characterized by fibrillations, chondrocyte clusters, and degenerative matrix changes, contains a variable proportion of cells that immunostain for IL‐1β, TNFα, and 6 different MMPs. These observations support the concept that cytokine–MMP associations reflect a modified chondrocyte phenotype and an intrinsic process of cartilage degradation in OA.

Objective.To examine by immunohistochemistry the relative distributions of 6 matrix metalloproteinases (MMPs 1, 2, 3, 8, 9, and 13) and the 2 proinflammatory cytokines interleukin-1␤ (IL-1␤) and tumor necrosis factor ␣ (TNF␣) in osteoarthritic (OA) cartilage compared with normal, age-matched articular cartilage.Methods. Articular cartilage samples were obtained from the tibial plateau of OA knees removed at arthroplasty and from normal, nonarthritic, knees obtained at autopsy. Specimens were promptly fixed in Carnoy's fixative, processed, embedded in paraffin, sectioned, and examined by immunohistochemistry for MMP and cytokine production. In addition, human articular chondrocytes (HAC) were treated in vitro with either IL-1␤, TNF␣, or phorbol myristate acetate (PMA) to assess their potential to produce each of the MMPs, as determined by Western blotting and gelatin zymography.Results. Immunodetection of the collagenases (MMPs 1, 8, and 13) and stromelysin 1 (MMP-3) was demonstrated in a proportion of chondrocytes in the superficial zone of almost all of the OA specimens that had degenerative matrix changes. The gelatinases (MMPs 2 and 9) were also demonstrated by immunohistochemistry but were not so prominent. IL-1␤-and TNF␣-positive chondrocytes were also observed in a proportion of cells in the superficial zones of OA specimens. Much less immunostaining for MMPs and cytokines was observed in the deep zone of all OA specimens, where the cartilage matrix and chondrocyte morphology appeared normal. In contrast, fullthickness normal cartilage specimens showed virtually no immunostaining for these MMPs or cytokines. Confirmation that chondrocytes can produce these 6 MMPs was obtained from HAC cultures treated with either IL-1␤, TNF␣, or PMA; conditioned medium from activated HAC contained all the MMPs demonstrated by immunohistochemistry. Dual immunolocalization studies of OA cartilage specimens demonstrated the coexpression of IL-1 with MMP-8 by individual chondrocytes in situ.Conclusion. These results indicate that the superficial zone of OA cartilage specimens, which is characterized by fibrillations, chondrocyte clusters, and degenerative matrix changes, contains a variable proportion of cells that immunostain for IL-1␤, TNF␣, and 6 different MMPs. These observations support the concept that cytokine-MMP associations reflect a modified chondrocyte phenotype and an intrinsic process of cartilage degradation in OA.Osteoarthritis (OA) is the most prevalent disease of articular joints and is the major cause of disability in the elderly (1). Degeneration and loss of articular cartilage are characteristic features of OA. The appearance of fibrillations, matrix depletion, cell clusters, and changes in matrix composition reflect the aberrant behavior of resident chondrocytes (2). Although biomechanical factors are strongly implicated, at present it is unclear which stimuli regulate the hyperactive pheno-

The use of injectable pH-responsive doubly cross-linked microgels (DX microgels) to improve the mechanical properties of degenerated intervertebral discs is demonstrated for the first time. The microgel comprised methyl methacrylate (MMA), methacrylic acid (MAA), ethyleneglycol dimethacrylate (EGD) and glycidyl methacrylate (GM) and was poly(MMA/MAA/EGD)-GM. The GM facilitated covalent interparticle cross-linking. The DX microgels are shown to have tunable mechanical properties. Degeneration of model bovine intervertebral discs (IVDs) was induced using collagenase. When injected into degenerated IVDs the DX microgels were shown to improve the strain, modulus, toughness and resilience. The extent of mechanical property improvement was an increasing function of DX microgel concentration, suggesting tunability. Cytotoxicity studies showed that the DX microgel was biocompatible under the conditions investigated. The results of this study imply that injectable DX microgels have good potential as a future regenerative medicine strategy for restoring the mechanical properties of degenerated load-bearing soft tissue, such as IVDs.

The potential of various pH-responsive alkyl (meth)acrylate ester- and (meth)acrylic acid-based copolymers, including poly(methyl methacrylate-co-acrylic acid) (PMMA-AA) and poly(n-butyl acrylate-co-methacrylic acid) (PBA-MAA), to form pH-sensitive biocompatible and biodegradable hollow particle gel scaffolds for use in non-load-bearing soft tissue regeneration have been explored. The optimal copolymer design criteria for preparation of these materials have been established. Physical gels which are both pH- and redox-sensitive were formed only from PMMA-AA copolymers. MMA is the optimal hydrophobic monomer, whereas the use of various COOH-containing monomers, e.g., MAA and AA, will always induce a pH-triggered physical gelation. The PMMA-AA gels were prepared at physiological pH range from concentrated dispersions of swollen, hollow, polymer-based particles cross-linked with either cystamine (CYS) or 3,3'-dithiodipropionic acid dihydrazide (DTP). A linear relationship between particle swelling ratios, gel elasticity, and ductility was observed. The PMMA-AA gels with lower AA contents feature lower swelling ratios, mechanical strengths, and ductilities. Increasing the swelling ratio (e.g., through increasing AA content) decreased the intraparticle elasticity; however, intershell contact and gel elasticity were found to increase. The mechanical properties and performance of the gels were tuneable upon varying the copolymers' compositions and the structure of the cross-linker. Compared to PMMA-AA/CYS, the PMMA-AA/DTP gels were more elastic and ductile. The biodegradability and cytotoxicity of the new hollow particle gels were tested for the first time and related to their composition, mechanical properties, and morphology. The new PMMA-AA/CYS and PMMA-AA/DTP gels have shown good biocompatibility, biodegradability, strength, and interconnected porosity and therefore have good potential as a tissue repair agent.

Fabrication strategies for programmed hydrogels that provide precise spatial control with predetermined responses to external stimuli are highly desirable. In this study, a partially reversible light‐driven assembly (PRLDA) method is introduced to construct multiresponsive hydrogels utilizing microgel (MG) particle building blocks (swollen diameter of 107 nm). No other material is required to prepare the gels beyond the MGs themselves. Facile preparation of multiresponsive hydrogels that are reversibly responsive to light, pH, and temperature using phototriggered covalent interlinking of coumarin‐based MGs is demonstrated. The gels have phototuneable moduli and swelling ratios and show light‐assisted healing and reshaping. Remarkably, the intrinsic fluorescence of the gels undergoes a reversible light‐triggered wavelength‐shift. The emission peak blueshifted from 420 to 390 nm upon irradiation with 365 nm light. The PRLDA gels can be constructed using either positive or negative photopatterning. It is shown that the gels can be exploited for multiresponsive cytocompatible actuators, grippers, and ON/OFF circuit components as well as anticounterfeit gels. The PRLDA method provides new insight into programmed gel property control and has excellent potential for biomaterial and optoelectronic applications.