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The CDC (2025) reported Osteoarthritis (OA) affects 33 million US adults. We are characterizing normal and OA human articular cartilage to advance tissue‑engineering and cartilage regeneration strategies. We are: (1) generating layer‑resolved bulk RNA‑seq and spatial transcriptomic (GeoMx DSP) profiles; 2) characterizing zonal protein expression by immunostaining; and (3) comparing transcriptomic signatures of normal/OA cartilage to mesenchymal stromal cell (MSC)–derived hyaline‑like cartilage constructs cultured within scaffolds in a sheared environment while adding oscillating hydrostatic pressure (OHP) in a novel bioreactor. 

Abstract Differentiating mesenchymal stromal cells into healthy chondrocytes for articular cartilage (AC) requires a variety of biochemical and biomechanical stimuli. While the effects of such stimuli on chondrogenic markers are well-studied, no bioreactor has thus far collected real-time data from mRNA expression for use in closed-loop control. Here, we present the design of an automated bioreactor that applies perfusion, oscillating hydrostatic pressure (OHP), and varying concentrations of multiple biochemical growth factors to the cell cultures while measuring fluorescent markers of gene expression in real-time. Our system achieves a gradated hydrodynamic environment across the length of cell cultures with flow rates between 0.46 and 3.2 cm s⁻¹and shear stresses between 5.0 and 60 mPa, can apply OHP at up to 0.5 Hz and 4.83 MPa, and can dynamically control the concentrations of two biochemical growth factors. The integrated fluorescent fiberscope system can measure fluorescent dye concentrations as low as 3 nm within the cell chamber. Our LNA/DNA nano-biosensor is designed to track Sox9 mRNA, a gene critical to the chondrogenic process. The reactor is controlled via MATLAB Simulink and includes remote observation and control features to increase flexibility and minimize downtime. This novel bioreactor provides a platform for further AC research with the ultimate design goal of generating a transfer function that maps growth factor inputs to mRNA expression outputs for real-time control. 

Recreating articular cartilage tri-layered patterning for an engineered in vitro cell construct holds promise for advancing cartilage repair efforts. Our approach involves the development of a mul-tichambered perfusion tissue bioreactor that regulates fluid shear stress levels similar to the gradated hydrodynamic environment in articular cartilage. COMSOL modeling reveals our ta-pered cell chamber design will produce three different shear levels, high in the 22 – 41 mPa range, medium in the 4.5 – 8.4 mPa range, and low in the 2.2 – 3.8 mPa range and distributed across the surface of our mesenchymal stromal cell (MSC) encapsulated construct. In a 14-day bioreactor culture, we assess how fluid shear magnitude and cell vertical location within a 3D construct influence cell chondrogenesis. Notably, Sox9 expression for MSCs cultivated in our reactor shows spatially patterned gene upregulations coding for key chondrogenic marker pro-teins. Beginning with the high shear stress region, lubricin and type II collagen gene increases of 410 and 370-fold indicate cell movement towards a superficial zone architype which is further supported by histological and immunohistochemical stains illustrating the formation of a dense proteoglycan matrix enriched with lubricin, versican, and collagen types I and II molecules. For the medium shear stress region high aggrecan and type II collagen gene expressions of 2.3 and 400-fold, respectively, along with high proteoglycan analyses show movement toward a superfi-cial/mid-zone cartilage architype. For low shear stress regions higher collagen types II and X gene upregulations of 550 and 8,300-fold, the latter being 2x of that for the high shear regime, indicate cell movement with deep zone characteristics. Collectively, biochemical analysis, histol-ogy, and gene expression data demonstrated that our fluid shear bioreactor induced a stratified structure within tissue engineered constructs, demonstrating the feasibility of using this ap-proach to recapitulate the structure of native articular cartilage. 

Abstract Post‐traumatic osteoarthritis develops following an inciting injury to a joint and results in cartilage degeneration. Mechanical loading, including articulation, drives anabolic responses in cartilage clinically, in vivo, and in vitro. Tribological articulation, or sliding of cartilage on a glass counterface, has long been used as an in vitro tool to study cartilage tissue behavior. However, it is unclear if tribological articulation affects chondrocyte fate following injury, and if the timing of articulation impacts the resultant effect. The goal of this study was to investigate the effect of tribological articulation on injured cartilage tissue at two time points: (i) performed immediately after injury and (ii) 24 h after injury. Neonatal bovine femoral cartilage explants were injured using a rapid spring‐loaded impactor and subsequently subjected to tribological articulation. Cell death due to impact injury was highest near the articular surface, suggesting a strain‐dependent mechanism. Immediate articulation following injury mitigated cell death compared to injury alone or delayed articulation; markers for both general cell death and early‐stage apoptosis were markedly decreased in the explants that were immediately slid. Interestingly, mitigation of cell death due to sliding was most predominant at the cartilage surface. Tribological articulation is known to create fluid flow within the tissue, predominantly at the articular surface, which could drive the protective response seen here. Altogether, this work shows that perturbations to the cellular environment immediately following cartilage injury significantly impact chondrocyte fate. 

Articular cartilage is a load-bearing, hierarchically organized tissue composed of a network of type II collagen embedded in an aggrecan-rich polyelectrolyte gel. Its ability to resist deformation and dissipate energy arises from spatially varying matrix composition and architecture. Here, we review experimental and theoretical advances that elucidate the mechanistic basis of cartilage shear mechanics. Recent studies have shown that the tissue operates near a rigidity transition, in which small changes in collagen density, cross-linking, or osmotic stress can produce large, nonlinear changes in shear stiffness. We discuss how this behavior is captured by models rooted in rigidity percolation, continuum elasticity, and micromechanics, and how these frameworks connect depth-dependent composition to macroscale mechanical response. Throughout, we emphasize physical principles that describe observations across native, degraded, and engineered tissues, and we highlight emerging strategies for designing cartilage-inspired materials with tunable, anisotropic mechanics, with applications in soft robotics, synthetic gels, and load-bearing biomaterials. 

Abstract Intra‐articular injections of hyaluronic acid (HA) are the cornerstone of osteoarthritis (OA) treatments. However, the mechanism of action and efficacy of HA viscosupplementation are debated. As such, there has been recent interest in developing synthetic viscosupplements. Recently, a synthetic 4 wt% polyacrylamide (pAAm) hydrogel was shown to effectively lubricate and bind to the surface of cartilage in vitro. However, its ability to localize to cartilage and alter the tribological properties of the tissue in a live articulating large animal joint is not known. The goal of this study was to quantify the distribution and extent of localization of pAAm in the equine metacarpophalangeal or metatarsophalangeal joint (fetlock joint), and determine whether preferential localization of pAAm influences the tribological properties of the tissue. An established planar fluorescence imaging technique was used to visualize and quantify the distribution of fluorescently labeled pAAm within the joint. While the pAAm hydrogel was present on all surfaces, it was not uniformly distributed, with more material present near the site of the injection. The lubricating ability of the cartilage in the joint was then assessed using a custom tribometer across two orders of magnitude of sliding speed in healthy synovial fluid. Cartilage regions with a greater coverage of pAAm, that is, higher fluorescent intensities, exhibited friction coefficients nearly 2‐fold lower than regions with lesser pAAm (R_rm = −0.59,p < 0.001). Collectively, the findings from this study indicate that intra‐articular viscosupplement injections are not evenly distributed inside a joint, and the tribological outcomes of these materials is strongly determined by the ability of the material to localize to the articulating surfaces in the joint.