Diarthrodial joints are essential for load bearing and locomotion. Physiologically, articular cartilage sustains millions of cycles of mechanical loading. Chondrocytes, the cells in cartilage, regulate their metabolic activities in response to mechanical loading. Pathological mechanical stress can lead to maladaptive cellular responses and subsequent cartilage degeneration. We sought to deconstruct chondrocyte mechanotransduction by identifying mechanosensitive ion channels functioning at injurious levels of strain. We detected robust expression of the recently identified mechanosensitive channels, PIEZO1 and PIEZO2. Combined directed expression of Piezo1 and -2 sustained potentiated mechanically induced Ca 2+ signals and electrical currents compared with single-Piezo expression. In primary articular chondrocytes, mechanically evoked Ca 2+ transients produced by atomic force microscopy were inhibited by GsMTx4, a PIEZO-blocking peptide, and by Piezo1-or Piezo2-specific siRNA. We complemented the cellular approach with an explant-cartilage injury model. GsMTx4 reduced chondrocyte death after mechanical injury, suggesting a possible therapy for reducing cartilage injury and posttraumatic osteoarthritis by attenuating Piezo-mediated cartilage mechanotransduction of injurious strains.A rticular cartilage is a hydrated connective tissue that supports loads and minimizes friction in the diarthrodial joints. It has a highly differentiated extracellular matrix (ECM) composed primarily of type II collagen, the large aggregating proteoglycan, aggrecan, and water. Chondrocytes are the only cells in cartilage and are responsible for maintaining and remodeling cartilage through a homeostatic balance of anabolic and catabolic activities. Under normal physiologic conditions, chondrocytes are exposed to millions of cycles of mechanical loading per year (1). These mechanical signals play an important role in regulating chondrocyte anabolic and biosynthetic activity, as evidenced by cartilage atrophy following periods of disuse or immobilization (2-7). However, under abnormal loading conditions (e.g., due to obesity, trauma, or joint instability), mechanical factors play a critical role in the onset and progression of osteoarthritis (1). Such "injurious" loading has been modeled in vitro using explant culture systems that replicate many of the early cellular and molecular events characteristic of osteoarthritis (8). Osteoarthritis is a painful and debilitating disease of weight-bearing joints that affects over 26 million people in the United States (9) with posttraumatic arthritis being responsible for ∼12% of the incidence of osteoarthritis (10).Despite the critical importance of mechanical loading in health and disease of synovial joints, the mechanisms of mechanotransduction of chondrocytes are not fully understood and are likely to differ under physiologic and pathologic conditions (11)(12)(13)(14). Although many different mechanisms have been shown to be involved in chondrocyte mechanotransduction (13,(15)(16)(17), recent st...

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ObjectiveTo assess the role of the glycoprotein PRG4 in joint lubrication and chondroprotection by measuring friction, stiffness, surface topography, and subsurface histology of the hip joints of Prg4−/− and wild-type (WT) mice.MethodsFriction and elastic modulus were measured in cartilage from the femoral heads of Prg4−/− and WT mice ages 2, 4, 10, and 16 weeks using atomic force microscopy, and the surface microstructure was imaged. Histologic sections of each femoral head were stained and graded.ResultsHistologic analysis of the joints of Prg4−/− mice showed an enlarged, fragmented surface layer of variable thickness with Safranin O–positive formations sometimes present, a roughened underlying articular cartilage surface, and a progressive loss of pericellular proteoglycans. Friction was significantly higher on cartilage of Prg4−/− mice at age 16 weeks, but statistically significant differences in friction were not detected at younger ages. The elastic modulus of the cartilage was similar between cartilage surfaces of Prg4−/− and WT mice at young ages, but cartilage of WT mice showed increasing stiffness with age, with significantly higher moduli than cartilage of Prg4−/− mice at older ages.ConclusionDeletion of the gene Prg4 results in significant structural and biomechanical changes in the articular cartilage with age, some of which are consistent with osteoarthritic degeneration. These findings suggest that PRG4 plays a significant role in preserving normal joint structure and function.

Lubricin and hyaluronic acid (HA), molecular constituents of synovial fluid, have long been theorized to play a role in joint lubrication and wear protection. While lubricin has been shown to function as a boundary lubricant, conflicting evidence exists as to the boundary lubricating ability of hyaluronic acid. Here, we use colloidal force microscopy to explore the friction behavior of these two molecules on the microscale between chemically uniform hydrophilic (hydroxyl-terminated) and hydrophobic (methyl-terminated) surfaces in physiological buffer solution. Behaviors on both surfaces are physiologically relevant since the heterogeneous articular cartilage surface contains both hydrophilic and hydrophobic elements. Friction between hydrophobic surfaces was initially high (μ=1.1, at 100nN of applied normal load) and was significantly reduced by lubricin addition while friction between hydrophilic surfaces was initially low (μ=0.1) and was slightly increased by lubricin addition. At lubricin concentrations above 200 µg/ml, friction behavior on the two surfaces was similar (μ=0.2) indicating that nearly all interaction between the two surfaces was between adsorbed lubricin molecules rather than between the surfaces themselves. In contrast, addition of HA did not appreciably alter the frictional behavior between the model surfaces. No synergistic effect on friction behavior was seen in a physiological mixture of lubricin and HA. Lubricin can equally mediate the frictional response between both hydrophilic and hydrophobic surfaces, likely fully preventing direct surface-to-surface contact at sufficient concentrations, whereas HA provides considerably less boundary lubrication.

Articular cartilage provides a low-friction, wear-resistant surface for the motion of diarthrodial joints. The objective of this study was to develop a method for in situ friction measurement of murine cartilage using a colloidal probe attached to the cantilever of an atomic force microscope. Sliding friction was measured between a chemically functionalized microsphere and the cartilage of the murine femoral head. Friction was measured at normal loads ranging incrementally from 20 to 100 nN with a sliding speed of 40 microm/s and sliding distance of 64 microm. Under these test conditions, hydrostatic pressurization and biphasic load support in the cartilage were minimized, providing frictional measurements that predominantly reflect boundary lubrication properties. Friction coefficients measured on murine tissue (0.25+/-0.11) were similar to those measured on porcine tissue (0.23+/-0.09) and were in general agreement with measurements of boundary friction on cartilage by other researchers. Using the colloidal probe as an indenter, the elastic mechanical properties and surface roughness were measured in the same configuration. Interfacial shear was found to be the principal mechanism of friction generation, with little to no friction resulting from plowing forces, collision forces, or energy losses due to normal deformation. This measurement technique can be applied to future studies of cartilage friction and mechanical properties on genetically altered mice or other small animals.