Mechanical compression influences intracellular Ca²⁺signaling in chondrocytes seeded in agarose constructs

Abstract

Ca2+ signaling forms part of a possible mechanotransduction pathway by which chondrocytes may alter their metabolism in response to mechanical loading. In this study, a well-characterized model system utilizing bovine articular chondrocytes embedded in 4% agarose constructs was used to investigate the effect of physiological mechanical compressive strain applied after 1 and 3 days in culture. The intracellular Ca2+ concentration was measured by use of the ratiometric Ca2+ indicator indo 1-AM and confocal microscopy. A positive Ca2+ response was defined as a percent increase in Ca2+ ratio above a preset threshold. A significantly greater percentage of cells exhibited a positive Ca2+ response in strained constructs compared with unstrained controls at both time points. In strained constructs, treatment with either Ga3+ or EGTA significantly reduced the number of positive Ca2+ responders compared with untreated controls. These results represent an important step in understanding the physiological role of intracellular Ca2+ in chondrocytes under mechanical compression.

Mechanically induced calcium signaling in chondrocytes in situ

“…These response times are much faster than those reported for in vitro experiments using agarose constructs. For example, Roberts et al 16 found a delay in calcium signaling of approximately 200 s after applying a static, nominal compression of 20% to a chondrocyte/agarose construct. They suggested that calcium signaling is a downstream event in the mechanotransduction cascade.…”

“…[10][11][12][13] Calcium is an ubiquitous second messenger which regulates many cellular processes including metabolism, proliferation, gene transcription, contraction, and mechanotransduction. 14 Previous studies have suggested that Ca 2þ signals form part of an intricate mechanotransduction pathway in chondrocytes: physical stimuli, such as compression, [15][16][17][18][19] fluid flow, [20][21][22] hydrostatic pressure, 23,24 and osmotic stress, [25][26][27][28][29] influence Ca 2þ signaling in chondrocytes. Therefore, insight into mechanically induced Ca 2þ signaling may provide novel understanding of chondrocyte mechanotransduction.…”

“…16,18,30 However, these studies neglect the potential role of signaling pathways involving transmembrane proteins linked to the ECM, as well as the specialized peri-cellular matrices that affect chondrocyte mechanics, 31,32 and thus, the biological responses of chondrocytes to mechanical loading conditions. The aim of this study was to develop a method to investigate biological responses of chondrocytes in intact articular cartilage attached to its native bone in response to mechanical perturbations.…”

Computational investigation of in situ chondrocyte deformation and actin cytoskeleton remodelling under physiological loading

“…The first phase simulates the formation of an equilibrium actin cytoskeleton distribution in the cell in the absence of externally applied loading. As previously mentioned, an exponentially decaying signal in the cytoplasm (as observed for chondrocytes [50]) drives the formation of the actin cytoskeleton, while tension reduction in the cytoplasm leads to dissociation. The second phase then involves the application of nominal strains to the RVE.…”

Cellular contractility and substrate elasticity: a numerical investigation of the actin cytoskeleton and cell adhesion

“…Cellular signalling drives SF remodelling and the exponentially decaying signal in the current study is based on the observations of Roberts et al (2001) and Ruwhof et al (2001). In these studies, an approximately exponentially decaying calcium signal is detected following mechanical loading.…”